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Nuclear factors TOX and TOX2 upregulate TIM3 expression and lead to T-cell exhaustion in malignancies. Here, we demonstrate two distinct TIM3 expression patterns (high & low) with high TOX and TOX2 levels in T-cell acute lymphoblastic leukemia (T-ALL) specimens and cell lines. However, the mechanisms regulated by TOX and TIM3 signaling in leukemogenesis are unclear. We found that TOX and TOX2 proteins each directly upregulated HAVCR2 transcription, while the cellular localization of TOX2 was different in Jurkat and MOLT3 cells (nucleus) and lymphoblastic cell T2 and normal T cells (cytoplasm). Nuclear TOX and TOX2 formed a protein complex and repressed HAVCR2 promoter activity by recruiting transcriptional corepressor LCOR and deacetylase HDAC3. The nuclear-cytosol translocation of TOX2 was deacetylation-dependent and cooperatively mediated by deacetylase Sirt1 and kinase TBK1. Radiation damage induced TOX2 nuclear translocation and decreased Sirt1, TIM3, and caspase 1 expression in normal T cells. Accordingly, knockdown of TOX, TOX2 or LCOR; HDAC3 inhibition; or TIM3 overexpression induced Jurkat cell apoptosis in vitro and slow growth in vivo. Thus, our findings demonstrate a novel regulatory mechanism involving TOX-TOX2 and the TIM3 pathway in the leukemogenesis of T-ALL., (© 2024. The Author(s).)

During DNA replication, core histones that form nucleosomes on template strands are evicted and associate with newly synthesized strands to reform nucleosomes. Mcm2, a subunit of the Mcm2-7 complex, which is a core component of the replicative helicase, interacts with histones in the amino-terminal region (Mcm2N) and is involved in the parental histone recycling to lagging strands. Herein, the interaction of Mcm2N with histones was biochemically analyzed to reveal the molecular mechanisms underlying histone recycling by Mcm2N. With the addition of Mcm2N, a histone hexamer, comprising an H3-H4 tetramer and an H2A-H2B dimer, was excised from the histone octamer to form a complex with Mcm2N. The histone hexamer, but not H3-H4 tetramer was released from Mcm2N in the presence of Nap1, a histone chaperone. FACT, another histone chaperone, stabilized Mcm2N-histone hexamer complex to protect from Nap1-dependent dissociation. This study indicates cooperative histone transfer via Mcm2N and histone chaperones., (© The Author(s) 2024. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.)

Saccharomyces cerevisiae HMO1 is an architectural nuclear DNA-binding protein that stimulates the activity of some remodelers and regulates the transcription of ribosomal protein genes, often binding to a DNA motif called IFHL. However, the molecular mechanism dictating this sequence specificity is unclear. Our circular dichroism spectroscopy studies show that the HMO1:DNA complex forms without noticeable changes in the structure of DNA and HMO1. Molecular modeling/molecular dynamics studies of the DNA complex with HMO1 Box B reveal two extended sites at the N-termini of helices I and II of Box B that are involved in the formation of the complex and stabilize the DNA bend induced by intercalation of the F114 side chain between base pairs. A comparison of the affinities of HMO1 for 24 bp DNA fragments containing either randomized or IFHL sequences reveals a twofold increase in the stability of the complex in the latter case, which may explain the selectivity in the recognition of the IFHL-containing promoter regions.

In this issue of Molecular Cell, Engeholm et al. 1 present cryo-EM structures of the chromatin remodeler Chd1 bound to a hexasome-nucleosome complex, an intermediate state during transcription either with or without FACT to restore the missing H2A-H2B dimer. These two binding modes reveal how Chd1 and FACT cooperate in nucleosome re-establishment during transcription., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

To maintain the nucleosome organization of transcribed genes, ATP-dependent chromatin remodelers collaborate with histone chaperones. Here, we show that at the 5' ends of yeast genes, RNA polymerase II (RNAPII) generates hexasomes that occur directly adjacent to nucleosomes. The resulting hexasome-nucleosome complexes are then resolved by Chd1. We present two cryoelectron microscopy (cryo-EM) structures of Chd1 bound to a hexasome-nucleosome complex before and after restoration of the missing inner H2A/H2B dimer by FACT. Chd1 uniquely interacts with the complex, positioning its ATPase domain to shift the hexasome away from the nucleosome. In the absence of the inner H2A/H2B dimer, its DNA-binding domain (DBD) packs against the ATPase domain, suggesting an inhibited state. Restoration of the dimer by FACT triggers a rearrangement that displaces the DBD and stimulates Chd1 remodeling. Our results demonstrate how chromatin remodelers interact with a complex nucleosome assembly and suggest how Chd1 and FACT jointly support transcription by RNAPII., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Faithful transfer of parental histones to newly replicated daughter DNA strands is critical for inheritance of epigenetic states. Although replication proteins that facilitate parental histone transfer have been identified, how intact histone H3-H4 tetramers travel from the front to the back of the replication fork remains unknown. Here, we use AlphaFold-Multimer structural predictions combined with biochemical and genetic approaches to identify the Mrc1/CLASPIN subunit of the replisome as a histone chaperone. Mrc1 contains a conserved histone-binding domain that forms a brace around the H3-H4 tetramer mimicking nucleosomal DNA and H2A-H2B histones, is required for heterochromatin inheritance, and promotes parental histone recycling during replication. We further identify binding sites for the FACT histone chaperone in Swi1/TIMELESS and DNA polymerase α that are required for heterochromatin inheritance. We propose that Mrc1, in concert with FACT acting as a mobile co-chaperone, coordinates the distribution of parental histones to newly replicated DNA., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The histone variant macroH2A is generally linked to transcriptionally inactive chromatin, but how macroH2A regulates chromatin structure and functions in the transcriptional process remains elusive. This study reveals that while the integration of human macroH2A1.2 into nucleosomes does not affect their stability or folding dynamics, it notably hinders the maintenance of facilitates chromatin transcription's (FACT's) function. We show that FACT effectively diminishes the stability of macroH2A1.2-nucleosomes and expedites their depletion subsequent to the initial unfolding process. Furthermore, we identify the residue S139 in macroH2A1.2 as a critical switch to modulate FACT's function in nucleosome maintenance. Genome-wide analyses demonstrate that FACT-mediated depletion of macroH2A-nucleosomes allows the correct localization of macroH2A, while the S139 mutation reshapes macroH2A distribution and influences stimulation-induced transcription and cellular response in macrophages. Our findings provide mechanistic insights into the intricate interplay between macroH2A and FACT at the nucleosome level and elucidate their collective role in transcriptional regulation and immune response of macrophages., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Exhausted CD8 T (T ex ) cells in chronic viral infection and cancer have sustained co-expression of inhibitory receptors (IRs). T ex cells can be reinvigorated by blocking IRs, such as PD-1, but synergistic reinvigoration and enhanced disease control can be achieved by co-targeting multiple IRs including PD-1 and LAG-3. To dissect the molecular changes intrinsic when these IR pathways are disrupted, we investigated the impact of loss of PD-1 and/or LAG-3 on T ex cells during chronic infection. These analyses revealed distinct roles of PD-1 and LAG-3 in regulating T ex cell proliferation and effector functions, respectively. Moreover, these studies identified an essential role for LAG-3 in sustaining TOX and T ex cell durability as well as a LAG-3-dependent circuit that generated a CD94/NKG2 + subset of T ex cells with enhanced cytotoxicity mediated by recognition of the stress ligand Qa-1b, with similar observations in humans. These analyses disentangle the non-redundant mechanisms of PD-1 and LAG-3 and their synergy in regulating T ex cells., Competing Interests: Declaration of interests E.J.W. is a member of the Parker Institute for Cancer Immunotherapy which supported this study. E.J.W. is an advisor for Arsenal Bioscience, Coherus, Danger Bio, Janssen, New Limit, Marengo, Pluto Immunotherapeutics, Prox Biosciences, Related Sciences, Santa Ana Bio, and Synthekine. E.J.W. is a founder of and holds stock in Arsenal Biosciences. A.H.S. has patents/pending royalties on the PD-1 pathway from Roche and Novartis and has research funding from IOME, AbbVie, and Taiwan Bio. A.H.S. serves as an advisor for Selecta, Elpiscience, Monopteros, Bicara, Fibrogen, IOME, Bioentre, Corner Therapeutics, Alixia, GlaxoSmithKline, Amgen, and Janssen. D.A.A.V. has patents covering LAG-3, with others pending. D.A.A.V. is a co-founder and stockholder for Novasenta, Tizona, and Trishula; a stockholder for Oncorus and Werewolf; and has patents licensed to Bristol Myers Squibb (BMS) and Novasenta. D.A.A.V. is a scientific advisor for Tizona, Werewolf, F-Star, Bicara, Apeximmune, and T7/Imreg Bio and is a consultant for BMS, Incyte, Regeneron, Ono Pharma, Peptone, and Avidity Partners. D.A.A.V. receives research funding from BMS and Novasenta. O.K. holds stock in Arsenal Biosciences and is an employee of Orange Grove Bio. A.C.H. consults for Immunai and received research funding from BMS and Merck. J.R.G. consults for Arsenal Biosciences., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Chromatin condensation state is the key for retrieving genetic information. High-mobility group protein (HMG) proteins exhibit DNA-binding and bending activities, playing an important role in the regulation of chromatin structure. We have shown that nucleosomes tightly packaged into heterochromatin undergo considerable dynamic histone H2A-H2B maintenance via the direct interaction between HP1/Swi6 and facilitate chromatin transcription (FACT), which is composed of the Spt16/Pob3 heterodimer and Nhp6. In this study, we analyzed the role of Nhp6, an HMG box protein, in the FACT at heterochromatin. Pob3 mutant strains showed derepressed heterochromatin-dependent gene silencing, whereas Nhp6 mutant strains did not show significant defects in chromatin regulation or gene expression, suggesting that these two modules play different roles in chromatin regulation. We expressed a protein fusing Nhp6 to the C-terminus of Pob3, which mimics the multicellular FACT component Ssrp1. The chromatin-binding activity of FACT increased with the number of Nhp6 fused to Pob3, and the heterochromatin formation rate was promoted more strongly. Furthermore, we demonstrated that this promotion of heterochromatinization inhibited the heterochromatic variegation caused by epe1 + disruption. Heterochromatic variegation can be observed in a variety of regulatory steps; however, when it is caused by fluctuations in chromatin arrangement, it can be eliminated through the strong recruitment of the FACT complex., (© 2024 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.)

