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Tandem repeats are frequent across the human genome, and variation in repeat length has been linked to a variety of traits. Recent improvements in long read sequencing technologies have the potential to greatly improve tandem repeat analysis, especially for long or complex repeats. Here, we introduce LongTR, which accurately genotypes tandem repeats from high-fidelity long reads available from both PacBio and Oxford Nanopore Technologies. LongTR is freely available at https://github.com/gymrek-lab/longtr and https://zenodo.org/doi/10.5281/zenodo.11403979 .

The Genome Aggregation Database (gnomAD), widely recognized as the gold-standard reference map of human genetic variation, has largely overlooked tandem repeat (TR) expansions, despite the fact that TRs constitute ∼6% of our genome and are linked to over 50 human diseases. Here, we introduce the TR-gnomAD (https://wlcb.oit.uci.edu/TRgnomAD), a biobank-scale reference of 0.86 million TRs derived from 338,963 whole-genome sequencing (WGS) samples of diverse ancestries (39.5% non-European samples). TR-gnomAD offers critical insights into ancestry-specific disease prevalence using disparities in TR unit number frequencies among ancestries. Moreover, TR-gnomAD is able to differentiate between common, presumably benign TR expansions, which are prevalent in TR-gnomAD, from those potentially pathogenic TR expansions, which are found more frequently in disease groups than within TR-gnomAD. Together, TR-gnomAD is an invaluable resource for researchers and physicians to interpret TR expansions in individuals with genetic diseases.

Objectives To determine whether the information provided by short tandem repeat (STR) testing and bone marrow (BM) biopsy specimens following hematopoietic stem cell transplant (HSCT) provides redundant information, leading to test overutilization, without additional clinical benefit. Methods Cases with synchronous STR and flow cytometric immunophenotyping (FCI) testing, as part of the BM evaluation, were assessed for STR/FCI concordance. Results Of 1199 cases (410 patients), we found the overall concordance between STR and FCI was 93%, with most cases (1063) classified as STR–/FCI–. Of all discordant cases, 75 (6%) were STR+/FCI–, with only 5 (6.7%) cases best explained as identification of disease relapse. Eight cases were STR–/FCI+, representing relapsed/residual disease. Analysis of cases 1 year or more from transplant (54% of all cases) indicated only 9 (1.5%) were STR+/FCI–, and none uniquely identified relapse. Conclusions These data suggest that STR analysis performed 1 year or more post-HSCT does not identify unknown cases of relapse. Furthermore, while STR testing is critical for identifying graft failure/rejection within the first year posttransplant, FCI appears superior to STR at detecting late relapses with low-level disease. Therefore, STR testing from patients 1 year or more post-HSCT may be unnecessary, as BM biopsy evaluation is sufficient to identify disease relapse. [ABSTRACT FROM AUTHOR]

Tandem repeats (TRs) represent one of the largest sources of genetic variation in humans and are implicated in a range of phenotypes. Here we present a deep characterization of TR variation based on high coverage whole genome sequencing from 3550 diverse individuals from the 1000 Genomes Project and H3Africa cohorts. We develop a method, EnsembleTR, to integrate genotypes from four separate methods resulting in high-quality genotypes at more than 1.7 million TR loci. Our catalog reveals novel sequence features influencing TR heterozygosity, identifies population-specific trinucleotide expansions, and finds hundreds of novel eQTL signals. Finally, we generate a phased haplotype panel which can be used to impute most TRs from nearby single nucleotide polymorphisms (SNPs) with high accuracy. Overall, the TR genotypes and reference haplotype panel generated here will serve as valuable resources for future genome-wide and population-wide studies of TRs and their role in human phenotypes.

Borgs are huge, linear extrachromosomal elements associated with anaerobic methane-oxidizing archaea. Striking features of Borg genomes are pervasive tandem direct repeat (TR) regions. Here, we present six new Borg genomes and investigate the characteristics of TRs in all ten complete Borg genomes. We find that TR regions are rapidly evolving, recently formed, arise independently, and are virtually absent in host Methanoperedens genomes. Flanking partial repeats and A-enriched character constrain the TR formation mechanism. TRs can be in intergenic regions, where they might serve as regulatory RNAs, or in open reading frames (ORFs). TRs in ORFs are under very strong selective pressure, leading to perfect amino acid TRs (aaTRs) that are commonly intrinsically disordered regions. Proteins with aaTRs are often extracellular or membrane proteins, and functionally similar or homologous proteins often have aaTRs composed of the same amino acids. We propose that Borg aaTR-proteins functionally diversify Methanoperedens and all TRs are crucial for specific Borg-host associations and possibly cospeciation.

The adjacent -2 segregation is a rare separation mode in the gametic formation of balanced translocation carriers. A fetus with severe fetal growth restriction (FGR) and chromosomal abnormalities is reported, and the prenatal diagnosis was performed by G -banding karyotype analysis, chromosomal microarray analysis (CMA), fluorescence in situ hybridization (FISH), quantitive fluorescent polymerase chain reaction (QF-PCR). The CMA result of amniotic fluid was arr[hg19]18p11.32q11.2(141,354-21,994,637)×3, 21q11.2q21.3(15,502,777- 29,700,071)×1; Refer to the CMA results, the fetal karyotype was described as 46,XX, +der(18)t(18;21) (q11.2; q22.1),-21. In order to verify the source of abnormal karyotype, the chromosomes of parents′ peripheral blood were detected. The father′s karyotype was 46,XY,t(18;21)(q11.2;q22.1), and the pregnant woman had normal karyotype. FISH results showed that the amniotic fluid cells had two normal 18 chromosomes, one normal 21 chromosome and one derived 18 chromosome, and that the father was a cross-translocation carrier of the 18p and 21p. To rule out the possibility that the fetus′s two normal chromosomes 18 were uniparental disomy, this study used the results of identification of maternal blood contamination, and the results of short tandem repeat(STR) test confirmed that only one of the two normal chromosome 18 came from the mother. Therefore, the application of multiple detection techniques in the diagnosis of rare separation methods such as adjacent-2 segregation can help doctors to provide accurate prenatal genetic counseling for those balanced translocation carriers, and to identify the types of chromosomal abnormalities in their fetuses. [ABSTRACT FROM AUTHOR]

