Search

Basic helix-loop-helix domains in yeast regulatory proteins Ino2 and Ino4 mediate formation of a heterodimer which binds to and activates expression of phospholipid biosynthetic genes. The human proto-oncoprotein c-Myc (Myc) and its binding partner Max activate genes important for cellular proliferation and contain functional domains structure and position of which strongly resembles Ino2 and Ino4. Since Ino2-Myc and Ino4-Max may be considered as orthologs we performed functional comparisons in yeast. We demonstrate that Myc and Max could be stably synthesized in S. cerevisiae and together significantly activated a target gene of Ino2/Ino4 but nevertheless were unable to functionally complement an ino2 ino4 double mutant. We also map two efficient transcriptional activation domains in the N-terminus of Myc (TAD1: aa 1-41 and TAD2: aa 91-140), corresponding to TAD positions in Ino2. We finally show that coactivators such as TFIID subunits Taf1, Taf4, Taf6, Taf10 and Taf12 as well as ATPase subunits of chromatin remodelling complexes Swi2, Sth1 and Ino80 previously shown to interact with TADs of Ino2 were also able to bind TADs of Myc, supporting the view that heterodimers Ino2/Ino4 and Myc/Max are evolutionary related but have undergone transcriptional rewiring of target genes., Competing Interests: Declarations. Conflicts of interest: The authors declare no competing interests. Ethics approval: Not applicable (no studies with human participants or animals were performed in this study). Consent to participate: Not applicable. Consent for publication: All authors have read and approved the final manuscript., (© 2025. The Author(s).)

TAF8 is part of the transcription factor TFIID complex. TFIID is crucial for recruiting the transcription factor complex containing RNA polymerase II. TAF8 deficiency was recently reported as causing a severe neurodevelopmental disorder in eight patients. We have ascertained three Muslim Arab couples with fetal brain malformations. Clinical, imaging, pathological, biochemical, and molecular analyses were performed. Pre-natal ultrasound performed in four pregnancies revealed massive cerebellar atrophy, microcephaly, cerebral and corpus callosum (CC) anomalies. Pre-natal MRI studies of two of the affected fetuses confirmed microcephaly, small vermis, abnormal sulcation pattern with malformation, and shortening of CC. The fetuses were found to carry a novel likely pathogenic homozygous variant (c.45 + 5 G > A) of TAF8, predicted to affect splicing and presenting autosomal recessive inheritance. Post-mortem examinations confirmed the imaging studies in one fetus. Dysmorphic features including hypertelorism, wide nasal bridge, clinodactyly, and hirsutism were present. Western blotting analysis in fibroblasts of an affected fetus demonstrated a significant reduction of TAF8 protein. We determined high expression levels of TAF8 which progressively diminish in fetal brains of WT mice. We report for the first time the fetal presentation of TAF8 deficiency due to a novel genetic variant, and study TAF8 presence during fetal and neonatal periods in mouse brains. Our study may contribute to understanding the role of TAF8 in the developing human brain., Competing Interests: Competing interests: The authors declare no competing interests. Ethics approval and consent to participate: The guardians of the affected individuals signed an informed consent form for participation in this study. The Israeli Supreme Helsinki Committee approved the study (GMC- 991110, NHR-58-17)., (© 2024. The Author(s).)