Thymocyte selection-associated high-mobility group box (TOX) is a transcription factor that is crucial for T cell exhaustion during chronic antigenic stimulation, but its role in inflammation is poorly understood. Here, we report that TOX extracellularly mediates drastic inflammation upon severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection by binding to the cell surface receptor for advanced glycation end-products (RAGE). In various diseases, including COVID-19, TOX release was highly detectable in association with disease severity, contributing to lung fibroproliferative acute respiratory distress syndrome (ARDS). Recombinant TOX-induced blood vessel rupture, similar to a clinical signature in patients experiencing a cytokine storm, further exacerbating respiratory function impairment. In contrast, disruption of TOX function by a neutralizing antibody and genetic removal of RAGE diminished TOX-mediated deleterious effects. Altogether, our results suggest an insight into TOX function as an inflammatory mediator and propose the TOX-RAGE axis as a potential target for treating severe patients with pulmonary infection and mitigating lung fibroproliferative ARDS., Competing Interests: Competing interests statement:The authors declare no competing interest.

Facilitates chromatin transcription (FACT) is a histone chaperone that supports transcription through chromatin in vitro, but its functional roles in vivo remain unclear. Here, we analyze the in vivo functions of FACT with the use of multi-omics analysis after rapid FACT depletion from human cells. We show that FACT depletion destabilizes chromatin and leads to transcriptional defects, including defective promoter-proximal pausing and elongation, and increased premature termination of RNA polymerase II. Unexpectedly, our analysis revealed that promoter-proximal pausing depends not only on the negative elongation factor (NELF) but also on the +1 nucleosome, which is maintained by FACT., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Under stress conditions, ribosome biogenesis is downregulated. This process requires that expression of ribosomal RNA, ribosomal protein, and ribosome biogenesis genes be controlled in a coordinated fashion. The mechanistic Target of Rapamycin Complex 1 (mTORC1) participates in sensing unfavorable conditions to effect the requisite change in gene expression. In Saccharomyces cerevisiae, downregulation of ribosomal protein genes involves dissociation of the activator Ifh1p in a process that depends on Utp22p, a protein that also functions in pre-rRNA processing. Ifh1p has a paralog, Crf1p, which was implicated in communicating mTORC1 inhibition and hence was perceived as a repressor. We focus here on two ribosomal biogenesis genes, encoding Utp22p and the high mobility group protein Hmo1p, both of which are required for communication of mTORC1 inhibition to target genes. Crf1p functions as an activator on these genes as evidenced by reduced mRNA abundance and RNA polymerase II occupancy in a crf1Δ strain. Inhibition of mTORC1 has distinct effects on expression of HMO1 and UTP22; for example, on UTP22, but not on HMO1, the presence of Crf1p promotes the stable depletion of Ifh1p. Our data suggest that Crf1p functions as a weak activator, and that it may be required to prevent re-binding of Ifh1p to some gene promoters after mTORC1 inhibition in situations when Ifh1p is available. We propose that the inclusion of genes encoding proteins required for mTORC1-mediated downregulation of ribosomal protein genes in the same regulatory circuit as the ribosomal protein genes serves to optimize transcriptional responses during mTORC1 inhibition., (© 2024 John Wiley & Sons Ltd.)

In cancer therapy, DNA intercalators are mainly known for their capacity to kill cells by inducing DNA damage. Recently, several DNA intercalators have attracted much interest given their ability to inhibit RNA Polymerase I transcription (BMH-21), evict histones (Aclarubicin) or induce chromatin trapping of FACT (Curaxin CBL0137). Interestingly, these DNA intercalators lack the capacity to induce DNA damage while still retaining cytotoxic effects and stabilize p53. Herein, we report that these DNA intercalators impact chromatin biology by interfering with the chromatin stability of RNA polymerases I, II and III. These three compounds have the capacity to induce degradation of RNA polymerase II and they simultaneously enable the trapping of Topoisomerases TOP2A and TOP2B on the chromatin. In addition, BMH-21 also acts as a catalytic inhibitor of Topoisomerase II, resembling Aclarubicin. Moreover, BMH-21 induces chromatin trapping of the histone chaperone FACT and propels accumulation of Z-DNA and histone eviction, similarly to Aclarubicin and CBL0137. These DNA intercalators have a cumulative impact on general transcription machinery by inducing accumulation of topological defects and impacting nuclear chromatin. Therefore, their cytotoxic capabilities may be the result of compounding deleterious effects on chromatin homeostasis., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

HMG (high mobility group) proteins are a diverse family of nonhistone chromosomal proteins that interact with DNA and a wide range of transcriptional regulators to regulate the structural architecture of DNA. HMGXB4 (also known as HMG2L1) is an HMG protein family member that contains a single HMG box domain. Our previous studies have demonstrated that HMGXB4 suppresses smooth muscle differentiation and exacerbates endotoxemia by promoting a systemic inflammatory response in mice. However, the expression of Hmgxb4 in vivo has not fully examined. Herein, we generated a mouse model that harbors a gene trap in the form of a lacZ gene insertion into the Hmgxb4 gene. This mouse enables the visualization of endogenous HMGXB4 expression in different tissues via staining for the β-galactosidase activity of LacZ which is under the control of the endogenous Hmgxb4 gene promoter. We found that HMGXB4 is widely expressed in mouse tissues and is a nuclear protein. Furthermore, the Hmgxb4 gene trap mice exhibit normal cardiac function and blood pressure. Measurement of β-galactosidase activity in the Hmgxb4 gene trap mice demonstrated that the arterial injury significantly induces Hmgxb4 expression. In summary, the Hmgxb4 gene trap reporter mouse described here provides a valuable tool to examine the expression level of endogenous Hmgxb4 in both physiological and pathological settings in vivo., (© 2024 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

RNA polymerase I (Pol I) is responsible for synthesizing the three largest eukaryotic ribosomal RNAs (rRNAs), which form the backbone of the ribosome. Transcription by Pol I is required for cell growth and, therefore, is subject to complex and intricate regulatory mechanisms. To accomplish this robust regulation, the cell engages a series of trans-acting transcription factors. One such factor, high mobility group protein 1 (Hmo1), has long been established as a trans-acting factor for Pol I in Saccharomyces cerevisiae ; however, the mechanism by which Hmo1 promotes rRNA synthesis has not been defined. Here, we investigated the effect of the deletion of HMO1 on transcription elongation by Pol I in vivo. We determined that Hmo1 is an important activator of transcription elongation, and without this protein, Pol I accumulates across rDNA in a sequence-specific manner. Our results demonstrate that Hmo1 promotes efficient transcription elongation by rendering Pol I less sensitive to pausing in the G-rich regions of rDNA.