Yuting Kang,1,* Qiujie Li,2,* Yao Yao,3 Chao Xu,2 Zhuoran Qiu,2 Wei Jia,1,3 Gang Li,1,3 Pengtao Wang1 1Ningxia Key Laboratory of Clinical and Pathogenic Microbiology, Institute of Medical Sciences, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, 750004, People’s Republic of China; 2College of Clinical Medicine, Ningxia Medical University, Yinchuan, Ningxia, 750004, People’s Republic of China; 3Center of Medical Laboratory, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, 750004, People’s Republic of China*These authors contributed equally to this workCorrespondence: Gang Li; Pengtao Wang, General Hospital of Ningxia Medical University, Technology Building 601, 804 Shengli Road, Yinchuan, People’s Republic of China, Tel +86-0951-6744462 ; +86-0951-6743330, Email gone.lee@163.com; wangpengtao2021@163.comPurpose: The objective of this study was to determine the clinical distribution, in vitro antifungal susceptibility and underlying resistance mechanisms of Aspergillus fumigatus (A. fumigatus) isolates from the General Hospital of Ningxia Medical University between November 2021 and May 2023.Methods: Antifungal susceptibility testing was performed using the Sensititre YeastOne YO10, and isolates with high minimal inhibitory concentrations (MICs) were further confirmed using the standard broth microdilution assays established by the Clinical and Laboratory Standards Institute (CLSI) M38-third edition. Whole-Genome Resequencing and RT-qPCR in azole-resistant A. fumigatus strains were performed to investigate the underlying resistance mechanisms.Results: Overall, a total of 276 A. fumigatus isolates were identified from various clinical departments, showing an increasing trend in the number of isolates over the past 3 years. Two azole-resistant A. fumigatus strains (0.72%) were observed, one of which showed overexpression of cyp51A, cyp51B, cdr1B, MDR1/2, artR, srbA, erg24A, and erg4B, but no cyp51A mutation. However, the other strain harbored two alterations in the cyp51A sequences (L98H/S297T). Therefore, we first described two azole-resistant clinical A. fumigatus strains in Ningxia, China, and reported one azole-resistant strain that has the L98H/S297T mutations in the cyp51A gene without any tandem repeat (TR) sequences in the promoter region.Conclusions: This study emphasizes the importance of enhancing attention and surveillance of azole-resistant A. fumigatus, particularly those with non-TR point mutations of cyp51A or non-cyp51A mutations, in order to gain a better understanding of their prevalence and spread in the region.Keywords: Aspergillus fumigatus, azole resistance, cyp51A mutation, tandem repeat sequences

In genetic studies of psychiatric disorders in the pre-genome-wide association study (GWAS) era, one of the most commonly studied loci is the serotonin transporter (SLC6A4) promoter polymorphism, a 43-base-pair insertion/deletion polymorphism in the promoter region (5-HTTLPR). The genetic association signals between 5-HTTLPR and psychiatric phenotypes, however, have been inconsistent across many studies. Since the polymorphism cannot be tested via available SNP arrays, we had previously proposed an efficient machine learning algorithm to predict the genotypes of 5-HTTLPR based on the genotypes of eight nearby SNPs, which requires access to individual-level genotype and phenotype data. To utilize the advantage of publicly available GWAS summary statistics obtained from studies with very large sample sizes, we develop a GWAS summary-statistics-based approach for testing the variable number of tandem repeat (VNTR) associations with various phenotypes. We first cross-verify the accuracy of the summary-statistics-based approach for 61 phenotypes in the UK Biobank. Since we observed a strong similarity between the predicted individual-level 5-HTTLPR genotype-based approach and the summary-statistics-based approach, we applied our method to the available neurobehavioral GWAS summary statistics data obtained from large-scale GWAS. We found no genome-wide significant evidence for association between 5-HTTLPR and any of the neurobehavioral traits. We did observe, however, genome-wide significant evidence for association between this locus and human adult height, BMI, and total cholesterol. Our summary-statistics-based approach provides a systematic way to examine the role of VNTRs and related types of genetic polymorphisms in disease risk and trait susceptibility of phenotypes for which large-scale GWAS summary statistics data are available.

Background and Objectives: The study aimed to investigate the distribution of genes encoding integrons, extended spectrum beta-lactamase (ESBL) in E. coli isolated from UTIs, as well as the genetic diversity among the isolates. Materials and Methods: E. coli isolates were recovered from the patients with UTI in Kerman Iran. Antibiotic susceptibility was done according to CLSI guidelines. The presence of ESBL genes and integrons was evaluated using PCR. PCR and sequencing were applied for the evaluation of cassette content of integrons. Genotyping of the isolates was performed by multiple-locus variable-number tandem repeat analysis (MLVA). Results: Imipenem was the most effective antibiotic, while the highest resistance was observed to streptomycin. In total 40.2% of isolates were ESBL producers. Of 69 integron-positive isolates, 59 only had class I integrons, 4 only had class II integrons and 6 had both types. The most common gene cassette found within class I integrons was dfrA17-aadA5 (n=27). The E. coli isolates were divided into 16 MLVA clusters. Conclusion: The current study demonstrated the simultaneous presence of class I integrons and ESBLs involved in the resistance of UPEC isolates to antibacterial agents. Our finding also revealed that the E. coli isolates belonged to diverse clones. [ABSTRACT FROM AUTHOR]

We report here on a novel pro-leukemogenic role of FMS-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD) that interferes with microRNAs (miRNAs) biogenesis in acute myeloid leukemia (AML) blasts. We showed that FLT3-ITD interferes with the canonical biogenesis of intron-hosted miRNAs such as miR-126, by phosphorylating SPRED1 protein and inhibiting the "gatekeeper" Exportin 5 (XPO5)/RAN-GTP complex that regulates the nucleus-to-cytoplasm transport of pre-miRNAs for completion of maturation into mature miRNAs. Of note, despite the blockage of "canonical" miRNA biogenesis, miR-155 remains upregulated in FLT3-ITD+ AML blasts, suggesting activation of alternative mechanisms of miRNA biogenesis that circumvent the XPO5/RAN-GTP blockage. MiR-155, a BIC-155 long noncoding (lnc) RNA-hosted oncogenic miRNA, has previously been implicated in FLT3-ITD+ AML blast hyperproliferation. We showed that FLT3-ITD upregulates miR-155 by inhibiting DDX3X, a protein implicated in the splicing of lncRNAs, via p-AKT. Inhibition of DDX3X increases unspliced BIC-155 that is then shuttled by NXF1 from the nucleus to the cytoplasm, where it is processed into mature miR-155 by cytoplasmic DROSHA, thereby bypassing the XPO5/RAN-GTP blockage via "non-canonical" mechanisms of miRNA biogenesis.

SummaryA rich set of tools have recently been developed for performing genome-wide genotyping of tandem repeats (TRs). However, standardized tools for downstream analysis of these results are lacking. To facilitate TR analysis applications, we present TRTools, a Python library and suite of command line tools for filtering, merging and quality control of TR genotype files. TRTools utilizes an internal harmonization module, making it compatible with outputs from a wide range of TR genotypers.Availability and implementationTRTools is freely available at https://github.com/gymreklab/TRTools. Detailed documentation is available at https://trtools.readthedocs.io.Supplementary informationSupplementary data are available at Bioinformatics online.