Background: TAF1L may play an important role in the occurrence and development of gastric cancer (GC), but the correlation between the expression of TAF1L and the clinicopathological factors and prognosis of GC is still unclear., Methods: A total of 1053 GC patients in Zhejiang Cancer Hospital between January 1st, 2018 to December 31th, 2019 were screened. Finally, 120 patients met the inclusion criteria. TAF1L expression was detected by immunohistochemistry, and the correlations of TAF1L in clinicopathological characteristics and prognosis were analyzed. TCGA GC dataset was used to perform further bioinformatics analysis., Results: In this study, TAF1L expression was evaluated in 120 clinical samples of GC. TAF1L expression was higher in tumor tissues and was associated with tumor differentiation (p = 0.046), signet-ring cells (p = 0.043), dMMR status (p = 0.011), lympho-vascular invasion (p = 0.038), and neural invasion (p = 0.005) in our cohort. Cases with high expression of TAF1L presented worse mean OS than those with low expression (40.3 months vs. 51.8 months, p = 0.019), and the difference was also significant in HER2-positive cases (20.9 months vs. 51.2 months, p = 0.007) as well as pMMR cases (38.8 months vs. 51.6 months, p = 0.006). Multivariate Cox regression analysis showed that TAF1L (HR = 2.044, 95%CI = 1.007-4.147, p = 0.048) and HER2 status (HR = 2.383, 95%CI = 1.087-5.222, p = 0.030) were independent prognosis factors of these patients. In subgroup analysis, TAF1L was the independent prognostic risk factor in HER2-positive patients (HR = 6.736, 95%CI = 1.373-33.032, p = 0.019). and pMMR patients (HR = 2.291, 95%CI = 1.126-4.660, p = 0.022). Besides, HER2 status was the independent prognostic risk factor in TAF1L-H patients (HR = 4.832, 95%CI = 1.908-12.239, p = 0.001). TCGA dataset also indicated the higher expression of TAF1L in tumors than normal tissues (p < 0.001). High TAF1L expression is linked to worse survival in MSS (11.0 months vs. 35.0 months, p = 0.0046) groups, and is negatively associated with overall survival in HER2-positive cases (24.0 months vs. 57.0 months, p = 0.0039)., Conclusion: TAF1L is closely related to the occurrence and development of GC. Our results suggested that TAF1L is a significant biomarker for predicting prognosis of GC and may play an important role in immunotherapy and targeted therapy., Competing Interests: Declarations. Ethics approval: This study was performed in line with the principles of the Declaration of Helsinki. Approval was granted by Zhejiang Cancer Hospital Ethics Committee (No. IRB-2022-691). Human Ethics and Consent to Participate declarations: not applicable. Informed consent was waived by the Ethics Committee of the Zhejiang Cancer Hospital because of the retrospective study of this project. Consent for publication: Not Applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Clear cell renal cell carcinoma (ccRCC) is a malignant tumor needs more effective treatments. TATA box-binding protein-associated factor RNA polymerase I subunit D (TAF1D) is a member of the selective factor 1 complex and functions in RNA polymerase I-dependent transcription. Higher TAF1D expression was found in ccRCC tumor tissues and indicated worse survival. Our study aimed to investigate the therapeutic potential of TAF1D in ccRCC. The proliferation and migration of ccRCC cells were significantly inhibited after TAF1D knockdown, while TAF1D overexpressing had opposite effects. Moreover, TAF1D knockdown induced cells to undergo G 0 /G 1 cell cycle arrest and blockade of the epithelial-mesenchymal transition (EMT) process. Mechanistically, TAF1D affect the cell cycle and EMT through the PI3K/AKT/mTOR signaling pathway, thereby promoting the proliferation and metastasis of ccRCC cells in vivo and in vitro. The inhibitory effect of TAF1D knockdown could be reverted by the AKT activator SC79 in ccRCC cells, confirming this mechanism. Besides, TAF1D knockdown in ccRCC cells had a sensitizing effect on sunitinib and enhanced tumor cell inhibiting induced by sunitinib. In conclusion, TAF1D may be a promising target for the treatment of ccRCC and for overcoming sunitinib resistance., 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 © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Cyclophilin A (CypA) is a peptidyl-prolyl isomerase that participates in multiple cancer events, but the molecular mechanisms of abnormal expression and regulation of CypA in ovarian cancer (OC) have never been considered. This study identifies CypA as a key driver of epithelial-mesenchymal transition (EMT) in ovarian cancer and explores the mechanisms that underly this process. We show that CypA is upregulated in tissues and serum of ovarian cancer patients and that CypA overexpression correlates with poor prognosis. CypA facilitates tumor growth and metastasis in vivo in subcutaneous tumor xenograft and abdominal metastatic models, and in vitro studies suggest a mechanism, showing that CypA accelerates ovarian cancer cell epithelial-mesenchymal transition by activating a PI3K/AKT signaling pathway. Mechanistic studies showed that STAT5A binds pri-miR-514a-3p and inhibits its activity, whereas miR-514a-3p directly binds to the 3'-UTR of CypA to suppress its expression, resulting in STAT5A promoting the expression of CypA, forming the STAT5A/miR-514a-3p/CypA axis. Furthermore, immunoprecipitates and mass spectrometry analysis identifies a CypA interaction with TAF15 that stabilizes TAF15 by suppressing its proteasome degradation and promotes its entry into the nucleus. While STAT5A is positively regulated by TAF15. Our findings identify a novel feedback loop for CypA that drives EMT and ovarian tumor growth and metastasis via a TAF15/STAT5A/miR-514a-3p pathway in ovarian cancer and facilitates the release of CypA into the extracellular, which provides a promising therapeutic target for OC treatment and a diagnostic biomarker., Competing Interests: Competing interests: The authors declare no competing interests. Ethical approval: The informed consent was obtained from patients and the study was approved by the Medical Ethics Committee of the Affiliated Hospital of Xuzhou Medical University (XYFY2022-KL439-01). All the research was performed in accordance with government policies and the Helsinki declaration. The animal experiments were approved by the Institutional Animal Care and Use Committee of Xuzhou Medical University (202005A009)., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

