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Background: Anti-calcitonin gene-related peptide (CGRP) monoclonal antibodies have emerged as promising therapeutic options for the treatment of chronic migraine. However, treatment response varies considerably among individuals, suggesting a potential role for genetic factors. This study aimed to identify genetic variants affecting the efficacy of anti-CGRP monoclonal antibody therapy in chronic migraine among the Han Chinese population in Taiwan to enhance treatment precision and to understand the genetic architecture of migraine. Methods: We conducted a quantitative trait locus (QTL) association study in patients with chronic migraines from a tertiary medical center in Taiwan using the Taiwan Precision Medicine Array Chip. The patients received fremanezumab or galcanezumab for at least 12 weeks. Treatment efficacy was assessed based on the improvement rate in monthly migraine days. Genetic variants were identified, and their associations with treatment efficacy were examined through quantitative trait loci analysis, linkage disequilibrium studies, and functional annotations using the Gene Ontology database. Results: Six single nucleotide polymorphisms (SNPs) relative variants were significantly associated with anti-CGRP therapy response (p < 1 × 10− 7): rs116870564, rs75244870, rs56216870, rs12938101, rs74655790, and rs149540851. These variants are located in or near genes, including LRRC4C, ATAD2B, and OXR1, which are involved in neuronal development, DNA-dependent ATPase activity, and oxidation-reduction processes, respectively. The rs116870564 variant in LRRC4C showed the strongest association (β = -0.551, p = 6.65 × 10− 9). The functional impact of these variants is attributed to their regulatory effects on gene expression, which are influenced by intron splicing regulation, transcription factors, and changes in chromatin structure. Conclusion: The identification of key genetic markers associated with response to anti-CGRP therapy emphasizes the importance of genetic variability in treatment efficacy. This could lead to more personalized chronic migraine management strategies and tailored therapeutic approaches based on individual genetic profiles. Further research in larger, diverse populations is warranted to validate these findings and refine our understanding of the role of CGRP in chronic migraine pathophysiology. Trial registration: Not applicable. [ABSTRACT FROM AUTHOR]

Objective: Recurrent pregnancy loss (RPL) has a multifactorial etiology, with a majority of cases remaining unexplained. To account for these unexplained cases, possible male factors are being explored. Conventional semen analysis lacks a qualitative assessment of sperms and information regarding sperm DNA integrity. Sperm DNA fragmentation (SDF) has diagnostic value in unexplained RPL, and it may account for a number of unexplained cases. Hence, we planned a study to explore and evaluate the impact of sperm DNA fragmentation in couples with unexplained recurrent pregnancy losses. Study Design: Analytical cross‐sectional study was conducted at a tertiary‐level referral facility in India between August 2021 and July 2023. Participants (n = 70) were divided into two groups—male partners of couples with unexplained RPL (following spontaneous conceptions) (n = 35) and men with at least one previous live birth (spontaneous or following fertility treatments for female factor infertility such as ovulation induction or intrauterine insemination) as controls (n = 35). Neither of the two groups of couples recruited for this study had undergone ART as fertility treatment. Primary outcome assessed was mean DNA fragmentation index (DFI). Secondary outcomes included differences in semen parameters such as sperm concentration, progressive sperm motility and morphology, proportion of men with high (≥30%) and low DFI in the two groups, and the association between various semen parameters and DFI. Results: Univariate logistic regression revealed that sperm DNA fragmentation was higher in men with unexplained RPL (30.0; IQR (interquartile range) 19.0, 46.0) as compared to controls (22.0; IQR 14.0, 30.0) although it was not statistically significant (OR, odds ratio, 1.02; 95% CI 1.0–1.1, p = 0.08). A higher proportion of men with unexplained RPL had DFI ≥30% compared to controls (54.2% vs. 25.7%; OR 3.43 (95% CI 1.2–9.4); p = 0.02). No statistically significant differences were observed in semen volume, sperm concentration, progressive motility, and morphology between the two groups. Sperm DNA fragmentation index also showed a weak but significant inverse relationship with sperm morphology (r = −0.336, p = 0.004). Conclusion: The current study did not show any significant difference in the mean sperm DNA fragmentation levels in male partners of couples with unexplained RPL compared to controls. However, a higher proportion of men with DFI ≥30% were observed in unexplained RPL population when compared to controls. [ABSTRACT FROM AUTHOR]

Mutating replication-dependent (RD) histone genes is an important tool for understanding chromatin-based epigenetic regulation. Deploying this tool in metazoans is particularly challenging because RD histones in these organisms are typically encoded by many genes, often located at multiple loci. Such gene arrangements make the ability to generate homogenous histone mutant genotypes by site-specific gene editing quite difficult. Drosophila melanogaster provides a solution to this problem because the RD histone genes are organized into a single large tandem array that can be deleted and replaced with transgenes containing mutant histone genes. In the last ∼15 years several different RD histone gene replacement platforms were developed using this simple strategy. However, each platform contains weaknesses that preclude full use of the powerful developmental genetic capabilities available to Drosophila researchers. Here we describe the development of a newly engineered platform that rectifies many of these weaknesses. We used CRISPR to precisely delete the RD histone gene array (HisC), replacing it with a multifunctional cassette that permits site-specific insertion of either one or two synthetic gene arrays using selectable markers. We designed this cassette with the ability to selectively delete each of the integrated gene arrays in specific tissues using site-specific recombinases. We also present a method for rapidly synthesizing histone gene arrays of any genotype using Golden Gate cloning technologies. These improvements facilitate the generation of histone mutant cells in various tissues at different stages of Drosophila development and provide an opportunity to apply forward genetic strategies to interrogate chromatin structure and gene regulation. [ABSTRACT FROM AUTHOR]

The present study aimed to evaluate whether epigenetic markers are expressed in the dental follicles surrounding ectopically erupting teeth. Twenty‐one dental follicles were collected in 20 adolescent children through surgical exposure of ectopic teeth. The epigenetic modifications of DNA methylation and histone acetylation were evaluated by immunohistochemistry. The results showed cells positive for DNA‐methyltransferase 1 (DNMT1), DNA methyltransferase 3 beta (DNMT3B), ten‐eleven translocation‐2 (TET2), acetyl‐histone H3 (AcH3), acetyl‐histone H4 (AcH4), 5‐methylcytosine (5mC), and 5‐hydroxymethylcytosine (5hmC) were present in all the samples. The levels of epigenetic markers representing active chromatin (5hmC, AcH3, AcH4, and TET2) were statistically significantly higher than those of markers representing inactive chromatin (5mC, DNMT3B, DNMT1). In conclusion, follicles in ectopic teeth display major epigenetic modifications. In the follicles, epigenetic markers associated with the activation of bone‐related genes are more abundant than markers associated with the inactivation of bone‐related genes. [ABSTRACT FROM AUTHOR]

Organisms with differentiated sex chromosomes must accommodate unequal gene dosage in males and females. Male fruit flies increase X-linked gene expression to compensate for hemizygosity of their single X chromosome. Full compensation requires localization of the Male-Specific Lethal (MSL) complex to active genes on the male X, where it modulates chromatin to elevate expression. The mechanisms that identify X chromatin are poorly understood. The euchromatic X is enriched for AT-rich, ∼359 bp satellites termed the 1.688X repeats. Autosomal insertions of 1.688X DNA enable MSL recruitment to nearby genes. Ectopic expression of dsRNA from one of these repeats produces siRNA and partially restores X-localization of MSLs in males with defective X recognition. Surprisingly, expression of double-stranded RNA from three other 1.688X repeats failed to rescue males. We reconstructed dsRNA-expressing transgenes with sequence from two of these repeats and identified phasing of repeat DNA, rather than sequence or orientation, as the factor that determines rescue of males with defective X recognition. Small RNA sequencing revealed that siRNA was produced in flies with a transgene that rescues, but not in those carrying a transgene with the same repeat but different phasing. We demonstrate that pericentromeric X heterochromatin promotes X recognition through a maternal effect, potentially mediated by small RNA from closely related heterochromatic repeats. This suggests that the sources of siRNAs promoting X recognition are highly redundant. We propose that enrichment of satellite repeats on Drosophilid X chromosomes facilitates the rapid evolution of differentiated sex chromosomes by marking the X for compensation. [ABSTRACT FROM AUTHOR]

Background: The dilation and torsion of testicular veins in the plexus pampiniformis causes Varicocele, which is a surgically repairable cause of male infertility. This study assessed the impact of varicocelectomy on semen characteristics, total motile sperm count (TMSC) and sperm DNA integrity in patients with severe oligoasthenoteratozoospermia (OAT). Materials and Methods: In this prospective study, semen samples of 360 men with severe OAT who underwent varicocelectomy according to World Health Organization (WHO) criteria 2021 were studied (pre-operatively and at 6, 12, and 18 months post-operatively). Results: The average age of our patients was 38.5 years. The mean spermatozoa concentration was found to be 1.60 ± 0.83 million/ml pre-operatively, while the mean post-operative concentration was 5.17 ± 1.23 million/ml at 6 months, 8.32 ± 0.98 million/ml at 12 months, and 13.51 ± 1.48 million/ml at 18 months (P<0.0001). The mean percentage of A+B motile spermatozoa was 2.92 ± 1.17% pre-operatively, 6.10 ± 1.51% at six months, 9.58 ± 1.49% at 12 months and 13.92 ± 1.88% at 18 months postoperatively (P<0.0001). The mean Modified David's morphology score was 3.80 ± 1.43% pre-operatively, 5.95 ± 1.23% at 6 months, 7.94 ± 1.18% at 12 months, and 10.82 ± 1.91% at 18 months post-operatively (P<0.0001). The mean of total motile sperm count (TMSC) was statistically improved after varicocelectomy (P<0.001). The mean of DNA fragmentation index (DFI) of the spermatozoa was 31.40 ± 0.52% pre-operatively, and post-operatively at 28.20 ± 0.32% at 6 months, 25.90 ± 0.31% at 12 months and 20.50 ± 0.40% at 18 months (P<0.001). Conclusion: Varicocelectomy was associated with significant improvement of sperm parameters and DNA fragmentation resulting in significant improvement of spermatogenesis quality. We believe that universalization in the routinely used sperm dispersion chromatin (SDC) test could be beneficial in the treatment of infertility. [ABSTRACT FROM AUTHOR]