Triploidy is a genetic occurrence in which the chromosome count is 3n = 69 with a double (2n) chromosomal contribution to the conceptus from one parent. Such pregnancies are usually nonviable and are estimated to account for approximately 1% of recognized conceptions and 10% of recognized miscarriages. Majority opinion is that fetal losses due to triploidies are caused by the presence of 2 copies of paternal chromosomes. In this study, we present a digynic monoandric triploid miscarriage from a 32-year-old G7P1051 at approximately 13 weeks gestation, in which 2 copies of the maternal chromosomes are present in the fetus. This unusual phenomenon is supported by nonmolar placental histology, chromosomal microarray, and short tandem repeat assays, with the latter 2 being discussed in detail. Furthermore, this study includes discussion of recurrent miscarriage, recurrent triploidy, and long-term clinical follow-up of the patient. [ABSTRACT FROM AUTHOR]

Background and Objectives: The study aimed to investigate the distribution of genes encoding integrons, extended spectrum beta-lactamase (ESBL) in E. coli isolated from UTIs, as well as the genetic diversity among the isolates. Materials and Methods: E. coli isolates were recovered from the patients with UTI in Kerman Iran. Antibiotic susceptibility was done according to CLSI guidelines. The presence of ESBL genes and integrons was evaluated using PCR. PCR and sequencing were applied for the evaluation of cassette content of integrons. Genotyping of the isolates was performed by multiple-locus variable-number tandem repeat analysis (MLVA). Results: Imipenem was the most effective antibiotic, while the highest resistance was observed to streptomycin. In total 40.2% of isolates were ESBL producers. Of 69 integron-positive isolates, 59 only had class I integrons, 4 only had class II integrons and 6 had both types. The most common gene cassette found within class I integrons was dfrA17-aadA5 (n=27). The E. coli isolates were divided into 16 MLVA clusters. Conclusion: The current study demonstrated the simultaneous presence of class I integrons and ESBLs involved in the resistance of UPEC isolates to antibacterial agents. Our finding also revealed that the E. coli isolates belonged to diverse clones.

Centromeric variation has been linked to cancer and infertility, but centromere sequences contain multiple tandem repeats and can only be assembled manually from long error-prone reads. Here we describe the centroFlye algorithm for centromere assembly using long error-prone reads, and apply it to assemble human centromeres on chromosomes 6 and X. Our analyses reveal putative breakpoints in the manual reconstruction of the human X centromere, demonstrate that human X chromosome is partitioned into repeat subfamilies and provide initial insights into centromere evolution. We anticipate that centroFlye could be applied to automatically close remaining multimegabase gaps in the reference human genome.

Linking genomic variation to phenotypical traits remains a major challenge in evolutionary genetics. In this study, we use phylogenomic strategies to investigate a distinctive trait among mammals: the development of masculinizing ovotestes in female moles. By combining a chromosome-scale genome assembly of the Iberian mole, Talpa occidentalis, with transcriptomic, epigenetic, and chromatin interaction datasets, we identify rearrangements altering the regulatory landscape of genes with distinct gonadal expression patterns. These include a tandem triplication involving CYP17A1, a gene controlling androgen synthesis, and an intrachromosomal inversion involving the pro-testicular growth factor gene FGF9, which is heterochronically expressed in mole ovotestes. Transgenic mice with a knock-in mole CYP17A1 enhancer or overexpressing FGF9 showed phenotypes recapitulating mole sexual features. Our results highlight how integrative genomic approaches can reveal the phenotypic impact of noncoding sequence changes.

We are entering into an exciting era of genomics where truly complete, high-quality assemblies of human chromosomes are available end-to-end, or from 'telomere-to-telomere' (T2T). This technological advance offers a new opportunity to include endogenous human centromeric regions in high-resolution, sequence-based studies. These emerging reference maps are expected to reveal a new functional landscape in the human genome, where centromere proteins, transcriptional regulation, and spatial organization can be examined with base-level resolution across different stages of development and disease. Such studies will depend on innovative assembly methods of extremely long tandem repeats (ETRs), or satellite DNAs, paired with the development of new, orthogonal validation methods to ensure accuracy and completeness. This review reflects the progress in centromere genomics, credited by recent advancements in long-read sequencing and assembly methods. In doing so, I will discuss the challenges that remain and the promise for a new period of scientific discovery for satellite DNA biology and centromere function.

SUMMARY: Tandem DNA repeats can be sequenced with long-read technologies, but cannot be accurately deciphered due to the lack of computational tools taking high error rates of these technologies into account. Here we introduce Noise-Cancelling Repeat Finder (NCRF) to uncover putative tandem repeats of specified motifs in noisy long reads produced by Pacific Biosciences and Oxford Nanopore sequencers. Using simulations, we validated the use of NCRF to locate tandem repeats with motifs of various lengths and demonstrated its superior performance as compared to two alternative tools. Using real human whole-genome sequencing data, NCRF identified long arrays of the (AATGG)n repeat involved in heat shock stress response. AVAILABILITY AND IMPLEMENTATION: NCRF is implemented in C, supported by several python scripts, and is available in bioconda and at https://github.com/makovalab-psu/NoiseCancellingRepeatFinder. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

This randomized, open-label, phase 2b study (NCT01565668) evaluated the efficacy and safety of 2 dosing regimens of quizartinib monotherapy in patients with relapsed/refractory (R/R) FLT3-internal tandem duplication (ITD)-mutated acute myeloid leukemia (AML) who previously underwent transplant or 1 second-line salvage therapy. Patients (N = 76) were randomly assigned to 30- or 60-mg/day doses (escalations to 60 or 90 mg/day, respectively, permitted for lack/loss of response) of single-agent oral quizartinib dihydrochloride. Allelic frequency of at least 10% was defined as FLT3-ITD-mutated disease. Coprimary endpoints were composite complete remission (CRc) rates and incidence of QT interval corrected by Fridericia's formula (QTcF) of more than 480 ms (grade 2 or greater). Secondary endpoints included overall survival (OS), duration of CRc, bridge to transplant, and safety. CRc rates were 47% in both groups, similar to earlier reports with higher quizartinib doses. Incidence of QTcF above 480 ms was 11% and 17%, and QTcF above 500 ms was 5% and 3% in the 30- and 60-mg groups, respectively, which is less than earlier reports with higher doses of quizartinib. Median OS (20.9 and 27.3 weeks), duration of CRc (4.2 and 9.1 weeks), and bridge to transplant rates (32% and 42%) were higher in the 60-mg groups than in the 30-mg group. Dose escalation occurred in 61% and 14% of patients in the 30- and 60-mg groups, respectively. This high clinical activity of quizartinib at the evaluated doses is consistent with previous reports with an improved safety profile. Need to dose-escalate more than half of patients who received quizartinib 30 mg also supports further investigation of treatment with quizartinib 60 mg/day.

While mutations affecting protein-coding regions have been examined across many cancers, structural variants at the genome-wide level are still poorly defined. Through integrative deep whole-genome and -transcriptome analysis of 101 castration-resistant prostate cancer metastases (109X tumor/38X normal coverage), we identified structural variants altering critical regulators of tumorigenesis and progression not detectable by exome approaches. Notably, we observed amplification of an intergenic enhancer region 624 kb upstream of the androgen receptor (AR) in 81% of patients, correlating with increased AR expression. Tandem duplication hotspots also occur near MYC, in lncRNAs associated with post-translational MYC regulation. Classes of structural variations were linked to distinct DNA repair deficiencies, suggesting their etiology, including associations of CDK12 mutation with tandem duplications, TP53 inactivation with inverted rearrangements and chromothripsis, and BRCA2 inactivation with deletions. Together, these observations provide a comprehensive view of how structural variations affect critical regulators in metastatic prostate cancer.