RSC (remodels the structure of chromatin) is an essential ATP-dependent chromatin remodeling complex in Saccharomyces cerevisiae. RSC utilizes its ATPase subunit, Sth1, to slide or remove nucleosomes. RSC has been shown to regulate the width of the nucleosome-depleted regions (NDRs) by sliding the flanking nucleosomes away from NDRs. As such, when RSC is depleted, nucleosomes encroach NDRs, leading to transcription initiation defects. In this study, we examined the effects of the catalytic-dead Sth1 on transcription and compared them to those observed during acute and rapid Sth1 depletion by auxin-induced degron strategy. We found that rapid depletion of Sth1 reduces recruitment of TBP and Pol II in highly transcribed genes, as would be expected considering its role in regulating chromatin structure at promoters. In contrast, cells harboring the catalytic-dead Sth1 (sth1-K501R) exhibited a severe reduction in TBP binding, but, surprisingly, also displayed a substantial accumulation in Pol II occupancies within coding regions. The Pol II occupancies further increased upon depleting endogenous Sth1 in the catalytic-dead mutant, suggesting that the inactive Sth1 contributes to Pol II accumulation in coding regions. Notwithstanding the Pol II increase, the ORF occupancies of histone chaperones, FACT and Spt6 were significantly reduced in the mutant. These results suggest a potential role for RSC in recruiting/retaining these chaperones in coding regions. Pol II accumulation despite substantial reductions in TBP, FACT, and Spt6 occupancies in the catalytic-dead mutant could indicate severe transcription elongation and termination defects. Such defects would be consistent with studies showing that RSC is recruited to coding regions in a transcription-dependent manner. Thus, these findings imply a role for RSC in transcription elongation and termination processes, in addition to its established role in transcription initiation., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Chhabi Govind reports financial support was provided by National Institutes of Health. If there are other authors, they 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 Elsevier B.V. All rights reserved.)

Histone chaperone FACT (facilitates chromatin transcription) is well known to promote chromatin recovery during transcription. However, the mechanism how FACT regulates genome-wide chromatin accessibility and transcription factor binding has not been fully elucidated. Through loss-of-function studies, we show here that FACT component Ssrp1 is required for DNA replication and DNA damage repair and is also essential for progression of cell phase transition and cell proliferation in mouse embryonic fibroblast cells. On the molecular level, absence of the Ssrp1 leads to increased chromatin accessibility, enhanced CTCF binding, and a remarkable change in dynamic range of gene expression. Our study thus unequivocally uncovers a unique mechanism by which FACT complex regulates transcription by coordinating genome-wide chromatin accessibility and CTCF binding., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Malignant proliferation and metastasis are the main causes of breast cancer death. The transcription factor high mobility group (HMG) box-containing protein 1 (HBP1) is an important tumor suppressor whose deletion or mutation is closely related to the appearance of tumors. Here, we investigated the role of HBP1 in breast cancer suppression. HBP1 enhances the activity of the tissue inhibitors of metalloproteinases 3 (TIMP3) promoter, thereby increasing protein and mRNA levels of TIMP3. TIMP3 increases the phosphatase and tensin homolog (PTEN) protein level by inhibiting its degradation and acts as a metalloproteinase inhibitor to inhibit the protein levels of MMP2/9. In this study, we demonstrated that the HBP1/TIMP3 axis plays a crucial role in inhibiting the tumorigenesis of breast cancer. HBP1 deletion interferes with the regulation of the axis and induces the occurrence and malignant progression of breast cancer. In addition, the HBP1/TIMP3 axis promotes the sensitivity of breast cancer to radiation therapy and hormone therapy. Our study opens new perspectives on the treatment and prognosis of breast cancer., Competing Interests: Declaration of Competing Interest There's no financial/personal interest or belief that could affect their objectivity., (Copyright © 2023. Published by Elsevier Ltd.)

Purpose: Non-functioning pituitary neuroendocrine tumors are challengingly diagnosed tumors in the clinic. Transsphenoidal surgery remains the first-line treatment. Despite the development of state-of-the-art techniques, no drug therapy is currently approved for the treatment. There are also no randomized controlled trials comparing therapeutic strategies or drug therapy for the management after surgery. Therefore, novel therapeutic interventions for the therapeutically challenging NF-PitNETs are urgently needed., Methods: We integrated epigenome and transcriptome data (both coding and non-coding) that elucidate disease-specific signatures, in addition to biological and pharmacological data, to utilize rational pathway and drug prioritization in NF-PitNETs. We constructed an epigenome- and transcriptome-based PPI network and proposed hub genes. The signature-based drug repositioning based on the integration of multi-omics data was performed., Results: The construction of a disease-specific network based on three different biological levels revealed DCC, DLG5, ETS2, FOXO1, HBP1, HMGA2, PCGF3, PSME4, RBPMS, RREB1, SMAD1, SOCS1, SOX2, YAP1, ZFHX3 as hub proteins. Signature-based drug repositioning using hub proteins yielded repositioned drug candidates that were confirmed in silico via molecular docking. As a result of molecular docking simulations, palbociclib, linifanib, trametinib, eplerenone, niguldipine, and zuclopenthixol showed higher binding affinities with hub genes compared to their inhibitors and were proposed as potential repositioned therapeutics for the management of NF-PitNETs., Conclusion: The proposed systems' biomedicine-oriented multi-omics data integration for drug repurposing to provide promising results for the construction of effective clinical therapeutics. To the best of our knowledge, this is the first study reporting epigenome- and transcriptome-based drug repositioning for NF-PitNETs using in silico confirmations., (© 2022. The Author(s), under exclusive licence to Italian Society of Endocrinology (SIE).)

Background: Mitochondrial transcription factor A (TFAM) is a transcription factor that maintains mitochondrial DNA (mtDNA) stabilization and initiates mtDNA replication. However, little is known about the immune regulation function and TFAM expression in immune cells in the tumors., Methods: Mouse tumor models were applied to analyze the effect of TFAM deficiency in myeloid cell lineage on tumor progression and tumor microenvironment (TME) modification. In vitro, primary mouse bone marrow-derived dendritic cells (BMDCs) were used in the investigation of the altered function and the activated pathway. OVA was used as the model antigen to validate the activation of immune responses in vivo. STING inhibitors were used to confirm the STING activation provoked by Tfam deficient in DCs., Results: The deletion of TFAM in DCs led to mitochondrial dysfunction and mtDNA cytosolic leakage resulting in the cGAS-STING pathway activation in DCs, which contributed to the enhanced antigen presentation. The deletion of TFAM in DCs has interestingly reversed the immune suppressive TME and inhibited tumor growth and metastasis in tumor models., Conclusions: We have revealed that TFAM knockout in DCs ameliorated immune-suppressive microenvironment in tumors through STING pathway. Our work suggests that specific TFAM knockout in DCs might be a compelling strategy for designing novel immunotherapy methods in the future., Competing Interests: Competing interests: No, there are no competing interests., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

DNA is stored in the nucleus of a cell in a folded state; however, only the necessary genetic information is extracted from the required group of genes. The key to extracting genetic information is chromatin ambivalence. Depending on the chromosomal region, chromatin is characterized into low-density "euchromatin" and high-density "heterochromatin", with various factors being involved in its regulation. Here, we focus on chromatin regulation and gene expression by the yeast FACT complex, which functions in both euchromatin and heterochromatin. FACT is known as a histone H2A/H2B chaperone and was initially reported as an elongation factor associated with RNA polymerase II. In budding yeast, FACT activates promoter chromatin by interacting with the transcriptional activators SBF/MBF via the regulation of G1/S cell cycle genes. In fission yeast, FACT plays an important role in the formation of higher-order chromatin structures and transcriptional repression by binding to Swi6, an HP1 family protein, at heterochromatin. This FACT property, which refers to the alternate chromatin-regulation depending on the binding partner, is an interesting phenomenon. Further analysis of nucleosome regulation within heterochromatin is expected in future studies.