Simple Summary: Alternative splicing is a mechanism that allows, through the combination of different exons, for the yielding of several protein variants from a single gene. These variants may display different and often opposed functions. Mutations occurring in driver genes result in oncogenesis, but alternative splicing may also result in obtaining oncogenic variants in the absence of driver mutations. This review describes how the oncogenic potential of driver genes is activated through aberrant alternative splicing. Firstly, there are driver genes directly acting on alternative splicing and their dysregulation affects the splicing of other genes involved in malignant transformation. A second possibility is that aberrant alternative splicing of a proto-oncogene or tumour suppressor gene results in the appearance of an oncogenic variant. The oncogenic potential of a total of 199 driver genes may result from aberrant alternative splicing, and the molecular mechanisms involved are detailed for more than 40 genes. Cancer driver genes are either oncogenes or tumour suppressor genes that are classically activated or inactivated, respectively, by driver mutations. Alternative splicing—which produces various mature mRNAs and, eventually, protein variants from a single gene—may also result in driving neoplastic transformation because of the different and often opposed functions of the variants of driver genes. The present review analyses the different alternative splicing events that result in driving neoplastic transformation, with an emphasis on their molecular mechanisms. To do this, we collected a list of 568 gene drivers of cancer and revised the literature to select those involved in the alternative splicing of other genes as well as those in which its pre-mRNA is subject to alternative splicing, with the result, in both cases, of producing an oncogenic isoform. Thirty-one genes fall into the first category, which includes splicing factors and components of the spliceosome and splicing regulators. In the second category, namely that comprising driver genes in which alternative splicing produces the oncogenic isoform, 168 genes were found. Then, we grouped them according to the molecular mechanisms responsible for alternative splicing yielding oncogenic isoforms, namely, mutations in cis splicing-determining elements, other causes involving non-mutated cis elements, changes in splicing factors, and epigenetic and chromatin-related changes. The data given in the present review substantiate the idea that aberrant splicing may regulate the activation of proto-oncogenes or inactivation of tumour suppressor genes and details on the mechanisms involved are given for more than 40 driver genes. [ABSTRACT FROM AUTHOR]

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Simple Summary: Bortezomib-containing chemotherapy did not improve the clinical outcome in the AAML1031 study in terms of overall survival and event-free survival compared to standard chemotherapy. We characterized epigenetically distinct proteomic profiles in a large cohort of pediatric patients that participated in this study using the reverse-phase protein array. We observed in the patient group that received standard therapy that a higher expression of 16 histone-modulating enzymes (HMEs) was an independent variable that predicted higher relapse risk three years after a second induction therapy compared to those with a lower HME protein expression. Also, there was significantly improved overall survival for those with a high HME expression who were treated with the bortezomib-containing chemotherapy, compared to high-HME patients treated without bortezomib. We also demonstrated that patients with a higher expression of HME had more open chromatin surrounding promoter sides compared to those with lower HME protein levels using ATAC-seq. The addition of the proteasome inhibitor bortezomib to standard chemotherapy did not improve survival in pediatric acute myeloid leukemia (AML) when all patients were analyzed as a group in the Children's Oncology Group phase 3 trial AAML1031 (NCT01371981). Proteasome inhibition influences the chromatin landscape and proteostasis, and we hypothesized that baseline proteomic analysis of histone- and chromatin-modifying enzymes (HMEs) would identify AML subgroups that benefitted from bortezomib addition. A proteomic profile of 483 patients treated with AAML1031 chemotherapy was generated using a reverse-phase protein array. A relatively high expression of 16 HME was associated with lower EFS and higher 3-year relapse risk after AML standard treatment compared to low expressions (52% vs. 29%, p = 0.005). The high-HME profile correlated with more transposase-accessible chromatin, as demonstrated via ATAC-sequencing, and the bortezomib addition improved the 3-year overall survival compared with standard therapy (62% vs. 75%, p = 0.033). These data suggest that there are pediatric AML populations that respond well to bortezomib-containing chemotherapy. [ABSTRACT FROM AUTHOR]

Chromosome condensation is essential for the fidelity of chromosome segregation during mitosis and meiosis. Condensation is associated both with local changes in nucleosome structure and larger-scale alterations in chromosome topology mediated by the condensin complex. We examined the influence of linker histone H1 and variant histone H2A.Z on chromosome condensation in budding yeast cells. Linker histone H1 has been implicated in local and global compaction of chromatin in multiple eukaryotes, but we observe normal condensation of the rDNA locus in yeast strains lacking H1. However, deletion of the yeast HTZ1 gene, coding for variant histone H2A.Z , causes a significant defect in rDNA condensation. Loss of H2A.Z does not change condensin association with the rDNA locus or significantly affect condensin mRNA levels. Prior studies reported that several phenotypes caused by loss of H2A.Z are suppressed by eliminating Swr1 , a key component of the SWR complex that deposits H2A.Z in chromatin. We observe that an htz1Δ swr1Δ strain has near-normal rDNA condensation. Unexpectedly, we find that elimination of the linker histone H1 can also suppress the rDNA condensation defect of htz1Δ strains. Our experiments demonstrate that histone H2A.Z promotes chromosome condensation, in part by counteracting activities of histone H1 and the SWR complex. [ABSTRACT FROM AUTHOR]

Background: Sex determination is essential in forensic science, anthropology, and medical diagnostics. Traditional methods, such as DNA analysis, are resource intensive and time-consuming. This pilot study explores the utility of oral rinse-based cytology for detecting Barr bodies in exfoliated oral mucosal cells from saliva as a more accessible and efficient alternative. Aim: This study aimed to assess the viability of rinse-based cytology in identifying sex through the detection of Barr bodies. Materials and Methods: In the study, 60 participants aged 20-30, free of lesions, and nonsmokers/nondrinkers, were evenly divided into male and female groups. Unstimulated saliva was collected from each participant between 9 and 10 am Participants were instructed to tilt their heads slightly forward to accumulate saliva and then spit into disposable sterile containers. After centrifugation, two smears were prepared and stained with Papanicolaou (PAP) and hematoxylin and eosin (H and E) stains. The smears were labeled (single-blinded) and examined for the presence of Barr body by two oral pathologists. Results: H and E stain had a mean Barr body count of 2.38 and a standard deviation of approximately 2.731, whereas PAP stain had a higher mean count of 3.57 with a standard deviation of 3.933. Levene's test showed significant differences (P = 0.000) in variances between the staining methods (F = 21.001, df = 118, P < 0.001), but the average Barr body count difference of about 1.183 with P = 0.058 was not statistically significant (P > 0.05). Conclusion: Oral rinse-based cytology offers a noninvasive, cost-effective, and time-efficient approach to sex determination. While this pilot study demonstrates its potential, further research involving larger and more diverse populations is required to validate its reliability fully. This innovative approach has the potential to impact various scientific and forensic disciplines, offering a practical alternative to traditional methods. [ABSTRACT FROM AUTHOR]

Objective: The diminished expression of microRNA‐146a (miR‐146a) in systemic lupus erythematosus (SLE) contributes to the aberrant activation of the interferon pathway. Despite its significance, the underlying mechanism driving this reduced expression remains elusive. Considering the integral role of enhancers in steering gene expression, our study seeks to pinpoint the SLE‐affected enhancers responsible for modulating miR‐146a expression. Additionally, we aim to elucidate the mechanisms by which these enhancers influence the contribution of miR‐146a to the activation of the interferon pathway. Methods: Circular chromosome conformation capture sequencing and epigenomic profiles were applied to identify candidate enhancers of miR‐146a. CRISPR activation was performed to screen functional enhancers. Differential analysis of chromatin accessibility was used to identify SLE‐dysregulated enhancers, and the mechanism underlying enhancer dysfunction was investigated by analyzing transcription factor binding. The therapeutic value of a lupus‐related enhancer was further evaluated by targeting it in the peripheral blood mononuclear cells (PBMCs) of patients with SLE through a CRISPR activation approach. Results: We identified shared and cell‐specific enhancers of miR‐146a in distinct immune cells. An enhancer 32.5 kb downstream of miR‐146a possesses less accessibility in SLE, and its chromatin openness was negatively correlated with SLE disease activity. Moreover, CCAAT/enhancer binding protein α, a down‐regulated transcription factor in patients with SLE, binds to the 32.5‐kb enhancer and induces the epigenomic change of this locus. Furthermore, CRISPR‐based activation of this enhancer in SLE PBMCs could inhibit the activity of interferon pathway. Conclusion: Our work defines a promising target for SLE intervention. We adopted integrative approaches to define cell‐specific and functional enhancers of the SLE critical gene and investigated the mechanism underlying its dysregulation mediated by a lupus‐related enhancer. [ABSTRACT FROM AUTHOR]