Tau is a developmentally regulated axonal protein that stabilizes and bundles microtubules (MTs). Its hyperphosphorylation is thought to cause detachment from MTs and subsequent aggregation into fibrils implicated in Alzheimer's disease. It is unclear which tau residues are crucial for tau-MT interactions, where tau binds on MTs, and how it stabilizes them. We used cryo-electron microscopy to visualize different tau constructs on MTs and computational approaches to generate atomic models of tau-tubulin interactions. The conserved tubulin-binding repeats within tau adopt similar extended structures along the crest of the protofilament, stabilizing the interface between tubulin dimers. Our structures explain the effect of phosphorylation on MT affinity and lead to a model of tau repeats binding in tandem along protofilaments, tethering together tubulin dimers and stabilizing polymerization interfaces.

The human genome reference sequence remains incomplete owing to the challenge of assembling long tracts of near-identical tandem repeats in centromeres. We implemented a nanopore sequencing strategy to generate high-quality reads that span hundreds of kilobases of highly repetitive DNA in a human Y chromosome centromere. Combining these data with short-read variant validation, we assembled and characterized the centromeric region of a human Y chromosome.

Simple Summary: African swine fever (ASF) is one of the most important animal diseases affecting the domestic swine population globally. Whole-genome sequence analysis on the circulating African swine fever virus (ASFV) strains would provide valuable information in tracking the outbreaks of the disease. The aim of this study was to prepare a curated dataset of ASFV genome sequences and investigate genome-wide diversity of circulating ASFV strains. We prepared a curated dataset containing 123 high-quality ASFV genome sequences representing 10 genotypes collected from 28 countries between 1949 and 2020. Phylogenetic analysis based on whole-genome sequences provided high-resolution topology in genotyping ASFV isolates, which was supported by pairwise genome sequence similarity comparison. Wide distribution and high variation of tandem repeat sequences were found in ASFV genomes. Structural variation and highly variable poly G or poly C tracts were also identified. This study improved our understanding on the patterns of genetic variation of ASFV and facilitated future studies on ASFV molecular epidemiology. African swine fever (ASF) is a lethal contagious viral disease of domestic pigs and wild boars caused by the African swine fever virus (ASFV). The pandemic spread of ASF has had serious effects on the global pig industry. Virus genome sequencing and comparison play an important role in tracking the outbreaks of the disease and tracing the transmission of the virus. Although more than 140 ASFV genome sequences have been deposited in the public databases, the genome-wide diversity of ASFV remains unclear. Here we prepared a curated dataset of ASFV genome sequences by filtering genomes with sequencing errors as well as duplicated genomes. A total of 123 ASFV genome sequences were included in the dataset, representing 10 genotypes collected between 1949 and 2020. Phylogenetic analysis based on whole-genome sequences provided high-resolution topology in differentiating closely related ASFV isolates, and drew new clues in the classification of some ASFV isolates. Genome-wide diversity of ASFV genomes was explored by pairwise sequence similarity comparison and ORF distribution comparison. Tandem repeat sequences were found widely distributed and highly varied in ASFV genomes. Structural variation and highly variable poly G or poly C tracts also contributed to the genome diversity. This study expanded our knowledge on the patterns of genetic diversity and evolution of ASFV, and provided valuable information for diagnosis improvement and vaccine development. [ABSTRACT FROM AUTHOR]

Sarcomas characterized by BCOR gene alterations, are a distinct clinico-pathological group of high-grade tumors, that represent 5% of small round cell tumors without EWSR or FUS fusion. Diverse genetic alterations characterize this group, including BCOR-CCNB3 gene fusion being the most common alteration and less frequently internal tandem duplications (ITDs). We present a compelling case of a 3-year-old girl diagnosed with a high-grade nasoethmoidal sarcoma exhibiting BCOR -ITD. The diagnostic process illustrates the histological and immunophenotypic spectrum, requiring an extensive immunohistochemical panel and diverse molecular tests for accurate classification. Additionally, this case highlights the challenges in detecting BCOR -ITDs using different NGS panels, advocating for alternative molecular approaches. Our patient after 10 months since diagnosis is alive with progressive disease. This emphasizes the urgency for ongoing research to refine diagnostic methods and develop effective therapeutic strategies for these rare and aggressive tumors., Competing Interests: Declaration of Conflicting InterestsThe authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Reverse transcription has frequently been co-opted for cellular functions and in prokaryotes is associated with protection against viral infection, but the underlying mechanisms of defense are generally unknown. Here, we show that in the DRT2 defense system, the reverse transcriptase binds a neighboring pseudoknotted noncoding RNA. Upon bacteriophage infection, a template region of this RNA is reverse transcribed into an array of tandem repeats that reconstitute a promoter and open reading frame, allowing expression of a toxic repetitive protein and an abortive infection response. Biochemical reconstitution of this activity and cryo-electron microscopy provide a molecular basis for repeat synthesis. Gene synthesis from a noncoding RNA is a previously unknown mode of genetic regulation in prokaryotes.

Tandem repeat proteins (TRPs) are widely distributed and bind to a wide variety of ligands. DNA-binding TRPs such as zinc finger (ZNF) and transcription activator-like effector (TALE) play important roles in biology and biotechnology. In this study, we first conducted an extensive analysis of TRPs in public databases, and found that the enormous diversity of TRPs is largely unexplored. We then focused our efforts on identifying novel TRPs possessing DNA-binding capabilities. We established a protein language model for DNA-binding protein prediction (PLM-DBPPred), and predicted a large number of DNA-binding TRPs. A subset was then selected for experimental screening, leading to the identification of 11 novel DNA-binding TRPs, with six showing sequence specificity. Notably, members of the STAR (Short TALE-like Repeat proteins) family can be programmed to target specific 9 bp DNA sequences with high affinity. Leveraging this property, we generated artificial transcription factors using reprogrammed STAR proteins and achieved targeted activation of endogenous gene sets. Furthermore, the members of novel families such as MOON (Marine Organism-Originated DNA binding protein) and pTERF (prokaryotic mTERF-like protein) exhibit unique features and distinct DNA-binding characteristics, revealing interesting biological clues. Our study expands the diversity of DNA-binding TRPs, and demonstrates that a systematic approach greatly enhances the discovery of new biological insights and tools., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