The SOX transcription factor family is pivotal in controlling aspects of development. To identify genotype-phenotype relationships of SOX proteins, we performed a non-biased study of SOX using 1890 open-reading frame and 6667 amino acid sequences in combination with structural dynamics to interpret 3999 gnomAD, 485 ClinVar, 1174 Geno2MP, and 4313 COSMIC human variants. We identified, within the HMG (High Mobility Group)- box, twenty-seven amino acids with changes in multiple SOX proteins annotated to clinical pathologies. These sites were screened through Geno2MP medical phenotypes, revealing novel SOX15 R104G associated with musculature abnormality and SOX8 R159G with intellectual disability. Within gnomAD, SOX18 E137K (rs201931544), found within the HMG box of ~0.8% of Latinx individuals, is associated with seizures and neurological complications, potentially through blood-brain barrier alterations. A total of 56 highly conserved variants were found at sites outside the HMG-box, including several within the SOX2 HMG-box-flanking region with neurological associations, several in the SOX9 dimerization region associated with Campomelic Dysplasia, SOX14 K88R (rs199932938) flanking the HMG box associated with cardiovascular complications within European populations, and SOX7 A379V (rs143587868) within an SOXF conserved far C-terminal domain heterozygous in 0.716% of African individuals with associated eye phenotypes. This SOX data compilation builds a robust genotype-to-phenotype association for a gene family through more robust ortholog data integration.

Background: Zhen Wu Decoction (ZWD) is a prescription from the classical text "Treatise on Exogenous Febrile Disease" and has been extensively used to control kidney diseases since the time of the Eastern Han Dynasty., Hypothesis: We hypothesized that ZWD limits tubular fibrogenesis by reinvigorating tubular bio-energetic capacity., Study Design / Methods: A mouse model of chronic kidney disease (CKD) was established using unilateral ureteral obstruction (UUO). Three concentrations of ZWD, namely 25.2 g/kg (high dosage), 12.6 g/kg (middle dosage), and 6.3 g/kg (low dosage), were included to study the dose-effect relationship. Real-time qPCR was used to observe gene transcription in blood samples from patients with CKD. Different siRNAs were designed to study the role of mitochondrial transcription factor A (TFAM) and nuclear factor (erythroid-derived 2)-related factor 2 (NRF2) in transforming growth factor (TGF)-β1 induced fibrogenesis and mitochondrial damage., Results: We showed that ZWD efficiently attenuates renal function impairment and reduces renal interstitial fibrosis. TFAM and NRF2 were repressed, and the stimulator of interferon genes (STING) was activated in CKD patient blood sample. We further confirmed that ZWD activated TFAM depended on NRF2 as an important negative regulator of STING in mouse kidneys. Treatment with ZWD significantly reduced oxidative stress and inflammation by regulating the levels of oxidative phosphorylation (OXPHOS) and pro-inflammatory factors, such as interleukin-6, interleukin-1β, tumor necrosis factor receptor 1, and mitochondrial respiratory chain subunits. NRF2 inhibitors can weaken the ability of ZWD to increase TFAM expression and heal injured mitochondria, playing a similar role to that of STING inhibitors. Our study showed that ZWD elevates the expression of TFAM and mitochondrial respiratory chain subunits by promoting NRF2 activation, after suppressing mitochondrial membrane damage and cristae breakdown and restricting mitochondrial DNA (mtDNA) leakage into the cytoplasm to reduce STING activation., Conclusion: ZWD maintains mitochondrial integrity and improves OXPHOS which represents an innovative insight into "strengthening Yang-Qi" theory. ZWD limits tubular fibrogenesis by reinvigorating tubular bioenergetic capacity., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2022 Elsevier GmbH. All rights reserved.)

Background: Mitochondrial dysfunction is an important pathogenic event in acute kidney injury (AKI). GCN5L1 is a specific acetyltransferase in mitochondria, which regulates glucose and fatty acid metabolism. However, the role of GCN5L1 in mitochondrial dysfunction and the pathogenesis of ischemic AKI are not fully understood., Methods: The protein level of GCN5L1 was detected by western blot assay. Acetylated proteomics was used to explore the level of acetylated TFAM. Duolink proximity ligation assay and co-immunoprecipitation were used to detect the interaction of TFAM and translocase of outer membrane 70 (TOM70). mtDNA copy number, the expression of mitochondrial electron transport chain complexes, the number and morphology of mitochondria were measured. The renal injury of AKI mice was reflected by the levels of creatinine and urea nitrogen and the pathological changes of renal tissue., Results: We showed that GCN5L1 was highly expressed in vivo and in vitro and renal tubules specific knockdown of GCN5L1 could effectively attenuate AKI-induced mitochondrial impairment. Besides, acetylated proteomics revealed that acetylated TFAM was significantly upregulated in AKI mice kidney, which reminded us that TFAM might be an acetylating substrate of GCN5L1. Mechanistically, we evidenced that GCN5L1 could acetylate TFAM at its K76 site and subsequently inhibited its binding to TOM70, thereby reducing TFAM import into mitochondria and mitochondrial biogenesis. Clinically, GCN5L1 and acetylated TFAM were positively correlated with disease severity (all p < 0.05)., Conclusions: In sum, these data demonstrated an unrecognized regulating mechanism of GCN5L1 on TFAM acetylation and its intracellular trafficking, and a potential intervening target for AKI associated mitochondrial disorders as well., (© 2022. The Author(s).)

Background: Oral squamous cell carcinoma (OSCC) is the main type of oral cancer. Disturbing DNA repair is an invaluable way to improve the effectiveness of tumor treatment. Here, we aimed to explore the key enhancer drivers associated with DNA damage repair in OSCC cells., Methods: Gene Set Enrichment Analysis (GSEA), Gene Set Variation Analysis (GSVA) and Kaplan-Meier analysis were applied to explore the relationship among DNA repair-related genes expression and clinical phenotypes based on The Cancer Genome Atlas (TCGA) database. HOMER software and Integrative Genomics Viewer were applied to identify and visualize enhancers using GSE120634. Toolkit for Cistrome Data Browser was applied to predict transcription factors. Human Protein Atlas Database was used to analyze the protein levels of transcription factors in OSCC and control tissues. Seventy-two OSCC patients were included in this study. qRT-PCR was used to detect transcription factor expression in OSCC and adjacent control tissues collected in this study. qRT-PCR and ChIP-qPCR were used to verify the binding of transcription factors to enhancers, and regulation of target genes transcription. Transcription factor knockdown and control cells were treated with cisplatin. CCK8 was used to detect cell viability and proliferation. Western blotting was implemented to detect the levels of DNA repair-related proteins. Transwell assay was used to detect cell invasion., Results: DNA repair was positively associated with the OSCC metastatic phenotype. Patients in the cluster with high expression of DNA repair-related genes had a worse prognosis and a higher proportion of advanced stage, low-differentiation, alcohol consumption and smoking compared to the cluster with low DNA repair-related gene expression. Seventeen metastasis-specific enhancer-controlled upregulated DNA repair-related genes, with the top two upregulated genes being ADRM1 26 S proteasome ubiquitin receptor (ADRM1) and solute carrier family 12 member 7 (SLC12A7) were screened. High mobility group 20 A (HMG20A) was the key prognostic enhancer driver regulating metastasis-specific DNA repair-related genes, with higher expression in OSCC tissues than normal control tissues, and higher expression in metastatic OSCC tissues than non-metastatic OSCC tissues. HMG20A bound to the metastasis-specific enhancers of ADRM1 and SLC12A7, thereby promoting ADRM1 and SLC12A7 expression. Knockdown of HMG20A enhanced cisplatin sensitivity of cells, and inhibited OSCC cells from repairing DNA damage caused by cisplatin, as well as proliferation and invasion of OSCC cells., Conclusion: HMG20A was identified as the key prognostic enhancer driver regulating DNA repair in OSCC cells, providing a new therapeutic target for OSCC., (© 2022. The Author(s).)

SOX10 gene and SOX10 protein are responsible for the gliogenesis of neuroglia from the neural crest cells. Expression of SOX10 gene encodes SOX10 protein which binds with DNA at its minor groove via its HMG domain upon activation. SOX10 protein undergoes bending and changes its conformation after binding with DNA. Via its transactivation domain and HMG domain, it further activates several other transcription factors, these cause gliogenesis of the neural crest cells into neuroglia. In literature, it is stated that the SOX10 gene helps in the formation of schwann cells, oligodendrocytes, and enteric ganglia from neural crest cells. Altered expression of the SOX10 gene results in agliogenesis, dysmyelination, and demyelination in the nervous system as well as intestinal aganglionosis. This review highlighted that there is a role of the SOX10 gene and SOX10 protein in enteric gliogenesis from the neural crest cells., Competing Interests: Declarations of competing interest None, (Copyright © 2022 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.)