The N 6-methyladenosine (m6A) modification is a highly conserved RNA modification found in eukaryotic messenger RNAs (mRNAs). It plays a vital role in regulating various biological processes. Dysregulation of m6A modifications has been linked to a range of complex genetic diseases in humans. However, there has been a lack of comprehensive characterization and comparison of m6A modifications at the transcriptome-wide level within families. To address this gap, we profiled transcriptome-wide m6A methylation in 18 individuals across 6 Yoruba trio families. The m6A methylomes of these 18 individuals revealed that m6A modifications in children showed greater similarity to each other than to their parents. This suggests that m6A modifications are influenced by multiple factors rather than solely determined by genetic factors. Additionally, we found that mRNAs exhibiting m6A modifications specific to children were enriched in cell cycle control processes, while those with m6A modifications specific to parents were associated with chromatin modifications. Furthermore, our analysis on the interactions between differentially expressed m6A-related regulatory genes and age-related genes suggested that age might be one of the factors influencing m6A modifications. In summary, our study provided a valuable dataset that highlighted the differences and functional diversity of m6A modifications within and between trio families. [ABSTRACT FROM AUTHOR]

Chromatin remodeling is central to the dynamic changes in gene expression that drive cell fate determination. During development, the sets of enhancers that are accessible for use change globally as cells transition between stages. While transcription factors and nucleosome remodelers are known to work together to control enhancer accessibility, it is unclear how the short stretches of DNA that they individually unmask yield the kilobase-sized accessible regions characteristic of active enhancers. Here, we performed a genetic screen to investigate the role of nucleosome remodelers in control of dynamic enhancer activity. We find that the Drosophila Switch/Sucrose Non-Fermenting complex, BAP, is required for repression of a temporally dynamic enhancer, brdisc. Contrary to expectations, we find that the BAP-specific subunit Osa is dispensable for mediating changes in chromatin accessibility between the early and late stages of wing development. Instead, we find that Osa is required to constrain the levels of brdisc activity when the enhancer is normally active. Genome-wide profiling reveals that Osa directly binds brdisc as well as thousands of other developmentally dynamic regulatory sites, including multiple genes encoding components and targets of the Notch signaling pathway. Transgenic reporter analyses demonstrate that Osa is required for activation and for constraint of different sets of target enhancers in the same cells. Moreover, Osa loss results in hyperactivation of the Notch ligand Delta and development of ectopic sensory structures patterned by Notch signaling early in development. Together, these findings indicate that proper constraint of enhancer activity is necessary for regulation of dose-dependent developmental events. [ABSTRACT FROM AUTHOR]

The effect of in-vitro sperm incubation with Pentoxifylline (PTX) and Coenzyme Q10 (CoQ10) in Oligoasthenoteratozoospermia (OAT) patients was evaluated. Semen samples were obtained from men with Normozoospermia and men with OAT. Motile sperm from the two groups were subdivided into four subgroups: (i) without incubation with PTX + CoQ10; (ii) incubation with PTX; (iii) Incubation with CoQ10; and (iv) incubation with a combination of PTX + CoQ10. Then, sperm parameters, chromatin, DNA and membrane integrity, protamine deficiency, apoptosis, mitochondrial activity, sperm chromatin dispersion test (SCD), hypo-osmotic swelling test (HOS), chromomycin A3 (CMA3), Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL), and diaminobenzidine (DAB) assays were evaluated, respectively. Sperm incubated with CoQ10 and a combination of CoQ10 and PTX resulted in a significant increase in the sperm parameters. Also, a significant decrease was noted with a combination of PTX and CoQ10 in normal men. There was a significant difference between CoQ10 treated and CoQ10 + PTX treated groups in comparison with the OAT group in the percentage of the DNA fragmentation, sperm apoptosis, AB+, HOS test + and sperm mitochondrial activity. Incubated sperm with CoQ10, PTX, and in combination with each other can improve sperm parameters in OAT patients. [ABSTRACT FROM AUTHOR]

The fruit fly Drosophila melanogaster serves as a powerful model organism for advancing our understanding of biological processes, not just by studying its similarities with other organisms including ourselves but also by investigating its differences to unravel the underlying strategies that evolved to achieve a common goal. This is particularly true for centromeres, specialized genomic regions present on all eukaryotic chromosomes that function as the platform for the assembly of kinetochores. These multiprotein structures play an essential role during cell division by connecting chromosomes to spindle microtubules in mitosis and meiosis to mediate accurate chromosome segregation. Here, we will take a historical perspective on the study of fly centromeres, aiming to highlight not only the important similarities but also the differences identified that contributed to advancing centromere biology. We will discuss the current knowledge on the sequence and chromatin organization of fly centromeres together with advances for identification of centromeric proteins. Then, we will describe both the factors and processes involved in centromere organization and how they work together to provide an epigenetic identity to the centromeric locus. Lastly, we will take an evolutionary point of view of centromeres and briefly discuss current views on centromere drive. [ABSTRACT FROM AUTHOR]

Large genome structural variations can impact genome regulation and integrity. Repeat-rich regions like pericentric heterochromatin are vulnerable to structural rearrangements although we know little about how often these rearrangements occur over evolutionary time. Repetitive genome regions are particularly difficult to study with genomic approaches, as they are missing from most genome assemblies. However, cytogenetic approaches offer a direct way to detect large rearrangements involving pericentric heterochromatin. Here, we use a cytogenetic approach to reveal large structural rearrangements associated with the X pericentromeric region of Drosophila simulans. These rearrangements involve large blocks of satellite DNA—the 500-bp and Rsp-like satellites—which colocalize in the X pericentromeric heterochromatin. We find that this region is polymorphic not only among different strains, but between isolates of the same strain from different labs, and even within individual isolates. On the one hand, our observations raise questions regarding the potential impact of such variation at the phenotypic level and our ability to control for such genetic variability. On the other hand, this highlights the very rapid turnover of the pericentric heterochromatin most likely associated with genomic instability of the X pericentromere. It represents a unique opportunity to study the dynamics of pericentric heterochromatin, the evolution of associated satellites on a very short time scale, and to better understand how structural variation arises. [ABSTRACT FROM AUTHOR]

Background: Sperm chromatin abnormalities are defects in nuclear maturation and DNA integrity. These defects originated from defective spermatogenesis due to a lack of DNA repair during chromatin remodeling. Changes in semen elements can cause damage to chromatin. There is little information about the relationship between changes in trace metal elements and total antioxidant capacity (TAC) with sperm chromatin damage. The present study was conducted to determine the relationship between Selenium (Se), Iron (Fe), Zinc (Zn), Copper (Cu) and the TAC of semen with the status of human sperm chromatin. Materials and Methods: In this experimental study, semen samples (n=30) were collected from healthy men referred to Kermanshah Motazadi Hospital and stored in liquid nitrogen; after thawing and centrifugation, sperm were separated. The atomic absorption method was used to measure the concentration of metal elements. The TAC was evaluated using the ferric-reducing antioxidant capacity of the plasma method. Furthermore, the integrity of sperm chromatin was measured using the sperm DNA fragmentation (SDF) method. Results: The status of sperm chromatin had a non-significant correlation with body mass index (BMI, P=0.25, r=0.21) and a non-significant negative correlation with sperm count (P=0.71, r=-0.71) and motility (P=0.75, r=0.61). In addition, there was no significant relationship between sperm chromatin and the TAC of semen (P=0.92, r=0.01). Additionally, there was no significant correlation between Se, Zn, or Cu concentration (P>0.05) and Fe concentration, which had a partially positive relationship with the chromatin state of sperm (P=0.24, r=0.20). Conclusion: The trace metal elements in the seminal fluid did not play a significant role in the status of sperm chromatin. [ABSTRACT FROM AUTHOR]

The article discusses the results of a study on the use of H3K27me3 immunohistochemistry in evaluating the dotlike expression pattern for female origin in testicular germ cell tumors. Topics mentioned include the indication of the silenced X chromosome in normal and neoplastic female tissues, the presence of dotlike expression in epithelial and stromal/mesenchymal teratoma compartments and the presence of epigenetic mutation causing the inactivation of X chromosome.

The chromatin of animal cells contains two types of histones: canonical histones that are expressed during S phase of the cell cycle to package the newly replicated genome, and variant histones with specialized functions that are expressed throughout the cell cycle and in nonproliferating cells. Determining whether and how canonical and variant histones cooperate to regulate genome function is integral to understanding how chromatin-based processes affect normal and pathological development. Here, we demonstrate that variant histone H3.3 is essential for Drosophila development only when canonical histone gene copy number is reduced, suggesting that coordination between canonical H3.2 and variant H3.3 expression is necessary to provide sufficient H3 protein for normal genome function. To identify genes that depend upon, or are involved in, this coordinate regulation we screened for heterozygous chromosome 3 deficiencies that impair development of flies bearing reduced H3.2 and H3.3 gene copy number. We identified two regions of chromosome 3 that conferred this phenotype, one of which contains the Polycomb gene, which is necessary for establishing domains of facultative chromatin that repress master regulator genes during development. We further found that reduction in Polycomb dosage decreases viability of animals with no H3.3 gene copies. Moreover, heterozygous Polycomb mutations result in de-repression of the Polycomb target gene Ubx and cause ectopic sex combs when either canonical or variant H3 gene copy number is reduced. We conclude that Polycomb-mediated facultative heterochromatin function is compromised when canonical and variant H3 gene copy number falls below a critical threshold. [ABSTRACT FROM AUTHOR]

Switch/sucrose nonfermentable (SWI/SNF)-related intellectual disability disorders (SSRIDDs) and Cornelia de Lange syndrome are rare syndromic neurodevelopmental disorders with overlapping clinical phenotypes. SSRIDDs are associated with the BAF (Brahma-Related Gene-1 associated factor) complex, whereas CdLS is a disorder of chromatin modification associated with the cohesin complex. Here, we used RNA interference in Drosophila melanogaster to reduce the expression of six genes (brm, osa, Snr1, SMC1, SMC3, vtd) orthologous to human genes associated with SSRIDDs and CdLS. These fly models exhibit changes in sleep, activity, startle behavior (a proxy for sensorimotor integration), and brain morphology. Whole genome RNA sequencing identified 9,657 differentially expressed genes (FDR < 0.05), 156 of which are differentially expressed in both sexes in SSRIDD- and CdLS-specific analyses, including Bap60, which is orthologous to SMARCD1, an SSRIDD-associated BAF component. k-means clustering reveals genes co-regulated within and across SSRIDD and CdLS fly models. RNAi-mediated reduction of expression of six genes co-regulated with focal genes brm, osa, and/or Snr1 recapitulated changes in the behavior of the focal genes. Based on the assumption that fundamental biological processes are evolutionarily conserved, Drosophila models can be used to understand underlying molecular effects of variants in chromatin-modification pathways and may aid in the discovery of drugs that ameliorate deleterious phenotypic effects. [ABSTRACT FROM AUTHOR]