FMS-like tyrosine kinase 3 (FLT3) mutations occur in approximately one third of acute myeloid leukemia (AML) patients. FLT3-Internal tandem duplication (FLT3-ITD) mutations are the most common FLT3 mutations and are associated with a poor prognosis. Gilteritinib is a FLT3 inhibitor that is US FDA approved for treating adult patients with relapsed/refractory AML and a FLT3 mutation. While gilteritinib monotherapy has improved patient outcome, few patients achieve durable responses. Combining gilteritinib with venetoclax (VEN) appears to make further improvements, though early results suggest that patients with prior exposure to VEN fair much worse than those without prior exposure. MRX-2843 is a promising inhibitor of FLT3 and MERTK. We recently demonstrated that MRX-2843 is equally potent as gilteritinib in FLT3-ITD AML cell lines in vitro and primary patient samples ex vivo. In this study, we investigated the combination of VEN and MRX-2843 against FLT3-ITD AML cells. We found that VEN synergistically enhances cell death induced by MRX-2843 in FLT3-mutated AML cell lines and primary patient samples. Importantly, we found that VEN synergistically enhances cell death induced by MRX-2843 in FLT3-ITD AML cells with acquired resistance to cytarabine (AraC) or VEN+AraC. VEN and MRX-2843 significantly reduce colony-forming capacity of FLT3-ITD primary AML cells. Mechanistic studies show that MRX-2843 decreases Mcl-1 and c-Myc protein levels via transcriptional regulation and combined MRX-2843 and VEN significantly decreases oxidative phosphorylation in FLT3-ITD AML cells. Our findings highlight a promising combination therapy against FLT3-ITD AML, supporting further in vitro and in vivo testing., Competing Interests: Declaration of Competing Interest The authors have no relevant financial or non-financial interests to disclose., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background & Aims: Considering the lack of knowledge regarding the influence of the variable number of repeats of 27 pb in intron 4 (4b/4a VNTR - rs61722009) of the endothelial nitric oxide synthase (eNOS) on the drug response, we assessed the influence of this polymorphism for the risk of upper gastrointestinal bleeding (UGIB). Methods: A case-control study, including 200 cases and 706 controls, was conducted in a Brazilian hospital complex. Cases were participants with UGIB diagnosis. Controls were participants admitted to surgical procedures not related to gastrointestinal problems. The 4b/4a VNTR was determined through polymerase chain reaction followed by fragment analysis. Conditional logistic regression models were designed. The additive interaction between the presence of the 4b/4a VNTR variant and the use of low-dose aspirin (LDA) and nonsteroidal anti-inflammatory drugs (NSAIDs) was calculated by fitting the Cox regression model through the parameters of Synergism index (S) and Relative Excess Risk Due To Interaction (RERI). Results: The presence of the 4b/4a VNTR variant did not increase the risk of UGIB: carriers of the 4a/4a genotype (OR=0.37, 95%CI: 0.09-1.45) and of the variant allele "4a" (OR=0.91, 95%CI: 0.55-1.51). The risk of UGIB in LDA users carriers of the wild genotype (OR=4.96, 95%CI: 2.04- 2.06) and the variant allele "4a" (OR=3.49, 95%CI: 1.18-10.38) is similar, as well as for NSAID users carriers of the wild genotype (OR=5.73, 95%CI: 2.61-12.60) and variant allele "4a" (OR=5.51, 95%CI: 1.42-15.82). No additive interaction was identified between the presence of the genetic variant and the use of LDA [RERI: -1.44 (95%CI: -6.02-3.14; S: 0.63 (95%CI: -1.97-1.15)] and NSAIDs [RERI: -0.13 (95%CI: -6.79-6.53; S: 0.97 (95%CI: -0.23-4.19)] on the UGIB risk. Conclusion: Our data suggests that there is no increase in the magnitude of UGIB risk in LDA and NSAIDs users' carrying the variant allele "4a". [ABSTRACT FROM AUTHOR]

Aberrant cytokine signaling initiated from mutant receptor tyrosine kinases (RTKs) provides critical growth and survival signals in high risk acute myeloid leukemia (AML). Inhibitors to FLT3 have already been tested in clinical trials, however, drug resistance limits clinical efficacy. Mutant receptor tyrosine kinases are mislocalized in the endoplasmic reticulum (ER) of AML and play an important role in the non-canonical activation of signal transducer and activator of transcription 5 (STAT5). Here, we have tested a potent new drug called imipramine blue (IB), which is a chimeric molecule with a dual mechanism of action. At 200-300 nM concentrations, IB is a potent inhibitor of STAT5 through liberation of endogenous phosphatase activity following NADPH oxidase (NOX) inhibition. However, at 75-150 nM concentrations, IB was highly effective at killing mutant FLT3-driven AML cells through a similar mechanism as thapsigargin (TG), involving increased cytosolic calcium. IB also potently inhibited survival of primary human FLT3/ITD+ AML cells compared to FLT3/ITDneg cells and spared normal umbilical cord blood cells. Therefore, IB functions through a mechanism involving vulnerability to dysregulated calcium metabolism and the combination of fusing a lipophilic amine to a NOX inhibiting dye shows promise for further pre-clinical development for targeting high risk AML.

One common hypothesis to explain the impacts of tandem duplications is that whole gene duplications commonly produce additive changes in gene expression due to copy number changes. Here, we use genome wide RNA-seq data from a population sample of Drosophila yakuba to test this 'gene dosage' hypothesis. We observe little evidence of expression changes in response to whole transcript duplication capturing 5' and 3' UTRs. Among whole gene duplications, we observe evidence that dosage sharing across copies is likely to be common. The lack of expression changes after whole gene duplication suggests that the majority of genes are subject to tight regulatory control and therefore not sensitive to changes in gene copy number. Rather, we observe changes in expression level due to both shuffling of regulatory elements and the creation of chimeric structures via tandem duplication. Additionally, we observe 30 de novo gene structures arising from tandem duplications, 23 of which form with expression in the testes. Thus, the value of tandem duplications is likely to be more intricate than simple changes in gene dosage. The common regulatory effects from chimeric gene formation after tandem duplication may explain their contribution to genome evolution.

Gene clusters of recently duplicated genes are hotbeds for evolutionary change. However, our understanding of how mutational mechanisms and evolutionary forces shape the structural and functional evolution of these clusters is hindered by the high sequence identity among the copies, which typically results in their inaccurate representation in genome assemblies. The presumed testis-specific, chimeric gene Sdic originated, and tandemly expanded in Drosophila melanogaster, contributing to increased male-male competition. Using various types of massively parallel sequencing data, we studied the organization, sequence evolution, and functional attributes of the different Sdic copies. By leveraging long-read sequencing data, we uncovered both copy number and order differences from the currently accepted annotation for the Sdic region. Despite evidence for pervasive gene conversion affecting the Sdic copies, we also detected signatures of two episodes of diversifying selection, which have contributed to the evolution of a variety of C-termini and miRNA binding site compositions. Expression analyses involving RNA-seq datasets from 59 different biological conditions revealed distinctive expression breadths among the copies, with three copies being transcribed in females, opening the possibility to a sexually antagonistic effect. Phenotypic assays using Sdic knock-out strains indicated that should this antagonistic effect exist, it does not compromise female fertility. Our results strongly suggest that the genome consolidation of the Sdic gene cluster is more the result of a quick exploration of different paths of molecular tinkering by different copies than a mere dosage increase, which could be a recurrent evolutionary outcome in the presence of persistent sexual selection.

Partial tandem duplication of MLL (MLL-PTD) characterizes acute myeloid leukemia (AML) patients often with a poor prognosis. To understand the order of occurrence of MLL-PTD in relation to other major AML mutations and to identify novel mutations that may be present in this unique AML molecular subtype, exome and targeted sequencing was performed on 85 MLL-PTD AML samples using HiSeq-2000. Genes involved in the cohesin complex (STAG2), a splicing factor (U2AF1) and a poorly studied gene, MGA were recurrently mutated, whereas NPM1, one of the most frequently mutated AML gene, was not mutated in MLL-PTD patients. Interestingly, clonality analysis suggests that IDH2/1, DNMT3A, U2AF1 and TET2 mutations are clonal and occur early, and MLL-PTD likely arises after these initial mutations. Conversely, proliferative mutations (FLT3, RAS), typically appear later, are largely subclonal and tend to be unstable. This study provides important insights for understanding the relative importance of different mutations for defining a targeted therapeutic strategy for MLL-PTD AML patients.