Dystrophin Dp71 is the most abundant product of the Duchenne muscular dystrophy gene in the nervous system, and mutations impairing its function have been associated with the neurodevelopmental symptoms present in a third of DMD patients. Dp71 is required for the clustering of neurotransmitter receptors and the neuronal differentiation of cultured cells; nonetheless, its precise role in neuronal cells remains to be poorly understood. In this study, we analyzed the effect of two pathogenic DMD gene point mutations on the Dp71 function in neurons. We engineered C272Y and E299del mutations to express GFP-tagged Dp71 protein variants in N1E-115 and SH-SY5Y neuronal cells. Unexpectedly, the ectopic expression of Dp71 mutants resulted in protein aggregation, which may be mechanistically caused by the effect of the mutations on Dp71 structure, as predicted by protein modeling and molecular dynamics simulations. Interestingly, Dp71 mutant variants acquired a dominant negative function that, in turn, dramatically impaired the distribution of different Dp71 protein partners, including β-dystroglycan, nuclear lamins A/C and B1, the high-mobility group (HMG)-containing protein (BRAF35) and the BRAF35-family-member inhibitor of BRAF35 (iBRAF). Further analysis of Dp71 mutants provided evidence showing a role for Dp71 in modulating both heterochromatin marker H3K9me2 organization and the neuronal genes' expression, via its interaction with iBRAF and BRAF5.

Yeast Hmo1 is a high mobility group B (HMGB) protein that participates in the transcription of ribosomal protein genes and rDNA, and also stimulates the activities of some ATP-dependent remodelers. Hmo1 binds both DNA and nucleosomes and has been proposed to be a functional yeast analog of mammalian linker histones. We used EMSA and single particle Förster resonance energy transfer (spFRET) microscopy to characterize the effects of Hmo1 on nucleosomes alone and with the histone chaperone FACT. Hmo1 induced a significant increase in the distance between the DNA gyres across the nucleosomal core, and also caused the separation of linker segments. This was opposite to the effect of the linker histone H1, which enhanced the proximity of linkers. Similar to Nhp6, another HMGB factor, Hmo1, was able to support large-scale, ATP-independent, reversible unfolding of nucleosomes by FACT in the spFRET assay and partially support FACT function in vivo. However, unlike Hmo1, Nhp6 alone does not affect nucleosome structure. These results suggest physiological roles for Hmo1 that are distinct from Nhp6 and possibly from other HMGB factors and linker histones, such as H1.

Obesity and its associated metabolic disease do serious harm to human health. The transcriptional cascade network with transcription factors as the core is the focus of current research on adipogenesis and its mechanism. Previous studies have found that HMG domain protein 20A (HMG20A) is highly expressed in the early stage of adipogenic differentiation of porcine intramuscular fat (IMF), which may be involved in regulating adipogenesis. In this study, HMG20A was found to play a key negative regulatory role in adipogenesis. Gain- and loss-of-function studies revealed that HMG20A inhibited the differentiation of SVF cells and C3H10T1/2 cells into mature adipocytes. RNA-seq was used to screen differentially expressed genes after HMG20A knockdown. qRT-PCR and ChIP-PCR confirmed that MEF2C was the real target of HMG20A, and HMG20A played a negative regulatory role through MEF2C. HMG20A binding protein LSD1 was found to alleviate the inhibitory effect of HMG20A on adipogenesis. Further studies showed that HMG20A could cooperate with LSD1 to increase the H3K4me2 of the MEF2C promoter and then increase the expression of MEF2C. Collectively, these findings highlight a role for HMG20A-dependent transcriptional and epigenetic regulation in adipogenesis.

Patterning of the anterior-posterior axis is fundamental to animal development. The Wnt pathway plays a major role in this process by activating the expression of posterior genes in animals from worms to humans. This observation raises the question of whether the Wnt pathway or other regulators control the expression of the many anterior-expressed genes. We found that the expression of five anterior-specific genes in Caenorhabditis elegans embryos depends on the Wnt pathway effectors pop-1/TCF and sys-1/β-catenin. We focused further on one of these anterior genes, ref-2/ZIC, a conserved transcription factor expressed in multiple anterior lineages. Live imaging of ref-2 mutant embryos identified defects in cell division timing and position in anterior lineages. Cis-regulatory dissection identified three ref-2 transcriptional enhancers, one of which is necessary and sufficient for anterior-specific expression. This enhancer is activated by the T-box transcription factors TBX-37 and TBX-38, and surprisingly, concatemerized TBX-37/38 binding sites are sufficient to drive anterior-biased expression alone, despite the broad expression of TBX-37 and TBX-38. Taken together, our results highlight the diverse mechanisms used to regulate anterior expression patterns in the embryo., (Copyright © 2022 Elsevier Inc. All rights reserved.)

FACT (FAcilitates Chromatin Transcription) is a heterodimeric protein complex composed of SUPT16H and SSRP1, and a histone chaperone participating in chromatin remodeling during gene transcription. FACT complex is profoundly regulated, and contributes to both gene activation and suppression. Here we reported that SUPT16H, a subunit of FACT, is acetylated in both epithelial and natural killer (NK) cells. The histone acetyltransferase TIP60 contributes to the acetylation of SUPT16H middle domain (MD) at lysine 674 (K674). Such acetylation of SUPT16H is recognized by bromodomain protein BRD4, which promotes protein stability of SUPT16H in both epithelial and NK cells. We further demonstrated that SUPT16H-BRD4 associates with histone modification enzymes (HDAC1, EZH2), and further regulates their activation status and/or promoter association as well as affects the relevant histone marks (H3ac, H3K9me3 and H3K27me3). BRD4 is known to profoundly regulate interferon (IFN) signaling, while such function of SUPT16H has never been explored. Surprisingly, our results revealed that SUPT16H genetic knockdown via RNAi or pharmacological inhibition by using its inhibitor, curaxin 137 (CBL0137), results in the induction of IFNs and interferon-stimulated genes (ISGs). Through this mechanism, depletion or inhibition of SUPT16H is shown to efficiently inhibit infection of multiple viruses, including Zika, influenza, and SARS-CoV-2. Furthermore, we demonstrated that depletion or inhibition of SUPT16H also causes the remarkable activation of IFN signaling in NK cells, which promotes the NK-mediated killing of virus-infected cells in a co-culture system using human primary NK cells. Overall, our studies unraveled the previously un-appreciated role of FACT complex in coordinating with BRD4 and regulating IFN signaling in both epithelial and NK cells, and also proposed the novel application of the FACT inhibitor CBL0137 to treat viral infections., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Puf5, a Puf-family RNA-binding protein, binds to 3´ untranslated region of target mRNAs and negatively regulates their expression in Saccharomyces cerevisiae. The puf5Δ mutant shows pleiotropic phenotypes including a weakened cell wall, a temperature-sensitive growth, and a shorter lifespan. To further analyze a role of Puf5 in cell growth, we searched for a multicopy suppressor of the temperature-sensitive growth of the puf5Δ mutant in this study. We found that overexpression of CLB2 encoding B-type cyclin suppressed the temperature-sensitive growth of the puf5Δ mutant. The puf5Δ clb2Δ double mutant displayed a severe growth defect, suggesting that Puf5 positively regulates the expression of a redundant factor with Clb2 in cell cycle progression. We found that expression of CLB1 encoding a redundant B-type cyclin was decreased in the puf5Δ mutant, and that this decrease of the CLB1 expression contributed to the growth defect of the puf5Δ clb2Δ double mutant. Since Puf5 is a negative regulator of the gene expression, we hypothesized that Puf5 negatively regulates the expression of a factor that represses CLB1 expression. We found such a repressor, Ixr1, which is an HMGB (High Mobility Group box B) protein. Deletion of IXR1 restored the decreased expression of CLB1 caused by the puf5Δ mutation and suppressed the growth defect of the puf5Δ clb2Δ double mutant. The expression of IXR1 was negatively regulated by Puf5 in an IXR1 3´ UTR-dependent manner. Our results suggest that IXR1 mRNA is a physiologically important target of Puf5, and that Puf5 and Ixr1 contribute to the cell cycle progression through the regulation of the cell cycle-specific expression of CLB1., Competing Interests: The authors have declared that no competing interests exist.