Simple Summary: Myelofibrosis is a heterogeneous disease regarding its mutation landscape as well as its clinical presentation and outcome. A genomic classification with prognostic implications of myeloproliferative neoplasms has been previously proposed, however, this classification has hardly been implemented in myelofibrosis. The aim of our work is to evaluate this genomic classification in a large series of myelofibrosis patients from a single institution, taking into account whether cases are primary or secondary. We found that both primary and secondary myelofibrosis have distinctive molecular landscapes and that genomic profiling provides accurate information regarding prognosis and disease progression. Myelofibrosis (MF) is a heterogeneous disease regarding its mutational landscape, clinical presentation, and outcomes. The aim of our work is to evaluate the genomic classification of MF considering whether it is primary or secondary. One-hundred seventy-five patients, 81 with primary MF (PMF) and 94 with secondary MF (SMF) were hierarchically allocated into eight molecular groups. We found that TP53 disruption/aneuploidy (n = 16, 9%) was more frequent (12% versus 6%) and showed higher allele burden (57% versus 15%, p = 0.01) in SMF than in PMF, and was associated with shorter survival (median 3.5 years). Mutations in chromatin/spliceosome genes (n = 72, 41%) represented the most frequent genomic group in PMF. Homozygous JAK2 mutation (n = 40, 23%) was enriched with old patients with SMF after long-standing polycythemia vera, whereas MF with heterozygous JAK2 mutation (n = 22, 13%) was similarly distributed among PMF and SMF. MF with CALR mutation (n = 19, 11%) predominated in post-essential thrombocythemia MF. The remaining genomic groups were infrequent. TP53 disruption, chromatin/spliceosome mutation, and homozygous JAK2 mutation were associated with significantly shorter survival and higher risk of progression. In conclusion, genomic classification reveals different pathogenic pathways between PMF and SMF and provides relevant information regarding disease phenotype and outcomes. [ABSTRACT FROM AUTHOR]

Assisted reproductive technology (ART) has become common amongst couples with infertility issues. ART is known to be successful, but epidemiological data indicates that ART is associated with placental disorders. Additionally, reports show increased risks of short- and long-term complications in children born to mothers undergoing ART. However, the mechanisms responsible for these events are obscure. The placenta is considered as a key organ for programming of diseases and ART procedures are suggested to alter the placental function and intrauterine growth trajectories. Epigenetic changes in maternal and foetal tissues are suggested to be the underlying mechanisms for these outcomes. Epigenetic regulation is known to evolve following fertilisation and before implantation and subsequently across gestation. During these critical periods of epigenetic 'programming', DNA methylation and chromatin remodelling influence the placental structure and function by regulating the expression of various genes. ART treatment coinciding with epigenetic 'programming' events during gametogenesis and early embryo development may alter the programming phases leading to long-term consequences. Thus, disruptions in placental development observed in ART pregnancies could be associated with altered epigenetic regulation of vital genes in the placenta. The review summarises available literature on the influence of ART procedures on epigenetic changes in the placenta. [ABSTRACT FROM AUTHOR]

Simple Summary: Glioblastoma is the most malignant brain tumor. To date, there is no curative therapy available. Since EGFR is an interesting candidate in precision medicine, we performed an integrated molecular analysis in glioblastoma with and without EGFR amplification. We found that there are significant molecular differences in glioblastoma, depending on the EGFR amplification state. Analysis of top differences revealed DNA replication and packaging, and chromatin and gene silencing pathways. Targeting these altered pathways may open novel targets in precision medicine. Glioblastoma IDH wildtype is the most frequent brain tumor in adults. It shows a highly malignant behavior and devastating outcomes. To date, there is still no targeted therapy available; thus, patients' median survival is limited to 12–15 months. Epithelial growth factor receptor (EGFR) is an interesting targetable candidate in advanced precision medicine for brain tumor patients. In this study, we performed integrated epigenome-wide DNA-methylation profiling of 866,895 methylation specific sites in 50 glioblastoma IDH wildtype samples, comparing EGFR amplified and non-amplified glioblastomas. We found 9849 significantly differentially methylated CpGs (DMCGs) with Δβ ≥ 0.1 and p-value < 0.05 in EGFR amplified, compared to EGFR non-amplified glioblastomas. Of these DMCGs, 2380 were annotated with tiling (2090), promoter (117), gene (69) and CpG islands (104); 7460 are located at other loci. Interestingly, the list of differentially methylated genes allocated eleven functionally relevant RNAs: five miRNAs (miR1180, miR1255B1, miR126, miR128-2, miR3125), two long non-coding RNAs (LINC00474, LINC01091), and four antisense RNAs (EPN2-AS1, MNX1-AS2, NKX2-2-AS1, WWTR1-AS1). Gene ontology (GO) analysis showed enrichment of "DNA replication-dependent nucleosome assembly", "chromatin silencing at rDNA", "regulation of gene silencing by miRNA", "DNA packaging", "posttranscriptional gene silencing", "gene silencing by RNA", "negative regulation of gene expression, epigenetic", "regulation of gene silencing", "protein-DNA complex subunit organization", and "DNA replication-independent nucleosome organization" pathways being hypomethylated in EGFR amplified glioblastomas. In summary, dissecting the methylomes of EGFR amplified and non-amplified glioblastomas revealed altered DNA replication, DNA packaging, chromatin silencing and gene silencing pathways, opening potential novel targets for future precision medicine. [ABSTRACT FROM AUTHOR]

Simple Summary: Programmed death-ligand 1 (PD-L1) enables immune evasion of tumors. Antibodies targeting PD-L1/PD-1 exhibit durable responses in eligible patients. However, antibodies cause life-threatening toxicities. Small molecules targeting PD-L1, or the drivers of PD-L1 or PD-L1/PD-1 axis, are being explored as alternatives. Thus, identifying vitamin D/vitamin D receptor (VDR) as the driver of PD-L1 expression in AML significantly enhances our understanding of the origin of PD-L1-driven immune evasions in AML and malignancies of pancreas and ovaries, where similar transcriptional regulation has been observed. To target vitamin D/VDR, we have developed MeTC7, which inhibits PD-L1 expression in vitro and in vivo and provides a new approach to block PD-L1/PD-1-driven tumorigenesis. Small-molecule inhibitors of PD-L1 are postulated to control immune evasion in tumors similar to antibodies that target the PD-L1/PD-1 immune checkpoint axis. However, the identity of targetable PD-L1 inducers is required to develop small-molecule PD-L1 inhibitors. In this study, using chromatin immunoprecipitation (ChIP) assay and siRNA, we demonstrate that vitamin D/VDR regulates PD-L1 expression in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) cells. We have examined whether a VDR antagonist, MeTC7, can inhibit PD-L1. To ensure that MeTC7 inhibits VDR/PD-L1 without off-target effects, we examined competitive inhibition of VDR by MeTC7, utilizing ligand-dependent dimerization of VDR-RXR, RXR-RXR, and VDR-coactivators in a mammalian 2-hybrid (M2H) assay. MeTC7 inhibits VDR selectively, suppresses PD-L1 expression sparing PD-L2, and inhibits the cell viability, clonogenicity, and xenograft growth of AML cells. MeTC7 blocks AML/mesenchymal stem cells (MSCs) adhesion and increases the efferocytotic efficiency of THP-1 AML cells. Additionally, utilizing a syngeneic colorectal cancer model in which VDR/PD-L1 co-upregulation occurs in vivo under radiation therapy (RT), MeTC7 inhibits PD-L1 and enhances intra-tumoral CD8+T cells expressing lymphoid activation antigen-CD69. Taken together, MeTC7 is a promising small-molecule inhibitor of PD-L1 with clinical potential. [ABSTRACT FROM AUTHOR]

Simple Summary: Mitotane (MTT) is the treatment of choice for adrenocortical carcinoma. Male hypogonadism is often diagnosed in male patients treated with this drug. This research aims to consider possible side effects induced by MTT in the testis and a hypothetical detrimental effect deriving from androgenic deficiency. Furthermore, considering the increased potentiality of treatment for adrenocortical carcinoma, we want to provide male patients of childbearing age, as in other oncological diseases, the opportunity to consider a sperm cryopreservation program. Background: Clinical evidence has shown frequent hypogonadism following mitotane (MTT) treatment in male patients with adrenocortical carcinoma. This study aimed to evaluate the impact of MTT on male gonadal function. Methods: Morphological analysis of testes and testosterone assays were performed on adult CD1 MTT-treated and untreated mice. The expression of key genes involved in interstitial and tubular compartments was studied by real-time PCR. Moreover, quantitative and qualitative analysis of spermatozoa was performed. Results: Several degrees of damage to the testes and a significant testosterone reduction in MTT-treated mice were observed. A significant decline in 3βHsd1 and Insl3 mRNA expression in the interstitial compartment confirmed an impairment of androgen production. Fsh-R mRNA expression was unaffected by MTT, proving that Sertoli cells are not the drug's primary target. Sperm concentrations were significantly lower in MTT-treated animals. Moreover, the drug caused a significant increase in the percentage of spermatozoa with abnormal chromatin structures. Conclusion: MTT negatively affects the male reproductive system, including changes in the morphology of testicular tissue and reductions in sperm concentration and quality. [ABSTRACT FROM AUTHOR]