Highly repetitive regions have historically posed a challenge when investigating sequence variation and content. High-throughput sequencing has enabled researchers to use whole-genome shotgun sequencing to estimate the abundance of repetitive sequence, and these methodologies have been recently applied to centromeres. Previous research has investigated variation in centromere repeats across eukaryotes, positing that the highest abundance tandem repeat in a genome is often the centromeric repeat. To test this assumption, we used shotgun sequencing and a bioinformatic pipeline to identify common tandem repeats across a number of grass species. We find that de novo assembly and subsequent abundance ranking of repeats can successfully identify tandem repeats with homology to known tandem repeats. Fluorescent in-situ hybridization shows that de novo assembly and ranking of repeats from non-model taxa identifies chromosome domains rich in tandem repeats both near pericentromeres and elsewhere in the genome.

BackgroundReceptor-like kinases (RLKs) belong to a large protein family with over 600 members in Arabidopsis and over 1000 in rice. Among RLKs, the lectin receptor-like kinases (LecRLKs) possess a characteristic extracellular carbohydrate-binding lectin domain and play important roles in plant development and innate immunity. There are 75 and 173 LecRLKs in Arabidopsis and rice, respectively. However, little is known about LecRLKs in perennial woody plants.ResultsHere we report the genome-wide analysis of classification, domain architecture and expression of LecRLKs in the perennial woody model plant Populus. We found that the LecRLK family has expanded in Populus to a total of 231, including 180 G-type, 50 L-type and 1 C-type LecRLKs. Expansion of the Populus LecRLKs (PtLecRLKs) occurred partially through tandem duplication. Based on domain architecture and orientation features, we classified PtLecRLKs into eight different classes. RNA-seq-based transcriptomics analysis revealed diverse expression patterns of PtLecRLK genes among leaves, stems, roots, buds and reproductive tissues and organs.ConclusionsThis study offers a comprehensive view of LecRLKs in the perennial woody model plant Populus and provides a foundation for functional characterization of this important family of receptor-like kinases.

Protein folding and unfolding experiments are interpreted under the assumption of microscopic reversibility, that is, that at equilibrium one process is the reverse of the other. Single-domain proteins illustrate the validity of such an interpretation, although reversibility does not necessarily hold under the different conditions typically used for folding and unfolding experiments. In fact, more complex proteins, which often exhibit irreversible unfolding, are generally considered not amenable to folding kinetics studies. Here, the X11 PDZ1-PDZ2 tandem repeat allows us to reveal the different folding and unfolding pathways at play under different experimental conditions, thus reconciling the apparent contradiction between theory and experiment., (© 2024 The Author(s). Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.)

The adverse prognosis of internal tandem duplication in the FMS-like tyrosine kinase 3 gene(s) (FLT3-ITD) in patients with acute myelogenous leukemia (AML) may depend on allelic burden. We compared postremission treatment with chemotherapy and hematopoietic stem cell transplantation (HSCT) in 169 FLT3-ITDmut intermediate cytogenetic risk AML patients with allelic ratio evaluable at diagnosis who achieved first complete remission (CR1) with induction therapy. To minimize selection bias, the analysis was limited to patients who remained in CR1 for at least 4 months (median time to HSCT) after achieving CR1, and propensity score matching was implemented. Sensitivity analysis including patients who remained in CR1 for at least 3 months was applied as well. HSCT in CR1 was associated with longer relapse-free survival (RFS) and overall survival (OS), with 3-year estimated rates of 18% and 24%, respectively (P

The assembly of a reference genome sequence of bread wheat is challenging due to its specific features such as the genome size of 17 Gbp, polyploid nature and prevalence of repetitive sequences. BAC-by-BAC sequencing based on chromosomal physical maps, adopted by the International Wheat Genome Sequencing Consortium as the key strategy, reduces problems caused by the genome complexity and polyploidy, but the repeat content still hampers the sequence assembly. Availability of a high-resolution genomic map to guide sequence scaffolding and validate physical map and sequence assemblies would be highly beneficial to obtaining an accurate and complete genome sequence. Here, we chose the short arm of chromosome 7D (7DS) as a model to demonstrate for the first time that it is possible to couple chromosome flow sorting with genome mapping in nanochannel arrays and create a de novo genome map of a wheat chromosome. We constructed a high-resolution chromosome map composed of 371 contigs with an N50 of 1.3 Mb. Long DNA molecules achieved by our approach facilitated chromosome-scale analysis of repetitive sequences and revealed a ~800-kb array of tandem repeats intractable to current DNA sequencing technologies. Anchoring 7DS sequence assemblies obtained by clone-by-clone sequencing to the 7DS genome map provided a valuable tool to improve the BAC-contig physical map and validate sequence assembly on a chromosome-arm scale. Our results indicate that creating genome maps for the whole wheat genome in a chromosome-by-chromosome manner is feasible and that they will be an affordable tool to support the production of improved pseudomolecules.

UnlabelledCytogenetically normal acute myeloid leukemia (CN-AML) represents nearly 50% of human AML. Co-occurring mutations in the de novo DNA methyltransferase DNMT3A and the FMS related tyrosine kinase 3 (FLT3) are common in CN-AML and confer a poorer prognosis. We demonstrate that mice with Flt3-internal tandem duplication (Flt3(ITD)) and inducible deletion of Dnmt3a spontaneously develop a rapidly lethal, completely penetrant, and transplantable AML of normal karyotype. AML cells retain a single Dnmt3a floxed allele, revealing the oncogenic potential of Dnmt3a haploinsufficiency. FLT3(ITD)/DNMT3A-mutant primary human and murine AML exhibit a similar pattern of global DNA methylation associated with changes in the expression of nearby genes. In the murine model, rescuing Dnmt3a expression was accompanied by DNA remethylation and loss of clonogenic potential, suggesting that Dnmt3a-mutant oncogenic effects are reversible. Dissection of the cellular architecture of the AML model using single-cell assays, including single-cell RNA sequencing, identified clonogenic subpopulations that express genes sensitive to the methylation of nearby genomic loci and responsive to DNMT3A levels. Thus, Dnmt3a haploinsufficiency transforms Flt3(ITD) myeloproliferative disease by modulating methylation-sensitive gene expression within a clonogenic AML subpopulation.SignificanceDNMT3A haploinsufficiency results in reversible epigenetic alterations that transform FLT3(ITD)-mutant myeloproliferative neoplasm into AML. Cancer Discov; 6(5); 501-15. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 461.

Oncogenic KRAS mutations introduce discrete amino acid substitutions that reduce intrinsic Ras GTPase activity and confer resistance to GTPase-activating proteins (GAPs). Here we discover a partial duplication of the switch 2 domain of K-Ras encoding a tandem repeat of amino acids G60_A66dup in a child with an atypical myeloproliferative neoplasm. K-Ras proteins containing this tandem duplication or a similar five amino acid E62_A66dup mutation identified in lung and colon cancers transform the growth of primary myeloid progenitors and of Ba/F3 cells. Recombinant K-Ras(G60_A66dup) and K-Ras(E62_A66dup) proteins display reduced intrinsic GTP hydrolysis rates, accumulate in the GTP-bound conformation and are resistant to GAP-mediated GTP hydrolysis. Remarkably, K-Ras proteins with switch 2 insertions are impaired for PI3 kinase binding and Akt activation, and are hypersensitive to MEK inhibition. These studies illuminate a new class of oncogenic KRAS mutations and reveal unexpected plasticity in oncogenic Ras proteins that has diagnostic and therapeutic implications.