As the hybrid between cattle and yak, cattleyak is a typical male sterile mammal, and the underlying mechanism for its spermatogenic arrest is still unclear. In this study, the coding region of cattleyak TAF4B gene was cloned by RT-PCR and analysed by bioinformatics. To investigate the effects of TAF4B on cellular proliferation and differentiation, an expression vector was generated and introduced into undifferentiated spermatogonia (UDSPG) of cattleyak. The results showed that the protein encoded by TAF4B did not contain the signal peptide sequence. The expression level of TAF4B in UDSPG of cattleyak was lower than that in yak, while the overexpression of TAF4B in cattleyak promoted the proliferation activity of cattleyak UDSPG. Meanwhile, the expression of proliferation and meiosis-related genes was increased but the differentiation-related genes were decreased. Therefore, the aberrant expression of TAF4B in cattleyak UDSPG possibly impaired its proliferation and differentiation equilibrium and decreased its growth potentiality, thereby reducing the quantity of UDSPG and affecting spermatogenesis. This study provided a potential approach for further elucidation of the mechanism of spermatogenesis arrest and provided a new idea for solving the problem of male infertility in cattleyak., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

G-quadruplexes (G4s) are non-canonical nucleic acid structures that form in guanine (G)-rich genomic regions. X-linked dystonia parkinsonism (XDP) is an inherited neurodegenerative disease in which a SINE-VNTR-Alu (SVA) retrotransposon, characterised by amplification of a G-rich repeat, is inserted into the coding sequence of TAF1, a key partner of RNA polymerase II. XDP SVA alters TAF1 expression, but the cause of this outcome in XDP remains unknown. To assess whether G4s form in XDP SVA and affect TAF1 expression, we first characterised bioinformatically predicted XDP SVA G4s in vitro. We next showed that highly stable G4s can form and stop polymerase amplification at the SVA region from patient-derived fibroblasts and neural progenitor cells. Using chromatin immunoprecipitazion (ChIP) with an anti-G4 antibody coupled to sequencing or quantitative PCR, we showed that XDP SVA G4s are folded even when embedded in a chromatin context in patient-derived cells. Using the G4 ligands BRACO-19 and quarfloxin and total RNA-sequencing analysis, we showed that stabilisation of the XDP SVA G4s reduces TAF1 transcripts downstream and around the SVA, and increases upstream transcripts, while destabilisation using the G4 unfolder PhpC increases TAF1 transcripts. Our data indicate that G4 formation in the XDP SVA is a major cause of aberrant TAF1 expression, opening the way for the development of strategies to unfold G4s and potentially target the disease., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The recognition of core promoter sequences by TFIID is the first step in RNA polymerase II (Pol II) transcription initiation. Metazoan holo-TFIID is a trilobular complex, composed of the TATA binding protein (TBP) and 13 TBP-associated factors (TAFs). Why and how TAFs are necessary for the formation of TFIID domains and how they contribute to transcription initiation remain unclear. Inducible TAF7 or TAF10 depletion, followed by comprehensive analysis of TFIID subcomplex formation, chromatin binding, and nascent transcription in mouse embryonic stem cells, result in the formation of a TAF7-lacking TFIID or a minimal core-TFIID complex, respectively. These partial complexes support TBP recruitment at promoters and nascent Pol II transcription at most genes early after depletion, but importantly, TAF10 is necessary for efficient Pol II pausing. We show that partially assembled TFIID complexes can sustain Pol II transcription initiation but cannot replace holo-TFIID over several cell divisions and/or development., Competing Interests: Declaration of interests The authors declare that they have no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