Chromosome breakage plays an important role in the evolution of karyotypes and can produce deleterious effects within a single individual, such as aneuploidy or cancer. Forces that influence how and where chromosomes break are not fully understood. In humans, breakage tends to occur in conserved hotspots called common fragile sites (CFS), especially during replication stress. By following the fate of dicentric chromosomes in Drosophila melanogaster, we find that breakage under tension also tends to occur in specific hotspots. Our experimental approach was to induce sister chromatid exchange in a ring chromosome to generate a dicentric chromosome with a double chromatid bridge. In the following cell division, the dicentric bridges may break. We analyzed the breakage patterns of 3 different ring-X chromosomes. These chromosomes differ by the amount and quality of heterochromatin they carry as well as their genealogical history. For all 3 chromosomes, breakage occurs preferentially in several hotspots. Surprisingly, we found that the hotspot locations are not conserved between the 3 chromosomes: each displays a unique array of breakage hotspots. The lack of hotspot conservation, along with a lack of response to aphidicolin, suggests that these breakage sites are not entirely analogous to CFS and may reveal new mechanisms of chromosome fragility. Additionally, the frequency of dicentric breakage and the durability of each chromosome's spindle attachment vary significantly between the 3 chromosomes and are correlated with the origin of the centromere and the amount of pericentric heterochromatin. We suggest that different centromere strengths could account for this. [ABSTRACT FROM AUTHOR]

Simple Summary: Single-cell DNA sequencing is a laboratory technique that analyzes the genetic content of individual cells. In the context of genetically diverse hematological cancers such as acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), the traditional approach of analyzing genetic material, which typically involves bulk samples of leukemia cells, may miss important mutations that may not be present in all cells. The single-cell DNA technique can better identify genetic mutations associated with disease recurrence and death even after stem-cell transplantation. The paper discusses the background, pitfalls, and applications of this technology when used during transplantation procedures. The use of this technology could potentially translate to better outcomes in AML/MDS patients receiving stem-cell transplantation. Acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) are genetically complex and diverse diseases. Such complexity makes challenging the monitoring of response to treatment. Measurable residual disease (MRD) assessment is a powerful tool for monitoring response and guiding therapeutic interventions. This is accomplished through targeted next-generation sequencing (NGS), as well as polymerase chain reaction and multiparameter flow cytometry, to detect genomic aberrations at a previously challenging leukemic cell concentration. A major shortcoming of NGS techniques is the inability to discriminate nonleukemic clonal hematopoiesis. In addition, risk assessment and prognostication become more complicated after hematopoietic stem-cell transplantation (HSCT) due to genotypic drift. To address this, newer sequencing techniques have been developed, leading to more prospective and randomized clinical trials aiming to demonstrate the prognostic utility of single-cell next-generation sequencing in predicting patient outcomes following HSCT. This review discusses the use of single-cell DNA genomics in MRD assessment for AML/MDS, with an emphasis on the HSCT time period, including the challenges with current technologies. We also touch on the potential benefits of single-cell RNA sequencing and analysis of accessible chromatin, which generate high-dimensional data at the cellular resolution for investigational purposes, but not currently used in the clinical setting. [ABSTRACT FROM AUTHOR]

Simple Summary: R-loops, also referred to as RNA: DNA hybrids, are tripartite structures formed when a single-stranded RNA molecule forms a complex with a double-stranded DNA duplex via homology-directed base pairing. Many cellular functions are thought to be mediated by R-loops, including gene regulation, DNA replication, chromatin patterning, and DNA repair. R-loops are common and dynamic structures that account for up to 5% of mammalian genomes. In this review, we present an updated review of R-loops at a specific locus, telomeres. We summarize recent advances in the protein factors that regulate the formation and resolution of R-loops at telomeres. We also discuss the role of persistent or unscheduled R-loops in promoting genome and telomere instability and their links to human diseases. Telomeric repeat containing RNA (TERRA) is transcribed from subtelomeric regions to telomeres. TERRA RNA can invade telomeric dsDNA and form telomeric R-loop structures. A growing body of evidence suggests that TERRA-mediated R-loops are critical players in telomere length homeostasis. Here, we will review current knowledge on the regulation of R-loop levels at telomeres. In particular, we will discuss how the central player TERRA and its binding proteins modulate R-loop levels through various mechanisms. We will further provide an overview of the consequences of TERRA-mediated persistent or unscheduled R-loops at telomeres in human ALT cancers and other organisms, with a focus on telomere length regulation after replication interference-induced damage and DNA homologous recombination-mediated repair. [ABSTRACT FROM AUTHOR]

Objectives: In recent years, smoking water pipes or hookah has increased among adolescents in most countries. Although there is evidence in support of the negative effects of this type of smoking on human health, such as the increased risk of lung disease, little is known about the potential effects of hookah smoking on the male reproductive system, especially on the molecular aspects of sperm. Patients and methods: This cross-sectional study examined sperm DNA fragmentation index, protamine 1 and 2 (PRM1 and PRM2) genes expression, and oxidant status in normozoospermic hookah smokers in comparison with non-smoker controls. Results: Our results showed significantly higher rates of DNA fragmentation, protamine deficiency, and abnormal chromatin condensation in the spermatozoa of hookah smokers (P <.0001). Also, protamine gene expression showed a remarkable decrease in hookah smokers (1.55 ± 2.54 and 0.33 ± 0.54) compared to the controls (3.49 ± 5.41 and 1.22 ± 1.96), although the reduction was not statistically significant (P =.155 and P =.066, respectively). Moreover, a significantly higher level of semen MDA was observed in the case group compared to the controls (0.39 ± 1.04 vs 0.15 ± 0.21; P =.013). Conclusion: According to our study, although hookah smoking does not have a significant effect on sperm parameters, it may have deleterious effects on DNA integrity, oxidative status, and nuclear protein levels of spermatozoa. [ABSTRACT FROM AUTHOR]

Background: Increased sperm DNA damage is known as one of the causes of recurrent pregnancy loss (RPL) which can be due to increased levels of oxidative stress. Therefore, the aim of this study was to assess the effect of alphalipoic acid (ALA) on sperm parameters and sperm functions in couples with a history of RPL. Materials and Methods: In this post hoc analysis in clinical trial study, a total of 37 couples with RPL (n=12 and n=25 for placebo and ALA groups, respectively) were considered. Men were treated with ALA (600 mg/day) or placebo for 80 days. Semen samples were acquired from the participants before initiation and after completion of the medication course and assessed regarding conventional sperm parameters, chromatin damage/integrity, intracellular oxidative stress, lipid peroxidation, and seminal antioxidant characteristics. Individuals were further followed up for twelve months for pregnancy occurrence and outcomes. Finally, after excluding patients with no history of RPL, the data was analyzed. Results: No significant differences were observed between the baseline measures of the aforementioned parameters except for seminal volume. After the intervention, the mean sperm DNA damage, protamine deficiency, and persisted histones were significantly lower in the ALA group than in placebo receivers (P<0.05). A decrease in the mean of seminal total antioxidant capacity (P=0.03), malondialdehyde (P=0.02), and sperm DNA damage (P=0.004) as well as an increase in sperm total motility (P=0.04) after treatment with ALA was noticed. In addition, the mean of protamine deficiency and persisted histones were declined post-ALA therapy (P=0.003 and 0.002, respectively). The percentage of spontaneous pregnancy in the ALA group (4 of 25 cases; 16%) was higher than in the placebo group (1 of 12, 8.3%). Conclusion: ALA-therapy attenuates sperm DNA damage and lipid peroxidation while enhancing sperm total motility and chromatin compaction in the male partner of couples with PRL (registration number: IRCT20190406043177N1). [ABSTRACT FROM AUTHOR]

OBJECTIVES: In recent years, smoking water pipes or hookah has increased among adolescents in most countries. Although there is evidence in support of the negative effects of this type of smoking on human health, such as the increased risk of lung disease, little is known about the potential effects of hookah smoking on the male reproductive system, especially on the molecular aspects of sperm. PATIENTS AND METHODS: This cross-sectional study examined sperm DNA fragmentation index, protamine 1 and 2 (PRM1 and PRM2) genes expression, and oxidant status in normozoospermic hookah smokers in comparison with non-smoker controls. RESULTS: Our results showed significantly higher rates of DNA fragmentation, protamine deficiency, and abnormal chromatin condensation in the spermatozoa of hookah smokers (P < .0001). Also, protamine gene expression showed a remarkable decrease in hookah smokers (1.55 ± 2.54 and 0.33 ± 0.54) compared to the controls (3.49 ± 5.41 and 1.22 ± 1.96), although the reduction was not statistically significant (P= .155 and P = .066, respectively). Moreover, a significantly higher level of semen MDA was observed in the case group compared to the controls (0.39 ± 1.04 vs 0.15 ± 0.21; P = .013). CONCLUSION: According to our study, although hookah smoking does not have a significant effect on sperm parameters, it may have deleterious effects on DNA integrity, oxidative status, and nuclear protein levels of spermatozoa. [ABSTRACT FROM AUTHOR]