When asynchronously growing cells suffer from nutrient depletion and inactivation of target of rapamycin complex 1 (TORC1) protein kinase, the rDNA (rRNA gene) region is condensed in budding yeast Saccharomyces cerevisiae, which is executed by condensin and Cdc14 protein phosphatase. However, it is unknown whether these mitotic factors can condense the rDNA region in nutrient-starved interphase cells. Here, we show that condensin is not involved in TORC1 inactivation-induced rDNA condensation in G1 cells. Instead, the high-mobility group protein Hmo1 drove this process. The histone deacetylase Rpd3 and Cdc14, which repress rRNA transcription, were both required for the interphase rDNA condensation. Furthermore, interphase rDNA condensation necessitated CLIP and cohibin that tether rDNA to inner nuclear membranes. Finally, we showed that Hmo1, CLIP, Rpd3, and Cdc14 were required for survival in nutrient-starved G1 cells. Thus, this study disclosed novel features of interphase chromosome condensation., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Purpose: The specific mechanisms and biomarkersunderlying the progression of stable coronary artery disease (CAD) to acute myocardial infarction (AMI) remain unclear. The current study aims to explore novel gene biomarkers associated with CAD progression by analyzing the transcriptomic sequencing data of peripheral blood monocytes in different stages of CAD., Material and Methods: A total of 24 age- and sex- matched patients at different CAD stages who received coronary angiography were enrolled, which included 8 patients with normal coronary angiography, 8 patients with angiographic intermediate lesion, and 8 patients with AMI. The RNA from peripheral blood monocytes was extracted and transcriptome sequenced to analyze the gene expression and the differentially expressed genes (DEG). A Gene Oncology (GO) enrichment analysis was performed to analyze the biological function of genes. Weighted gene correlation network analysis (WGCNA) was performed to classify genes into several gene modules with similar expression profiles, and correlation analysis was carried out to explore the association of each gene module with a clinical trait. The dynamic network biomarker (DNB) algorithm was used to calculate the key genes that promote disease progression. Finally, the overlapping genes between different analytic methods were explored., Results: WGCNA analysis identified a total of nine gene modules, of which two modules have the highest positive association with CAD stages. GO enrichment analysis indicated that the biological function of genes in these two gene modules was closely related to inflammatory response, which included T-cell activation, cell response to inflammatory stimuli, lymphocyte activation, cytokine production, and the apoptotic signaling pathway. DNB analysis identified a total of 103 genes that may play key roles in the progression of atherosclerosis plaque. The overlapping genes between DEG/WGCAN and DNB analysis identified the following 13 genes that may play key roles in the progression of atherosclerosis disease: SGPP2, DAZAP2, INSIG1, CD82, OLR1, ARL6IP1, LIMS1, CCL5, CDK7, HBP1, PLAU, SELENOS, and DNAJB6., Conclusions: The current study identified a total of 13 genes that may play key roles in the progression of atherosclerotic plaque and provides new insights for early warning biomarkers and underlying mechanisms underlying the progression of CAD., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Song, Qiao, Chen, Ge, Zhang, Yuan, Bian, Wang, Liu, Jia, Fu and Dou.)

Thymocyte selection-associated high mobility group box (TOX) has been described to be a key regulator in the formation of CD8+ T cell exhaustion. Hepatitis C virus (HCV) infection with different lengths of antigen exposure in acute, chronic, and after resolution of HCV infection is the ideal immunological model to study the expression of TOX in HCV-specific CD8+ T cells with different exposure to antigen. HCV-specific CD8+ T cells from 35 HLA-A*01:01, HLA-A*02:01, and HLA-A*24:02 positive patients were analyzed with a 16-color FACS-panel evaluating the surface expression of lineage markers (CD3, CD8), ectoenzymes (CD39, CD73), markers of differentiation (CD45RO, CCR7, CD127), and markers of exhaustion and activation (TIGIT, PD-1, KLRG1, CD226) and transcription factors (TOX, Eomesodermin, T-bet). Here, we defined on-target T cells as T cells against epitopes without escape mutations and off-target T cells as those against a "historical" antigen mutated in the autologous sequence. TOX+HCV-specific CD8+ T cells from patients with chronic HCV and on-target T cells displayed co-expression of Eomesodermin and were associated with the formation of terminally exhausted CD127 - PD1 hi , CD39 hi , CD73 low CD8+ T cells. In contrast, TOX+HCV-specific CD8+ T cells in patients with off-target T cells represented a progenitor memory Tex phenotype characterized by CD127 hi expression and a CD39 low and CD73 hi phenotype. TOX+HCV-specified CD8+ T cells in patients with a sustained virologic response were characterized by a memory phenotype (CD127+, CD73 hi ) and co-expression of immune checkpoints and Eomesodermin, indicating a key structure in priming of HCV-specific CD8+ T cells in the chronic stage, which persisted as a residual after therapy. Overall, the occurrence of TOX+HCV-specific CD8+ T cells was revealed at each disease stage, which impacted the development of progenitor Tex, intermediate Tex, and terminally exhausted T cell through an individual molecular footprint. In sum, TOX is induced early during acute infection but is modulated by changes in viral sequence and antigen recognition. In the case of antigen persistence, the interaction with Eomesodermin leads to the formation of terminally exhausted virus-specific CD8+ T cells, and there was a direct correlation of the co-expression of TOX and Eomes and terminally exhausted phenotype of virus-specific CD8+ T cells., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wildner, Walker, Brauneck, Ditt, Peine, Huber, Haag, Beisel, Timm and Schulze zur Wiesch.)

Sepsis, a systemic inflammatory response disease, is the most severe complication of infection and a deadly disease. High mobility group proteins (HMGs) are non-histone nuclear proteins binding nucleosomes and regulate chromosome architecture and gene transcription, which act as a potent pro-inflammatory cytokine involved in the delayed endotoxin lethality and systemic inflammatory response. HMGs increase in serum and tissues during infection, especially in sepsis. A growing number of studies have demonstrated HMGs are not only cytokines which can mediate inflammation, but also potential therapeutic targets in sepsis. To reduce sepsis-related mortality, a better understanding of HMGs is essential. In this review, we described the structure and function of HMGs, summarized the definition, epidemiology and pathophysiology of sepsis, and discussed the HMGs-related mechanisms in sepsis from the perspectives of non-coding RNAs (microRNA, long non-coding RNA, circular RNA), programmed cell death (apoptosis, necroptosis and pyroptosis), drugs and other pathophysiological aspects to provide new targets and ideas for the diagnosis and treatment of sepsis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liang and He.)

Methods: We search PubMed and Web of Sciences with keywords "SSRP1" and "Cancer." Only English literature was included, and conference papers and abstract were all excluded., Results: Transcription factors are classified into three groups based on their DNA binding motifs: simple helix-loop-helix (bHLH), classical zinc fingers (ZF-TFs), and homeodomains. The tumor-suppressive miR-497 (microRNA-497) acted as an undesirable regulator of SSRP1 upregulation, which led to tumor growth. The siRNA (small interfering RNA) knockdown of SSRP1 hindered cell proliferation along with incursion and glioma cell migration. Through the AKT (also known as protein kinase B) signaling pathway, SSRP1 silencing affected cancer apoptosis and cell proliferation., Conclusion: The MAPK (mitogen-activated protein kinase) signaling pathway's phosphorylation was suppressed when SSRP1 was depleted. The effect of curaxins on p53 and NF-B (nuclear factor- κ B), and their toxicity to cancer cells, is attributable to the FACT (facilitates chromatin transcription) complex's chromatin trapping., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Shengnan Jia et al.)

Preservation of nucleosomes during replication has been extensively studied, while the maintenance of nucleosomes during transcription has gotten less attention. The histone chaperone FACT has a role in transcription elongation, although whether it disassembles or assembles nucleosomes during this process is unclear. To elucidate the function of FACT in mammals, we deleted the Ssrp1 subunit of FACT in adult mice. FACT loss is lethal, possibly due to the loss of the earliest progenitors in bone marrow and intestine, while more differentiated cells are not affected. Using cells isolated from several tissues, we show that FACT loss reduces the viability of stem cells but not of cells differentiated in vitro. FACT depletion increases chromatin accessibility in a transcription-dependent manner in adipose mesenchymal stem cells, indicating that nucleosomes are lost in these cells during transcription in the absence of FACT. We also observe activation of interferon (IFN) signaling and the accumulation of immunocytes in organs sensitive to FACT loss. Our data indicate that FACT maintains chromatin integrity during transcription in mammalian adult stem cells, suggesting that chromatin transcription in stem cells and differentiated cells is different., (© 2022 The Authors.)