TAF1A, a gene encoding a TATA-box binding protein involved in ribosomal RNA synthesis, is a candidate gene for pediatric cardiomyopathy as biallelic TAF1A variants were reported in two families with affected individuals. Here, we report a third family with two siblings who presented with infantile restrictive cardiomyopathy and carried biallelic missense variants in TAF1A (NM&#95;001201536.1:c.1021G>A p.(Gly341Arg) and c.781A>C p.(Thr261Pro)). Additional shared clinical features in the siblings included feeding intolerance, congenital leukoencephalopathy, ventriculomegaly and concern for primary immunodeficiency. The first-born sibling passed away at 6 months of age due to complications of hemophagocytic lymphohistiocytosis (HLH) whereas the second sibling underwent cardiac transplantation at 1 year of age and is currently well. We compare the clinical and molecular features of all the TAF1A associated cardiomyopathy cases. Our study adds evidence for the gene-disease association of TAF1A with autosomal recessive pediatric cardiomyopathy., 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 © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

SVA (SINE (short interspersed nuclear element)-VNTR (variable number of tandem repeats)-Alu) retrotransposons remain active in humans and contribute to individual genetic variation. Polymorphic SVA alleles harbor gene regulatory potential and can cause genetic disease. However, how SVA insertions are controlled and functionally impact human disease is unknown. Here we dissect the epigenetic regulation and influence of SVAs in cellular models of X-linked dystonia parkinsonism (XDP), a neurodegenerative disorder caused by an SVA insertion at the TAF1 locus. We demonstrate that the KRAB zinc finger protein ZNF91 establishes H3K9me3 and DNA methylation over SVAs, including polymorphic alleles, in human neural progenitor cells. The resulting mini-heterochromatin domains attenuate the cis-regulatory impact of SVAs. This is critical for XDP pathology; removal of local heterochromatin severely aggravates the XDP molecular phenotype, resulting in increased TAF1 intron retention and reduced expression. Our results provide unique mechanistic insights into how human polymorphic transposon insertions are recognized and how their regulatory impact is constrained by an innate epigenetic defense system., (© 2024. The Author(s).)

Abnormalities in coagulation and fibrinolytic status have been demonstrated to be relevant to inflammatory bowel disease. Nevertheless, there is no study to methodically examine the role of the coagulation and fibrinolysis-related genes in the diagnosis of ulcerative colitis (UC). UC-related datasets (GSE169568 and GSE94648) were originated from the Gene Expression Omnibus database. The biomarkers related to coagulation and fibrinolysis were identified through combining differentially expressed analysis and machine learning algorithms. Moreover, Gene Set Enrichment Analysis and immune analysis were carried out. A total of 4 biomarkers (MAP2K1, CREBBP, TAF1, and HP) were identified, and biomarkers were markedly enriched in pathways related to immunity, such as T-cell receptor signaling pathway, primary immunodeficiency, chemokine signaling pathway, etc. In total, the infiltrating abundance of 4 immune cells between UC and control was markedly different, namely eosinophils, macrophage M0, resting mast cells, and regulatory T cells. And all biomarkers were significantly relevant to eosinophils. Our findings detected 4 coagulation and fibrinolysis-related biomarkers (MAP2K1, CREBBP, TAF1, and HP) for UC, which contributed to the advancement of UC for further clinical investigation., Competing Interests: The authors have no funding and conflicts of interest to disclose., (Copyright © 2024 the Author(s). Published by Wolters Kluwer Health, Inc.)

SMA (spinal muscular atrophy) is an autosomal recessive neuromuscular disease that causes muscle atrophy and weakness. SMA is diagnosed by a homozygous deletion in exon 7 of the SMN1 gene. However, mutations in genes located in the SMA region, such as SMN2 , NAIP, SERF1, and GTF2H2, may also contribute to the severity of the disease. Within our study's scope, 58 SMA patients who applied in 2018-2021 and 40 healthy controls were analyzed. The study retrospectively included the SMN1 and SMN2 copy numbers previously determined by the MLPA method. Then, NAIP gene analyses with the multiplex PCR method and GTF2H2 gene analyses with the RFLP method were performed. There was a significant correlation ( p  = 0.00001) between SMN2 copy numbers and SMA subtypes. Also, the NAIP gene ( p  = 0.01) and the GTF2H2 gene ( p  = 0.0049) revealed a significant difference between healthy and SMA subjects, whereas the SMA subtypes indicated no significant differences. We detected a significant correlation between clinical subtypes and HFMSE scores in 32 pediatric SMA patients compared ( p  = 0.01). While pediatric patients with GTF2H2 deletions demonstrated higher motor functions, and those with NAIP deletions demonstrated lower motor functions. In this study, we examined the relationship between NAIP and GTF2H2 , called SMN region modifier genes, and the clinical severity of the disease in Turkish SMA patients. Despite its small scale, this research will benefit future investigations into the pathogenesis of SMA disease.