This study explores whether resveratrol effectively protects the reproductive system against isoflurane-induced toxicity in testicular tissue. In this experiment, we randomly divided 60 adult male C57BL/6 mice into six groups (n = 10). Five consecutive days per week, mice were exposed to 1.5% isoflurane for 1 h/day and were given 50 and 100 mg/kg resveratrol. After 35 days (the completion of the mouse spermatogenesis period), the left testis was removed for histomorphometric evaluations, while the right testis was used to determine the Capacity of total antioxidants and lipid peroxidation. To analyze the Parameters of sperm, chromatin maturation, and DNA fragmentation, the left caudal epididymis was used. Based on a one-way analysis of variance (ANOVA), we considered a difference in means of 0.05 to be significant (P0.05). Compared to the control group, the isoflurane group showed a significant decrease in testicular weight, volume, sperm parameters, and tissue histomorphometry. Comparatively, to the control group, malondialdehyde levels increased, and the total antioxidant capacity decreased significantly. Resveratrol improved all of the above parameters in the simultaneous treatment groups compared to the isoflurane group. It did not, however, reach the level of the control group in all cases. It has been demonstrated that resveratrol, with its powerful antioxidant properties, reduces the reproductive toxicity of isoflurane by inhibiting free radicals and increasing the testicular tissue's antioxidant capacity. [ABSTRACT FROM AUTHOR]

Genome-wide association studies have identified over 100 loci associated with osteoarthritis risk, but the majority of osteoarthritis risk variants are noncoding, making it difficult to identify the impacted genes for further study and therapeutic development. To address this need, we used a multiomic approach and genome editing to identify and functionally characterize potential osteoarthritis risk genes. Computational analysis of genome-wide association studies and ChIP-seq data revealed that chondrocyte regulatory loci are enriched for osteoarthritis risk variants. We constructed a chondrocyte-specific regulatory network by mapping 3D chromatin structure and active enhancers in human chondrocytes. We then intersected these data with our previously collected RNA-seq dataset of chondrocytes responding to fibronectin fragment, a known osteoarthritis trigger. Integration of the 3 genomic datasets with recently reported osteoarthritis genome-wide association study variants revealed a refined set of putative causal osteoarthritis variants and their potential target genes. One of the putative target genes identified was SOCS2, which was connected to a putative causal variant by a 170-kb loop and is differentially regulated in response to fibronectin fragment. CRISPR-Cas9-mediated deletion of SOCS2 in primary human chondrocytes from 3 independent donors led to heightened expression of inflammatory markers after fibronectin fragment treatment. These data suggest that SOCS2 plays a role in resolving inflammation in response to cartilage matrix damage and provides a possible mechanistic explanation for its influence on osteoarthritis risk. In total, we identified 56 unique putative osteoarthritis risk genes for further research and potential therapeutic development. [ABSTRACT FROM AUTHOR]

Summary: Background: Childhood obesity rates are on the rise, but there are currently no effective therapies available to slow or halt their progression. Although environmental and lifestyle factors have been implicated in its pathogenesis, childhood obesity is considered a complex disorder with a clear genetic component. Intense genome‐wide association study (GWAS) efforts through large‐scale collaborations have enabled the discovery of genetic loci robustly associated with childhood obesity beyond the classic FTO locus. That said, GWAS itself does not pinpoint the actual underlying causal effector genes, but rather just yields association signals in the genome. Objective: This review aims to outline what has been elucidated thus far on the genetic aetiology of commong childhood obesity and to describe strategies to identify and validate both causal common genetic variants and their corresponding effector genes. Results: Relevant cell types for molecular studies can be identified by gene set enrichment analysis and considering known biology of obesity‐related physiological processes. Putatively causal single nucleotide polymorphisms (SNPs) can be identified by several methods including statistical fine mapping and 'assay for transposase accessible chromatin sequencing' (ATAC‐seq). Variant to gene mapping can then nominate effector genes likely regulated by cis‐regulatory elements harbouring putatively causal SNPs. A SNP's cis‐regulatory activity can be functionally validated by several in vitro methods including luciferase assay and CRISPR approaches. These CRISPR approaches can also be used to investigate how dysregulatn of effector genes may confer obesity risk. Conclusion: Uncovering the causative genes related to GWAS signals and elucidating their functional contributions to paediatric obesity with these strategies will deepen our understanding of this disease and serve better treatment outcomes. [ABSTRACT FROM AUTHOR]

Xenopus tropicalis tadpoles have the capacity for scarless regeneration of appendages including the limb and tail. Following injury, transcriptional programs must be activated and inactivated with high spatial and temporal resolution to result in a properly patterned appendage. Functional studies have established that histone‐modifying enzymes that act to close chromatin are required for regeneration, but the genomic regions sensitive to these activities are not fully established. Here we show that early inhibition of HDAC or EZH2 activity results in incomplete tail regeneration. To identify how each of these perturbations impacts chromatin accessibility, we applied an assay for transposase‐accessible chromatin (ATAC‐seq) to HDAC or EZH2‐inhibited regenerating tadpoles. We find that neither perturbation results in a global increase in chromatin accessibility, but that both inhibitors have targeted effects on chromatin accessibility and gene expression. Upon HDAC inhibition, regulatory regions neighbouring genes associated with neuronal regeneration are preferentially accessible, whereas regions associated with immune response and apoptosis are preferentially accessible following EZH2 inhibition. Together, these results suggest distinct roles for these two chromatin‐closing activities in appendage regeneration. [ABSTRACT FROM AUTHOR]

Introduction: The aim of the study was to examine the protective efficacy of astaxanthin (ASTA) against the damage that occurs during sperm cryopreservation. Methods: This experimental study was carried out on waste semen samples of 30 normozoospermic individuals who applied for semen analysis. Semen samples were divided into four equal volumes and 0 µM (control group), 50 µM, 100 µM, and 500 µM ASTA were added to each group. All groups were stored frozen in a liquid nitrogen tank. Semen samples were removed from liquid nitrogen after 72 hours and were thawed. Motility evaluation of sperm was performed. In addition, sperm was stained with acidic aniline blue to detect DNA chromatin condensation. Results: The highest motility loss was found in the control group and the least motility loss was in the 100 µM ASTA group. When examined in terms of sperm chromatin condensation, condensed sperm count was higher in the 100 µM ASTA group than in the other groups. Conclusions: It has been observed that ASTA added to the cryoprotectant substance during sperm cryopreservation positively affects sperm motility and reduces the number of decondensed sperm. [ABSTRACT FROM AUTHOR]

In Drosophila chromosomal rearrangements can be maintained and are associated with karyotypic variability among populations from different geographic localities. The abundance of variability in gene arrangements among chromosomal arms is even greater when comparing more distantly related species and the study of these chromosomal changes has provided insights into the evolutionary history of species in the genus. In addition, the sequencing of genomes of several Drosophila species has offered the opportunity to establish the global pattern of genomic evolution, at both genetic and chromosomal level. The combined approaches of comparative analysis of syntenic blocks and direct physical maps on polytene chromosomes have elucidated changes in the orientation of genomic sequences and the difference between heterochromatic and euchromatic regions. Unfortunately, the centromeric heterochromatic regions cannot be studied using the cytological maps of polytene chromosomes because they are underreplicated and therefore reside in the chromocenter. In Drosophila melanogaster, a cytological map of the heterochromatin has been elaborated using mitotic chromosomes from larval neuroblasts. In the current work, we have expanded on that mapping by producing cytological maps of the mitotic heterochromatin in an additional 10 sequenced Drosophila species. These maps highlight 2 apparently different paths, for the evolution of the pericentric heterochromatin between the subgenera Sophophora and Drosophila. One path leads toward a progressive complexity of the pericentric heterochromatin (Sophophora) and the other toward a progressive simplification (Drosophila). These maps are also useful for a better understanding how karyotypes have been altered by chromosome arm reshuffling during evolution. [ABSTRACT FROM AUTHOR]

The functioning of the secretory organs is closely related to the activity of the immune system. As is well known, this participation is manifested in the fact that at certain stages of activity, the lymphoid cells migrating to the organ can be involved in the regulation of secretion. In addition, the products of the immune system and even its cellular elements can become components of a number of secrets. Colostrum and milk contain a large number of cells of a wide spectrum (up to 1/3 of the volume), of which the number of lymphocytes is up to 16% of leukocytes. Lymphocytes, in an immunologically active form, entering the newborn’s body with colostrum, activate the cellular immunity system. The transport of lymphokinin mediators plays a certain role in this process. Microphages, T- and B-lymphocytes, penetrating through the intercellular spaces into the lymphoid layer of the intestine, transmit immunoreceptors to the prolymphocytes of the newborn, "armed" with their activity to recognize genetically foreign ones. The lymphocytes contained in colostrum are the cells of the immune system that provide cellular and humoral immunity. They are mainly represented by T-cells, B-cells and killer cells. Milk T-cells produce a full spectrum of immune regulatory proteins such as interferon, tumor necrosis factor alpha. These cells are the cells of the immune memory. Newborns who received the first portion of colostrum no later than an hour after birth are characterized by an increased number of leukocytes, more pronounced phagocytosis, which indicates the stimulation of hemo- and lymphocytosis. When carrying out transmission and scanning electron microscopy in the epithelial layer of the intestine, cellular elements were found that got there from the intestinal lumen. Microsections show how cells of a lymphoid nature, pushing apart the structures of the epithelial layer, bypass natural barriers and, at the same time, retain their physiological usefulness. The possibility of penetration of immunocompetent cells of the mother’s colostrum into the bloodstream of the young is proved using the natural label of the female’s cells – sex chromatin. Naturally, sex chromatin-labeled cells were sought in male newborns. The detection of colostrum cells in the intestinal wall and bloodstream of the young is approximately 25% in the blood, 1% in the lymph, and about 70% in the intestine. There is no doubt that the leukocytes of colostrum are of exceptional importance in creating immunity in newborn animals.