Background: Gastric carcinoma (GC) is one of the most deadly diseases due to tumour metastasis and resistance to therapy. Understanding the molecular mechanism of tumour progression and drug resistance will improve therapeutic efficacy and develop novel intervention strategies., Methods: Differentially expressed long non-coding RNAs (lncRNAs) in clinical specimens were identified by LncRNA microarrays and validated in different clinical cohorts by quantitative real-time polymerase chain reaction (qRT-PCR), in situ hybridisation and bioinformatics analysis. Biological functions of lncRNA were investigated by using cell proliferation assays, migration assays, xenograft tumour models and bioinformatics analysis. Effects of lncSLCO1C1 on GC cell survival were assessed by comet assays and immunofluorescence assays. Underlying molecular mechanisms were further explored by using a number of technologies including RNA pull-down, mass spectrometry analysis, RNA immunoprecipitation, co-immunoprecipitation, miRNA sequencing, luciferase reporter assays and molecular modelling., Results: LncSLCO1C1 was highly upregulated in GC tissue samples and associated with GC patients' poor overall survival. Overexpression of lncSLCO1C1 promoted proliferation and migration, whereas decreased lncSLCO1C1 expression produced the opposite effects. lncSLCO1C1 also mediated tumour resistance to chemotherapy with oxaliplatin by reducing DNA damage and increasing cell proliferation. Despite sequence overlapping between lncSLCO1C1 and PDE3A, alternations of PDE3A expression had no effect on the GC cell progression, indicating that lncSLCO1C1, not PDE3A, related with the progression of GC cells. Mechanistically, lncSLCO1C1 serves as a scaffold for the structure-specific recognition protein 1 (SSRP1)/H2A/H2B complex and regulates the function of SSRP1 in reducing DNA damage. Meanwhile, lncSLCO1C1 functions as a sponge to adsorb miR-204-5p and miR-211-5p that target SSRP1 mRNA, and thus increases SSRP1 expression. Patients with high expressions of both lncSLCO1C1 and SSRP1 have poor overall survival, highlighting the role of lncSLCO1C1 in GC progression., Conclusions: LncSLCO1C1 promotes GC progression by enhancing cell growth and preventing DNA damage via interacting and scaffolding the SSRP1/H2A/H2b complex and absorbing both miR-211-5p and miR-204-5p to increase SSRP1 expression., (© 2022 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Alveolar macrophages (AMs) are major lung tissue-resident macrophages capable of proliferating and self-renewal in situ. AMs are vital in pulmonary antimicrobial immunity and surfactant clearance. The mechanisms regulating AM compartment formation and maintenance remain to be fully elucidated currently. In this study, we have explored the roles of mitochondrial transcription factor A (TFAM)-mediated mitochondrial fitness and metabolism in regulating AM formation and function. We found that TFAM deficiency in mice resulted in significantly reduced AM numbers and impaired AM maturation in vivo. TFAM deficiency was not required for the generation of AM precursors nor the differentiation of AM precursors into AMs, but was critical for the maintenance of AM compartment. Mechanistically, TFAM deficiency diminished gene programs associated with AM proliferation and self-renewal and promoted the expression of inflammatory genes in AMs. We further showed that TFAM-mediated AM compartment impairment resulted in defective clearance of cellular debris and surfactant in the lung and increased the host susceptibility to severe influenza virus infection. Finally, we found that influenza virus infection in AMs led to impaired TFAM expression and diminished mitochondrial fitness and metabolism. Thus, our data have established the critical function of TFAM-mediated mitochondrial metabolism in AM maintenance and function., (Copyright © 2022 by The American Association of Immunologists, Inc.)

DNA repair confers the resistance of tumor cells to DNA-damaging anticancer therapies, while how reprogrammed metabolism in tumor cells contributes to such process remains poorly understood. Pyruvate kinase M2 isoform (PKM2) catalyzes the conversion of phosphoenolpyruvate to pyruvate and regulates the last rate-limiting step of glycolysis. Here it is shown that the glycolytic metabolite pyruvate enhances DNA damage repair by facilitating chromatin loading of γH2AX, thereby promoting the radiation resistance of glioma cells. Mechanistically, PKM2 is phosphorylated at serine (S) 222 upon DNA damage and interacts with FACT complex, a histone chaperone comprising SPT16 and SSRP1 subunit. The pyruvate produced by PKM2 directly binds to SSRP1, which increases the association of FACT complex with γH2AX and subsequently facilitates FACT-mediated chromatin loading of γH2AX, ultimately promoting DNA repair and tumor cell survival. Intriguingly, the supplementation of exogenous pyruvate can also sufficiently enhance FACT-mediated chromatin loading of γH2AX and promotes tumor cell survival upon DNA damage. The levels of PKM2 S222 phosphorylation correlate with the malignancy and prognosis of human glioblastoma. The finding demonstrates a novel mechanism by which PKM2-produced pyruvate promotes DNA repair by regulating γH2AX loading to chromatin and establishes a critical role of this mechanism in glioblastoma radiation resistance., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Ginsenoside Rg3 (Rg3), amplified by iterative heating processing with fresh ginseng, has a broad range of pharmacological activities and improves mitochondrial biogenesis in skeletal muscle. However, thus far no study has examined how Rg3 affects myotube growth or muscle atrophy, to the best of the authors' knowledge. The present study was conducted to examine the myogenic effect of Rg3 on dexamethasone (DEX)‑induced myotube atrophy and the underlying molecular mechanisms. Rg3 activated Akt/mammalian target of rapamycin signaling to prevent DEX‑induced myotube atrophy thereby stimulating the expression of muscle‑specific genes, including myosin heavy chain and myogenin, and suppressing muscle‑specific ubiquitin ligases as demonstrated by immunoblotting and immunostaining assays. Furthermore, Rg3 efficiently prevented DEX‑triggered mitochondrial dysfunction of myotubes through peroxisome proliferator‑activated receptor‑γ coactivator1α activities and its mitochondrial biogenetic transcription factors, nuclear respiratory factor‑1 and mitochondrial transcription factor A. These were confirmed by immunoblotting, luciferase assays, RT‑qPCR and mitochondrial analysis measuring the levels of ROS, ATP and membrane potential. By providing a mechanistic insight into the effect of Rg3 on myotube atrophy, the present study suggested that Rg3 has potential as a therapeutic or nutraceutical remedy to intervene in muscle aging or diseases including cancer cachexia.

The identification of FACT as a histone chaperone enabling transcription through chromatin in vitro has strongly shaped how its roles are envisioned. However, FACT has been implicated in essentially all aspects of chromatin biology, from transcription to DNA replication, DNA repair, and chromosome segregation. In this review, we focus on recent literature describing the role and mechanisms of FACT during transcription. We highlight the prime importance of FACT in preserving chromatin integrity during transcription and challenge its role as an elongation factor. We also review evidence for FACT's role as a cell-type/gene-specific regulator of gene expression and briefly summarize current efforts at using FACT inhibition as an anti-cancer strategy.