Epstein-Barr virus (EBV) nuclear antigen 2 (EBNA2) plays an important role in driving immortalization of EBV-infected B cells through regulating the expression of many viral and cellular genes. We report a structural study of the tumor suppressor BS69/ZMYND11 C-terminal region, comprised of tandem coiled-coil-MYND domains (BS69CC-MYND), in complex with an EBNA2 peptide containing a PXLXP motif. The coiled-coil domain of BS69 self-associates to bring two separate MYND domains in close proximity, thereby enhancing the BS69 MYND-EBNA2 interaction. ITC analysis of BS69CC-MYND with a C-terminal fragment of EBNA2 further suggests that the BS69CC-MYND homodimer synergistically binds to the two EBNA2 PXLXP motifs that are respectively located in the conserved regions CR7 and CR8. Furthermore, we showed that EBNA2 interacts with BS69 and down-regulates its expression at both mRNA and protein levels in EBV-infected B cells. Ectopic BS69CC-MYND is recruited to viral target promoters through interactions with EBNA2, inhibits EBNA2-mediated transcription activation, and impairs proliferation of lymphoblastoid cell lines (LCLs). Substitution of critical residues in the MYND domain impairs the BS69-EBNA2 interaction and abolishes the BS69 inhibition of the EBNA2-mediated transactivation and LCL proliferation. This study identifies the BS69 C-terminal domains as an inhibitor of EBNA2, which may have important implications in development of novel therapeutic strategies against EBV infection.

Importance: Persistence of FLT3 internal tandem duplication (ITD) in adults with acute myeloid leukemia (AML) in first complete remission (CR) prior to allogeneic hematopoietic cell transplant (HCT) is associated with increased relapse and death after transplant, but the association between the level of measurable residual disease (MRD) detected and clinical outcome is unknown., Objective: To examine the association between pre-allogeneic HCT MRD level with relapse and death posttransplant in adults with AML in first CR., Design, Setting, and Participants: In this cohort study, DNA sequencing was performed on first CR blood from patients with FLT3-ITD AML transplanted from March 2013 to February 2019. Clinical follow-up was through May 2022. Data were analyzed from October 2022 to December 2023., Exposure: Centralized DNA sequencing for FLT3-ITD in pre-allogeneic HCT first CR blood using a commercially available kit., Main Outcomes and Measures: The primary outcomes were overall survival and cumulative incidence of relapse, with non-relapse-associated mortality as a competing risk post-allogeneic HCT. Kaplan-Meier estimations (log-rank tests), Cox proportional hazards models, and Fine-Gray models were used to estimate the end points., Results: Of 537 included patients with FLT3-ITD AML from the Pre-MEASURE study, 296 (55.1%) were female, and the median (IQR) age was 55.6 (42.9-64.1) years. Using the variant allele fraction (VAF) threshold of 0.01% or greater for MRD positivity, the results closely aligned with those previously reported. With no VAF threshold applied (VAF greater than 0%), 263 FLT3-ITD variants (median [range] VAF, 0.005% [0.0002%-44%]), and 177 patients (33.0%) with positive findings were identified. Multivariable analyses showed that residual FLT3-ITD was the variable most associated with relapse and overall survival, with a dose-dependent correlation. Patients receiving reduced-intensity conditioning without melphalan or nonmyeloablative conditioning had increased risk of relapse and death at any given level of MRD compared with those receiving reduced-intensity conditioning with melphalan or myeloablative conditioning., Conclusions and Relevance: This study provides generalizable and clinically applicable evidence that the detection of residual FLT3-ITD in the blood of adults in first CR from AML prior to allogeneic HCT is associated with an increased risk of relapse and death, particularly for those with a VAF of 0.01% or greater. While transplant conditioning intensification, an intervention not available to all, may help mitigate some of this risk, alternative approaches will be necessary for this high-risk population of patients who are underserved by the current standard of care.

Cryptosporidiosis is an infectious enteric disease caused by species (some of them zoonotic) of the genus Cryptosporidium that in many countries are under surveillance. Typing assays critical to the surveillance of cryptosporidiosis typically involve characterization of Cryptosporidium glycoprotein 60 genes (gp60). Here, we characterized the gp60 of Cryptosporidium suis from two samples-a human and a porcine faecal sample-based on which a preliminary typing scheme was developed. A conspicuous feature of the C. suis gp60 was a novel type of tandem repeats located in the 5' end of the gene and that took up 777/1635 bp (48%) of the gene. The C. suis gp60 lacked the classical poly-serine repeats (TCA/TCG/TCT), which is usually subject to major genetic variation, and the length of the tandem repeat made a typing assay incorporating this region based on Sanger sequencing practically unfeasible. We therefore designed a typing assay based on the post-repeat region only and applied it to C. suis-positive samples from suid hosts from Norway, Denmark, and Spain. We were able to distinguish three different subtypes; XXVa-1, XXVa-2, and XXVa-3. Subtype XXVa-1 had a wider geographic distribution than the other subtypes and was also observed in the human sample. We think that the present data will inform future strategies to develop a C. suis typing assay that could be even more informative by including a greater part of the gene, including the tandem repeat region, e.g., by the use of long-read next-generation sequencing., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Central nervous system tumor with BCOR internal tandem duplication (CNS tumor with BCOR-ITD) constitutes a molecularly distinct entity, characterized by internal tandem duplication within exon 15 of the BCOR transcriptional co-repressor gene (BCOR-ITD). The current study aimed to elucidate the clinical, pathological, and molecular attributes of CNS tumors with BCOR-ITD and explore their putative cellular origin. This study cohort comprised four pediatric cases, aged 23 months to 13 years at initial presentation. Magnetic resonance imaging revealed large, well-circumscribed intra-CNS masses localized heterogeneously throughout the CNS. Microscopically, tumors were composed of spindle to ovoid cells, exhibiting perivascular pseudorosettes and palisading necrosis, but lacking microvascular proliferation. Immunohistochemical staining showed diffuse tumor cell expression of BCOR, CD56, CD99, vimentin, and the stem cell markers PAX6, SOX2, CD133 and Nestin, alongside focal positivity for Olig-2, S100, SOX10, Syn and NeuN. Molecularly, all cases harbored BCOR-ITDs ranging from 87 to 119 base pairs in length, including one case with two distinct ITDs. Notably, the ITDs were interrupted by unique 1-3 bp insertions in all cases. In summary, CNS tumors with BCOR-ITD exhibit characteristic clinical, pathological, and molecular features detectable through BCOR immunohistochemistry and confirmatory molecular analyses. Their expression of stem cell markers raises the possibility of an origin from neuroepithelial stem cells rather than representing true embryonal neoplasms., Competing Interests: Declaration of Competing Interest The authors state that there are no conflicts of interest to disclose., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Opinion Statement: The internal tandem duplication (ITD) mutation of the FMS-like receptor tyrosine kinase 3 (FLT3-ITD) is the most common mutation observed in approximately 30% of acute myeloid leukemia (AML) patients. It represents poor prognosis due to continuous activation of downstream growth-promoting signaling pathways such as STAT5 and PI3K/AKT. Hence, FLT3 is considered an attractive druggable target; selective small FLT3 inhibitors (FLT3Is), such as midostaurin and quizartinib, have been clinically approved. However, patients possess generally poor remission rates and acquired resistance when FLT3I used alone. Various factors in patients could cause these adverse effects including altered epigenetic regulation, causing mainly abnormal gene expression patterns. Epigenetic modifications are required for hematopoietic stem cell (HSC) self-renewal and differentiation; however, critical driver mutations have been identified in genes controlling DNA methylation (such as DNMT3A, TET2, IDH1/2). These regulators cause leukemia pathogenesis and affect disease diagnosis and prognosis when they co-occur with FLT3-ITD mutation. Therefore, understanding the role of different epigenetic alterations in FLT3-ITD AML pathogenesis and how they modulate FLT3I's activity is important to rationalize combinational treatment approaches including FLT3Is and modulators of methylation regulators or pathways. Data from ongoing pre-clinical and clinical studies will further precisely define the potential use of epigenetic therapy together with FLT3Is especially after characterized patients' mutational status in terms of FLT3 and DNA methlome regulators., (© 2024. The Author(s).)