Simple Summary: Mantle cell lymphoma (MCL) is much more common in males than in females. The reason for this is not clear, but research has indicated that the female sex hormones, estrogens, have a protective effect on MCL development. To study this further, mice were transplanted with MCL cells and treated with an estrogen that selectively activates ESR2, the main nuclear estrogen receptor in lymphoma cells. The activation of ESR2 resulted in reduced MCL tumor growth of MCL tumors that were both sensitive and resistant to a newly developed drug (ibrutinib). The mechanism for this effect was investigated by analyzing gene expression and ESR2 binding to target genes. The results show that the affected genes were enriched in several malignancy-related biological processes, including MCL. Furthermore, the results suggested an interplay between the lymphoma cells and the tumor microenvironment in response to ESR2 activation. Altogether, the results clarify the mechanisms of ESR2-mediated MCL growth impairment by estrogens and provide a possible explanation for the sex difference in incidence. Furthermore, targeting ESR2 may be an option when considering the treatment of MCL. Mantle cell lymphoma (MCL) is a non-Hodgkin lymphoma with one of the highest male-to-female incidence ratios. The reason for this is not clear, but epidemiological as well as experimental data have suggested a role for estrogens, particularly acting through estrogen receptor β (ESR2). To study the ESR2 effects on MCL progression, MCL cells sensitive and resistant to the Bruton tyrosine kinase inhibitor ibrutinib were grafted to mice and treated with the ESR2-selective agonist diarylpropionitrile (DPN). The results showed that the DPN treatment of mice grafted with both ibrutinib-sensitive and -resistant MCL tumors resulted in impaired tumor progression. To identify the signaling pathways involved in the impaired tumor progression following ESR2 agonist treatment, the transcriptome and ESR2 binding to target genes were investigated by genome-wide chromatin immunoprecipitation in Granta-519 MCL tumors. DPN-regulated genes were enriched in several biological processes that included cell–cell adhesion, endothelial–mesenchymal transition, nuclear factor-kappaB signaling, vasculogenesis, lymphocyte proliferation, and apoptosis. In addition, downregulation of individual genes, such as SOX11 and MALAT1, that play a role in MCL progression was also observed. Furthermore, the data suggested an interplay between the lymphoma cells and the tumor microenvironment in response to the ESR2 agonist. In conclusion, the results clarify the mechanisms by which estrogens, via ESR2, impair MCL tumor progression and provide a possible explanation for the sex-dependent difference in incidence. Furthermore, targeting ESR2 with a selective agonist may be an additional option when considering the treatment of both ibrutinib-sensitive and -resistant MCL tumors. [ABSTRACT FROM AUTHOR]

Simple Summary: Tumors of the brain and nerves develop frequently in patients with neurofibromatosis type 1. Many are benign growths, such as pilocytic astrocytomas in the brain and neurofibromas in the nerves. However, in some patients, the tumors become malignant and may cause local damage, disseminate to distant sites and result in death. We studied changes in the levels of chromatin proteins and changes in telomeres, in cells obtained from mouse gliomas that are deficient in neurofibromin as well as excess brain and nerve tumor tissue from patients with neurofibromatosis type 1 or sporadic tumors lacking neurofibromin expression. A decrease in the levels of these proteins in experimental cell lines resulted in susceptibility to a class of specific drugs knowns as ATR inhibitors, which may represent a specific vulnerability of these tumor subgroups. We expect our data to provide the required rationale for the development of more accurate animal models to study neurofibromatosis, as well as specific molecularly based drugs for treatment as alternatives to the current, often devastating approaches of surgery, radiation, and chemotherapy. Subsets of Neurofibromatosis Type 1 (NF1)-associated solid tumors have been shown to display high frequencies of ATRX mutations and the presence of alternative lengthening of telomeres (ALT). We studied the phenotype of combined NF1 and ATRX deficiency in malignant solid tumors. Cell lines derived from NF1-deficient sporadic glioblastomas (U251, SF188), an NF1-associated ATRX mutant glioblastoma cell line (JHH-NF1-GBM1), an NF1-derived sarcoma cell line (JHH-CRC65), and two NF1-deficient MPNST cell lines (ST88-14, NF90.8) were utilized. Cancer cells were treated with ATR inhibitors, with or without a MEK inhibitor or temozolomide. In contrast to the glioma cell line SF188, combined ATRX knockout (KO) and TERC KO led to ALT-like properties and sensitized U251 glioma cells to ATR inhibition in vitro and in vivo. In addition, ATR inhibitors sensitized U251 cells to temozolomide, but not MEK inhibition, irrespective of ATRX level manipulation; whereas, the JHH-NF1-GBM1 cell line demonstrated sensitivity to ATR inhibition, but not temozolomide. Similar effects were noted using the MPNST cell line NF90.8 after combined ATRX knockdown and TERC KO; however, not in ST88-14. Taken together, our study supports the feasibility of targeting the ATR pathway in subsets of NF1-deficient and associated tumors. [ABSTRACT FROM AUTHOR]

Specific gene transcriptional programs are required to ensure the proper proliferation and differentiation processes underlying the production of specialized cells during development. Gene activity is mainly regulated by the concerted action of transcription factors and chromatin proteins. In the nematode Caenorhabditis elegans, mechanisms that silence improper transcriptional programs in germline and somatic cells have been well studied, however, how are tissue-specific sets of genes turned on is less known. LSL-1 is herein defined as a novel crucial transcriptional regulator of germline genes in C. elegans. LSL-1 is first detected in the P4 blastomere and remains present at all stages of germline development, from primordial germ cell proliferation to the end of meiotic prophase. lsl-1 loss-of-function mutants exhibit many defects including meiotic prophase progression delay, a high level of germline apoptosis, and production of almost no functional gametes. Transcriptomic analysis and ChIP-seq data show that LSL-1 binds to promoters and acts as a transcriptional activator of germline genes involved in various processes, including homologous chromosome pairing, recombination, and genome stability. Furthermore, we show that LSL-1 functions by antagonizing the action of the heterochromatin proteins HPL-2/HP1 and LET-418/Mi2 known to be involved in the repression of germline genes in somatic cells. Based on our results, we propose LSL-1 to be a major regulator of the germline transcriptional program during development. [ABSTRACT FROM AUTHOR]

Simple Summary: Alternative lengthing of telomere (ALT) is an important mechanism for maintaining telomere length and cell proliferation in telomerase-negative tumor cells. However, the molecular mechanism of ALT is still poorly understood. ALT occurs in a wide range of tumor types and usually associated with a worse clinical consequence. Here, we review the recent findings of ALT mechanisms, which promise ALT could be a valuable drug target for clinical telomerase-negative tumor treatment. Telomeres are DNA–protein complexes that protect eukaryotic chromosome ends from being erroneously repaired by the DNA damage repair system, and the length of telomeres indicates the replicative potential of the cell. Telomeres shorten during each division of the cell, resulting in telomeric damage and replicative senescence. Tumor cells tend to ensure cell proliferation potential and genomic stability by activating telomere maintenance mechanisms (TMMs) for telomere lengthening. The alternative lengthening of telomeres (ALT) pathway is the most frequently activated TMM in tumors of mesenchymal and neuroepithelial origin, and ALT also frequently occurs during experimental cellular immortalization of mesenchymal cells. ALT is a process that relies on homologous recombination (HR) to elongate telomeres. However, some processes in the ALT mechanism remain poorly understood. Here, we review the most recent understanding of ALT mechanisms and processes, which may help us to better understand how the ALT pathway is activated in cancer cells and determine the potential therapeutic targets in ALT pathway-stabilized tumors. [ABSTRACT FROM AUTHOR]

Objective: To explore susceptibility loci associated with uveitis in Behçet's disease (BD). Methods: We conducted a 2‐stage study, consisting of a genome‐wide association study (GWAS) stage and a replication stage, in a Chinese population. The GWAS stage included 978 cases with BD‐related uveitis and 4,388 controls, and the replication stage included 953 cases with BD‐related uveitis and 2,129 controls. Luciferase reporter analysis and chromatin immunoprecipitation assay were performed to explore the functional role of susceptibility genetic variants near ZMIZ1. Results: Three independent HLA alleles (HLA–B51 [3.75 × 10−190], HLA–A26 [1.50 × 10−18], and HLA–C0704 [3.44 × 10−16]) were identified as having a genome‐wide association with BD‐related uveitis. In the non‐HLA region, in addition to confirming 7 previously reported loci, we identified 22 novel susceptibility variants located in 16 loci. Meta‐analysis of the Chinese cohort consisting of 1,931 cases and 6,517 controls and a published Japanese cohort of 611 cases and 737 controls showed genome‐wide significant associations with ZMIZ1, RPS6KA4, IL10RA, SIPA1‐FIBP‐FOSL1, and VAMP1. Functional experiments demonstrated that genetic variants of ZMIZ1 were associated with enhanced transcription activity and increased expression of ZMIZ1. Conclusion: This GWAS study identified a novel set of genetic variants that are associated with susceptibility to uveitis in BD. These findings enrich our understanding of the contribution of genetic factors to the disease. [ABSTRACT FROM AUTHOR]

Background: Today, vitamin D deficiency (VDD) is one of the major health issues around the world and VDD is associated with several diseases. This study was conducted to find the relationship between vitamin D status in male's serum with sperm function and clinical outcomes in infertile men candidate for intracytoplasmic sperm injection (ICSI). Materials and Methods: In this cohort study, different parameters of male fertility such as sperm parameters, oxidative stress, and sperm chromatin status were evaluated in sperm samples of 30 infertile couples candidate for ICSI. Clinical outcomes like fertilization, embryo quality, and implantation were also assessed. Data were analyzed using SPSS Statistics 25.0 software. Besides, assessment of the correlation between aforementioned parameters with the level of serum vitamin D, in this study, ICSI candidates were divided into three groups [individuals with sufficient vitamin D levels (>30 ng/ml), insufficient vitamin D levels (between 20-29 ng/ml), and VDD (<20 ng/ml)]. The aforementioned parametesr were also compared between these study groups. Results: Analysis of all the data revealed a significant correlation between the level of vitamin D with sperm concentration (P=0.000, r=0.5), sperm count (P=0.03, r=0.31) and sperm reactive oxygen species (ROS) level (P=0.000, r=-0.77). Moreover, comparing clinical outcomes within study groups showed a significant difference in implantation rate between sufficient and other groups (insufficient and deficient) (P=0.02). Conclusion: Considering the association between sperm concentration and level of ROS with vitamin D and, higher implantation rate in individuals with vitamin D sufficient group compared to other two groups, our data call for vitamin D supplementation as part of male infertility treatment. But considering our sample size, further research is needed to verify these findings. [ABSTRACT FROM AUTHOR]