The purpose of our study is to determine the protective effects of mitophagy enhancers against phosphorylated tau (P-tau)-induced mitochondrial and synaptic toxicities in Alzheimer's disease (AD). Mitochondrial abnormalities, including defective mitochondrial dynamics, biogenesis, axonal transport and impaired clearance of dead mitochondria are linked to P-tau in AD. Mitophagy enhancers are potential therapeutic candidates to clear dead mitochondria and improve synaptic and cognitive functions in AD. We recently optimized the doses of mitophagy enhancers urolithin A, actinonin, tomatidine, nicotinamide riboside in immortalized mouse primary hippocampal (HT22) neurons. In the current study, we treated mutant Tau expressed in HT22 (mTau-HT22) cells with mitophagy enhancers and assessed mRNA and protein levels of mitochondrial/synaptic genes, cell survival and mitochondrial respiration. We also assessed mitochondrial morphology in mTau-HT22 cells treated and untreated with mitophagy enhancers. Mutant Tau-HT22 cells showed increased fission, decreased fusion, synaptic & mitophagy genes, reduced cell survival and defective mitochondrial respiration. However, these events were reversed in mitophagy enhancers treated mTau-HT22 cells. Cell survival was increased, mRNA and protein levels of mitochondrial fusion, synaptic and mitophagy genes were increased, and mitochondrial fragmentation is reduced in mitophagy enhancers treated mTau-HT22 cells. Further, urolithin A showed strongest protective effects among all enhancers tested in AD. Our combination treatments of urolithin A + EGCG, addition to urolithin A and EGCG individual treatment revealed that combination treatments approach is even stronger than urolithin A treatment. Based on these findings, we cautiously propose that mitophagy enhancers are promising therapeutic drugs to treat mitophagy in patients with AD., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Medulloblastoma (MB) is a malignant pediatric brain tumor with a poor prognosis. Post-surgical radiation and cisplatin-based chemotherapy have been a mainstay of treatment, which often leads to substantial neurocognitive impairments and morbidity, highlighting the need for a novel therapeutic target to enhance the sensitivity of MB tumors to cytotoxic therapies. We performed a comprehensive study using a cohort of 71 MB patients' samples and pediatric MB cell lines and found that MB tumors have elevated levels of nucleosome remodeling FACT (FAcilitates Chromatin Transcription) complex and DNA repair enzyme AP-endonuclease1 (APE1). FACT interacts with APE1 and facilitates recruitment and acetylation of APE1 to promote repair of radiation and cisplatin-induced DNA damage. Further, levels of FACT and acetylated APE1 both are correlate strongly with MB patients' survival. Targeting FACT complex with CBL0137 inhibits DNA repair and alters expression of a subset of genes, and significantly improves the potency of cisplatin and radiation in vitro and in MB xenograft. Notably, combination of CBL0137 and cisplatin significantly suppressed MB tumor growth in an intracranial orthotopic xenograft model. We conclude that FACT complex promotes chemo-radiation resistance in MB, and FACT inhibitor CBL0137 can be used as a chemo-radiation sensitizer to augment treatment efficacy and reduce therapy-related toxicity in high-risk pediatric patients., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The histone chaperone FACT occupies transcribed regions where it plays prominent roles in maintaining chromatin integrity and preserving epigenetic information. How it is targeted to transcribed regions, however, remains unclear. Proposed models include docking on the RNA polymerase II (RNAPII) C-terminal domain (CTD), recruitment by elongation factors, recognition of modified histone tails, and binding partially disassembled nucleosomes. Here, we systematically test these and other scenarios in Saccharomyces cerevisiae and find that FACT binds transcribed chromatin, not RNAPII. Through a combination of high-resolution genome-wide mapping, single-molecule tracking, and mathematical modeling, we propose that FACT recognizes the +1 nucleosome, as it is partially unwrapped by the engaging RNAPII, and spreads to downstream nucleosomes aided by the chromatin remodeler Chd1. Our work clarifies how FACT interacts with genes, suggests a processive mechanism for FACT function, and provides a framework to further dissect the molecular mechanisms of transcription-coupled histone chaperoning., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Renal cell carcinoma (RCC) is a common tumor of the urinary system, and its global incidence is increasing annually. Circular RNAs (circRNAs) are involved in RCC tumorigenesis; however, the role of circ-EGLN3 (hsa&#95;circ&#95;0031594) derived from the Egl nine homolog 3 (EGLN3) gene in RCC remains undetermined., Methods: Circ-EGNL3 expression was examined before and after RNase R and actinomycin treatments in RCC cells and tissues. Cell proliferation, migration, and invasion were assessed using the CCK-8 assay, EdU staining, and wound-healing and Transwell assays. The interactions between microRNA (miR)-1224-3p and circ-EGLN3, and between miR-1224-3p and HMG box domain containing 3 (HMGXB3) were predicted by bioinformatics analysis and validated by dual-luciferase reporter assay., Results: Circ-EGLN3 was identified using RNase R and actinomycin treatments. Circ-EGLN3 was upregulated in RCC cells and tissues and correlated with poor overall survival. Silencing of circ-EGNL3 decreased RCC cell proliferation, migration, and invasion. Mechanistic studies indicated that circ-EGNL3 acts as a sponge for miR-1224-3p, which targeted HMGXB3. Circ-EGNL3 indirectly upregulated HMGXB3 by targeting miR-1224-3p, and overexpression of circ-EGLN3 reversed the repressive effects of miR-1224-3p on RCC., Conclusion: Circ-EGLN3 regulated RCC progression through the miR-1224-3p/HMGXB3 axis, suggesting its potential as a therapeutic target., (Copyright © 2021 Elsevier GmbH. All rights reserved.)

Sense-antisense mRNA pairs generated by convergent transcription is a way of gene regulation. c-fms gene is closely juxtaposed to the HMGXB3 gene in the opposite orientation, in chromosome 5. The intergenic region (IR) between c-fms and HMGXB3 genes is 162 bp. We found that a small portion (∼4.18%) of HMGXB3 mRNA is transcribed further downstream, including the end of the c-fms gene generating antisense mRNA against c-fms mRNA. Similarly, a small portion (∼1.1%) of c-fms mRNA is transcribed further downstream, including the end of the HMGXB3 gene generating antisense mRNA against the HMGXB3 mRNA. Insertion of the strong poly(A) signal sequence in the IR results in decreased c-fms and HMGXB3 antisense mRNAs, resulting in up-regulation of both c-fms and HMGXB3 mRNA expression. miR-324-5p targets HMGXB3 mRNA 3' UTR, and as a result, regulates c-fms mRNA expression. HuR stabilizes c-fms mRNA, and as a result, down-regulates HMGXB3 mRNA expression. UALCAN analysis indicates that the expression pattern between c-fms and HMGXB3 proteins are opposite in vivo in breast cancer tissues. Together, our results indicate that the mRNA encoded by the HMGXB3 gene can influence the expression of adjacent c-fms mRNA, or vice versa., (© 2021 Woo and Chambers; Published by Cold Spring Harbor Laboratory Press for the RNA Society.)

Mitochondrial diseases are genetic disorders that lead to impaired mitochondrial function, resulting in exercise intolerance and muscle weakness. In patients, muscle fatigue due to defects in mitochondrial oxidative capacities commonly precedes muscle weakness. In mice, deletion of the fast-twitch skeletal muscle-specific Tfam gene (Tfam KO) leads to a deficit in respiratory chain activity, severe muscle weakness and early death. Here, we performed a time-course study of mitochondrial and muscular dysfunctions in 11- and 14-week-old Tfam KO mice, i.e. before and when mice are about to enter the terminal stage, respectively. Although force in the unfatigued state was reduced in Tfam KO mice compared to control littermates (wild type) only at 14 weeks, during repeated submaximal contractions fatigue was faster at both ages. During fatiguing stimulation, total phosphocreatine breakdown was larger in Tfam KO muscle than in wild-type muscle at both ages, whereas phosphocreatine consumption was faster only at 14 weeks. In conclusion, the Tfam KO mouse model represents a reliable model of lethal mitochondrial myopathy in which impaired mitochondrial energy production and premature fatigue occur before muscle weakness and early death., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

Mitochondrial transcription factor A (TFAM) is compacting mitochondrial DNA (dmtDNA) into nucleoids and directly controls mtDNA copy number. Here, we show that the TFAM-to-mtDNA ratio is critical for maintaining normal mtDNA expression in different mouse tissues. Moderately increased TFAM protein levels increase mtDNA copy number but a normal TFAM-to-mtDNA ratio is maintained resulting in unaltered mtDNA expression and normal whole animal metabolism. Mice ubiquitously expressing very high TFAM levels develop pathology leading to deficient oxidative phosphorylation (OXPHOS) and early postnatal lethality. The TFAM-to-mtDNA ratio varies widely between tissues in these mice and is very high in skeletal muscle leading to strong repression of mtDNA expression and OXPHOS deficiency. In the heart, increased mtDNA copy number results in a near normal TFAM-to-mtDNA ratio and maintained OXPHOS capacity. In liver, induction of LONP1 protease and mitochondrial RNA polymerase expression counteracts the silencing effect of high TFAM levels. TFAM thus acts as a general repressor of mtDNA expression and this effect can be counterbalanced by tissue-specific expression of regulatory factors., (© 2021 Bonekamp et al.)