A case of 69, XXX homozygous triploid was reported. This is a 24-year-old, 21-week-pregnant woman with adverse pregnancy and childbirth history. The serum prenatal screening showed the high risk of 18- trisomy syndrome. The four -dimensional color ultrasound suggested some abnormality, and the non -invasive prenatal testing (NIPT) suggested the high risk of sex chromosome aneuploidy. After that, the in-depth genetic testing and analysis by the short tandem repeat (STR) fragment analysis and fetal chromosome karyotype analysis were done with the informed consent. In this case, the fetal chromosome karyotype was 69, XXX, and the STR fragment analysis showed fetal triploid. The possible cause of this case was double female fertilization, since no maternal contamination was found in amniotic fluid samples. Therefore, this fetal case was considered as the homozygous triploid. The pregnant woman chose to terminate the pregnancy after genetic counseling. In conclusion, the prenatal diagnosis combining with STR fragment analysis can not only eliminate the influence of maternal contamination, but also find chromosome number abnormalities. [ABSTRACT FROM AUTHOR]

Live cell RNA imaging using genetically encoded fluorescent labels is an important tool for monitoring RNA activities. A recently reported RNA aptamer-fluorogen system, the Spinach, in which an RNA aptamer binds and induces the fluorescence of a GFP-like 3,5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI) ligand, can be readily tagged to the RNA of interest. Although the aptamer-fluorogen system is sufficient for imaging highly abundant non-coding RNAs (tRNAs, rRNAs, etc.), it performs poorly for mRNA imaging due to low brightness. In addition, whether the aptamer-fluorogen system may perturb the native RNA characteristics has not been systematically characterized at the levels of RNA transcription, translation and degradation. To increase the brightness of these aptamer-fluorogen systems, we constructed and tested tandem arrays containing multiple Spinach aptamers (8-64 aptamer repeats). Such arrays enhanced the brightness of the tagged mRNA molecules by up to ~17 fold in living cells. Strong laser excitation with pulsed illumination further increased the imaging sensitivity of Spinach array-tagged RNAs. Moreover, transcriptional fusion to the Spinach array did not affect mRNA transcription, translation or degradation, indicating that aptamer arrays might be a generalizable labeling method for high-performance and low-perturbation live cell RNA imaging.

The human genome is arguably the most complete mammalian reference assembly, yet more than 160 euchromatic gaps remain and aspects of its structural variation remain poorly understood ten years after its completion. To identify missing sequence and genetic variation, here we sequence and analyse a haploid human genome (CHM1) using single-molecule, real-time DNA sequencing. We close or extend 55% of the remaining interstitial gaps in the human GRCh37 reference genome--78% of which carried long runs of degenerate short tandem repeats, often several kilobases in length, embedded within (G+C)-rich genomic regions. We resolve the complete sequence of 26,079 euchromatic structural variants at the base-pair level, including inversions, complex insertions and long tracts of tandem repeats. Most have not been previously reported, with the greatest increases in sensitivity occurring for events less than 5 kilobases in size. Compared to the human reference, we find a significant insertional bias (3:1) in regions corresponding to complex insertions and long short tandem repeats. Our results suggest a greater complexity of the human genome in the form of variation of longer and more complex repetitive DNA that can now be largely resolved with the application of this longer-read sequencing technology.

African swine fever (ASF) is a lethal contagious viral disease of domestic pigs and wild boars caused by the African swine fever virus (ASFV). The pandemic spread of ASF has had serious effects on the global pig industry. Virus genome sequencing and comparison play an important role in tracking the outbreaks of the disease and tracing the transmission of the virus. Although more than 140 ASFV genome sequences have been deposited in the public databases, the genome-wide diversity of ASFV remains unclear. Here we prepared a curated dataset of ASFV genome sequences by filtering genomes with sequencing errors as well as duplicated genomes. A total of 123 ASFV genome sequences were included in the dataset, representing 10 genotypes collected between 1949 and 2020. Phylogenetic analysis based on whole-genome sequences provided high-resolution topology in differentiating closely related ASFV isolates, and drew new clues in the classification of some ASFV isolates. Genome-wide diversity of ASFV genomes was explored by pairwise sequence similarity comparison and ORF distribution comparison. Tandem repeat sequences were found widely distributed and highly varied in ASFV genomes. Structural variation and highly variable poly G or poly C tracts also contributed to the genome diversity. This study expanded our knowledge on the patterns of genetic diversity and evolution of ASFV, and provided valuable information for diagnosis improvement and vaccine development.

We have used whole genome paired-end Illumina sequence data to identify tandem duplications in 20 isofemale lines of Drosophila yakuba and 20 isofemale lines of D. simulans and performed genome wide validation with PacBio long molecule sequencing. We identify 1,415 tandem duplications that are segregating in D. yakuba as well as 975 duplications in D. simulans, indicating greater variation in D. yakuba. Additionally, we observe high rates of secondary deletions at duplicated sites, with 8% of duplicated sites in D. simulans and 17% of sites in D. yakuba modified with deletions. These secondary deletions are consistent with the action of the large loop mismatch repair system acting to remove polymorphic tandem duplication, resulting in rapid dynamics of gain and loss in duplicated alleles and a richer substrate of genetic novelty than has been previously reported. Most duplications are present in only single strains, suggesting that deleterious impacts are common. Drosophila simulans shows larger numbers of whole gene duplications in comparison to larger proportions of gene fragments in D. yakuba. Drosophila simulans displays an excess of high-frequency variants on the X chromosome, consistent with adaptive evolution through duplications on the D. simulans X or demographic forces driving duplicates to high frequency. We identify 78 chimeric genes in D. yakuba and 38 chimeric genes in D. simulans, as well as 143 cases of recruited noncoding sequence in D. yakuba and 96 in D. simulans, in agreement with rates of chimeric gene origination in D. melanogaster. Together, these results suggest that tandem duplications often result in complex variation beyond whole gene duplications that offers a rich substrate of standing variation that is likely to contribute both to detrimental phenotypes and disease, as well as to adaptive evolutionary change.