Simple Summary: In bladder cancer, cisplatin remains the front-line therapy, but drug resistance is common. Previously, we showed that cancer cells can spontaneously convert to an aggressive drug-resistant phenotype without mutational events. In the current work, we explored the epigenetic mechanism behind the conversion to the drug-resistant phenotype. We discovered that drug-resistant cells have differentially accessible enhancers, which are bound by FOXC1, a transcription factor that is overexpressed in these cells. Accordingly, FOXC1 knockout significantly attenuates the emergence of the drug-resistant phenotype and reduces cell survival upon cisplatin treatment. These findings suggest that FOXC1 binding at accessible enhancers promotes cisplatin drug resistance in bladder cancer cells. Therefore, FOXC1 targeting may be a new therapeutic avenue to mitigate cisplatin resistance and improve treatment efficacy in bladder cancer. Chemotherapy resistance is traditionally attributed to DNA mutations that confer a survival advantage under drug selection pressure. However, in bladder cancer and other malignancies, we and others have previously reported that cancer cells can convert spontaneously to an aggressive drug-resistant phenotype without prior drug selection or mutational events. In the current work, we explored possible epigenetic mechanisms behind this phenotypic plasticity. Using Hoechst dye exclusion and flow cytometry, we isolated the aggressive drug-resistant cells and analyzed their chromatin accessibility at regulatory elements. Compared to the rest of the cancer cell population, the aggressive drug-resistant cells exhibited enhancer accessibility changes. In particular, we found that differentially accessible enhancers were enriched for the FOXC1 transcription factor motif, and that FOXC1 was the most significantly overexpressed gene in aggressive drug-resistant cells. ChIP-seq analysis revealed that differentially accessible enhancers in aggressive drug-resistant cells had a higher FOXC1 binding, which regulated the expression of adjacent cancer-relevant genes like ABCB1 and ID3. Accordingly, cisplatin treatment of bladder cancer cells led to an increased FOXC1 expression, which mediated cell survival and conversion to a drug-resistant phenotype. Collectively, these findings suggest that FOXC1 contributes to phenotypic plasticity by binding enhancers and promoting a mutation-independent shift towards cisplatin resistance in bladder cancer. [ABSTRACT FROM AUTHOR]

Objective: Genetic variants spanning UBE2L3 are associated with increased expression of the UBE2L3‐encoded E2 ubiquitin‐conjugating enzyme H7 (UbcH7), which facilitates activation of proinflammatory NF‐κB signaling and susceptibility to autoimmune diseases. We undertook this study to delineate how genetic variants carried on the UBE2L3/YDJC autoimmune risk haplotype function to drive hypermorphic UBE2L3 expression. Methods: We used bioinformatic analyses, electrophoretic mobility shift assays, and luciferase reporter assays to identify and functionally characterize allele‐specific effects of risk variants positioned in chromatin accessible regions of immune cells. Chromatin conformation capture with quantitative polymerase chain reaction (3C‐qPCR), chromatin immunoprecipitation (ChIP)–qPCR, and small interfering RNA (siRNA) knockdown assays were performed on patient‐derived Epstein‐Barr virus–transformed B cells homozygous for the UBE2L3/YDJC nonrisk or risk haplotype to determine if the risk haplotype increases UBE2L3 expression by altering the regulatory chromatin architecture in the region. Results: Of the 7 prioritized variants, 5 demonstrated allele‐specific increases in nuclear protein binding affinity and regulatory activity. High‐throughput sequencing of chromosome conformation capture coupled with ChIP (HiChIP) and 3C‐qPCR uncovered a long‐range interaction between the UBE2L3 promoter (rs140490, rs140491, rs11089620) and the downstream YDJC promoter (rs3747093) that was strengthened in the presence of the UBE2L3/YDJC risk haplotype, and correlated with the loss of CCCTC‐binding factor (CTCF) and gain of YY1 binding at the risk alleles. Depleting YY1 by siRNA disrupted the long‐range interaction between the 2 promoters and reduced UBE2L3 expression. Conclusion: The UBE2L3/YDJC autoimmune risk haplotype increases UBE2L3 expression through strengthening a YY1‐mediated interaction between the UBE2L3 and YDJC promoters. [ABSTRACT FROM AUTHOR]

Propofol is a commonly used drug for sedation and general anesthesia during cancer surgery. Previous studies indicate that propofol exerts anti-tumor effect in various cancers. The aim of this study was to investigate the underlying molecular mechanism of propofol in liver cancer. The effects of propofol on liver cancer cells were evaluated by cell viability assay, colony formation assay, and tumor xenograft model. Dysregulated lncRNAs of propofol-treated liver cancer cells were evaluated by transcriptome RNA sequencing. The underlying molecular mechanisms of lncRNA cancer susceptibility candidate 9 (CASC9) in propofol-induced anti-tumor effects were evaluated by western blot, quantitative real-time polymerase chain reaction (qRT-PCR), wound scratch healing assay, transwell cell migration and invasion assay, TUNEL staining, fluorescence in situ hybridization, RNA immunoprecipitation (RIP), and chromatin immunoprecipitation (ChIP). We found that propofol suppressed proliferation, migration, invasion, and tumor xenograft growth of liver cancer cells in a dose-dependent manner. Exosomes transfer from propofol-treated cells inhibited proliferation, migration, and invasion and promoted apoptosis of liver cancer cells. Transcriptional profiling of propofol-treated liver cancer cells identified CASC9 as significantly downregulated lncRNA in cells and exosomes. Enforced CASC9 expression partially rescued the inhibitory effects of propofol on liver cancer cells. Furthermore, CASC9 was found to interact directly with EZH2 and epigenetically regulated PTEN expression. Restoration of CASC9 partially abrogated the inhibition of propofol on Akt/mTOR signaling. Our results indicated that propofol exerted anti-tumor effects by downregulating CASC9, and subsequently suppressed Akt/mTOR signaling. Our findings provided a novel insight into propofol-induced anti-tumor effects in liver cancer. [ABSTRACT FROM AUTHOR]

Objective: In a recent genome‐wide association study, a significant genetic association between rs34330 of CDKN1B and risk of systemic lupus erythematosus (SLE) in Han Chinese was identified. This study was undertaken to validate the reported association and elucidate the biochemical mechanisms underlying the effect of the variant. Methods: We performed an allelic association analysis in patients with SLE, followed by a meta‐analysis assessing genome‐wide association data across 11 independent cohorts (n = 28,872). In silico bioinformatics analysis and experimental validation in SLE‐relevant cell lines were applied to determine the functional consequences of rs34330. Results: We replicated a genetic association between SLE and rs34330 (meta‐analysis P = 5.29 × 10−22, odds ratio 0.84 [95% confidence interval 0.81–0.87]). Follow‐up bioinformatics and expression quantitative trait locus analysis suggested that rs34330 is located in active chromatin and potentially regulates several target genes. Using luciferase and chromatin immunoprecipitation–real‐time quantitative polymerase chain reaction, we demonstrated substantial allele‐specific promoter and enhancer activity, and allele‐specific binding of 3 histone marks (H3K27ac, H3K4me3, and H3K4me1), RNA polymerase II (Pol II), CCCTC‐binding factor, and a critical immune transcription factor (interferon regulatory factor 1 [IRF‐1]). Chromosome conformation capture revealed long‐range chromatin interactions between rs34330 and the promoters of neighboring genes APOLD1 and DDX47, and effects on CDKN1B and the other target genes were directly validated by clustered regularly interspaced short palindromic repeat (CRISPR)–based genome editing. Finally, CRISPR/dead CRISPR‐associated protein 9–based epigenetic activation/silencing confirmed these results. Gene‐edited cell lines also showed higher levels of proliferation and apoptosis. Conclusion: Collectively, these findings suggest a mechanism whereby the rs34330 risk allele (C) influences the presence of histone marks, RNA Pol II, and IRF‐1 transcription factor to regulate expression of several target genes linked to proliferation and apoptosis. This process could potentially underlie the association of rs34330 with SLE. [ABSTRACT FROM AUTHOR]

Embryonic patterning is critically dependent on zygotic genome activation (ZGA). In Drosophila melanogaster embryos, the pioneer factor Zelda directs ZGA, possibly in conjunction with other factors. Here, we have explored the novel involvement of Chromatin-Linked Adapter for MSL Proteins (CLAMP) during ZGA. CLAMP binds thousands of sites genome-wide throughout early embryogenesis. Interestingly, CLAMP relocates to target promoter sequences across the genome when ZGA is initiated. Although there is a considerable overlap between CLAMP and Zelda binding sites, the proteins display distinct temporal dynamics. To assess whether CLAMP occupancy affects gene expression, we analyzed transcriptomes of embryos zygotically compromised for either clamp or zelda and found that transcript levels of many zygotically activated genes are similarly affected. Importantly, compromising either clamp or zelda disrupted the expression of critical segmentation and sex determination genes bound by CLAMP (and Zelda). Furthermore, clamp knockdown embryos recapitulate other phenotypes observed in Zelda-depleted embryos, including nuclear division defects, centrosome aberrations, and a disorganized actomyosin network. Based on these data, we propose that CLAMP acts in concert with Zelda to regulate early zygotic transcription. [ABSTRACT FROM AUTHOR]