X-linked dystonia-parkinsonism (XDP) is a severe neurodegenerative disorder resulting from an inherited intronic SINE-Alu-VNTR (SVA) retrotransposon in the TAF1 gene that causes dysregulation of TAF1 transcription. The specific mechanism underlying this dysregulation remains unclear, but it is hypothesized to involve the formation of G-quadruplexes (G4) structures within the XDP-SVA that impede transcription. In this study, we show that ZNF91, a critical repressor of SVA retrotransposons, specifically binds to G4-forming DNA sequences. Further, we found that genetic deletion of ZNF91 exacerbates the molecular phenotype associated with the XDP-SVA insertion in patient cells, while no difference was observed when ZNF91 was deleted from isogenic control cells. Additionally, we observed a significant age-related reduction in ZNF91 expression in whole blood and brain, indicating a progressive loss of repression of the XDP-SVA in XDP. These findings indicate that ZNF91 plays a crucial role in controlling the molecular phenotype associated with XDP. Since ZNF91 binds to G4-forming DNA sequences in SVAs, this suggests that interactions between ZNF91 and G4-forming sequences in the XDP-SVA minimize the severity of the molecular phenotype. Our results showing that ZNF91 expression levels significantly decrease with age provide a potential explanation for the age-related progressive neurodegenerative character of XDP. Collectively, our study provides important insights into the protective role of ZNF91 in XDP pathogenesis and suggests that restoring ZNF91 expression, destabilization of G4s, or targeted repression of the XDP-SVA could be future therapeutic strategies to prevent or treat XDP., Competing Interests: Competing interests statement:The authors declare no competing interest.

Mammalian RNA polymerase II preinitiation complexes assemble adjacent to a nucleosome whose proximal edge (NPE) is typically 40 to 50 bp downstream of the transcription start site. At active promoters, that +1 nucleosome is universally modified by trimethylation on lysine 4 of histone H3 (H3K4me3). The Pol II preinitiation complex only extends 35 bp beyond the transcription start site, but nucleosomal templates with an NPE at +51 are nearly inactive in vitro with promoters that lack a TATA element and thus depend on TFIID for promoter recognition. Significantly, this inhibition is relieved when the +1 nucleosome contains H3K4me3, which can interact with TFIID subunits. Here, we show that H3K4me3 templates with both TATA and TATA-less promoters are active with +35 NPEs when transcription is driven by TFIID. Templates with +20 NPE are also active but at reduced levels compared to +35 and +51 NPEs, consistent with a general inhibition of promoter function when the proximal nucleosome encroaches on the preinitiation complex. Remarkably, dinucleosome templates support transcription when H3K4me3 is only present in the distal nucleosome, suggesting that TFIID-H3K4me3 interaction does not require modification of the +1 nucleosome. Transcription reactions performed with an alternative protocol retaining most nuclear factors results primarily in early termination, with a minority of complexes successfully traversing the first nucleosome. In such reactions, the +1 nucleosome does not substantially affect the level of termination even with an NPE of +20, indicating that a nucleosome barrier is not a major driver of early termination by Pol II., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

CRISPR-Cas technology has transformed functional genomics, yet understanding of how individual exons differentially shape cellular phenotypes remains limited. Here, we optimized and conducted massively parallel exon deletion and splice-site mutation screens in human cell lines to identify exons that regulate cellular fitness. Fitness-promoting exons are prevalent in essential and highly expressed genes and commonly overlap with protein domains and interaction interfaces. Conversely, fitness-suppressing exons are enriched in nonessential genes, exhibiting lower inclusion levels, and overlap with intrinsically disordered regions and disease-associated mutations. In-depth mechanistic investigation of the screen-hit TAF5 alternative exon-8 revealed that its inclusion is required for assembly of the TFIID general transcription initiation complex, thereby regulating global gene expression output. Collectively, our orthogonal exon perturbation screens established a comprehensive repository of phenotypically important exons and uncovered regulatory mechanisms governing cellular fitness and gene expression., Competing Interests: Declaration of interests The authors declare no competing interests., (Published by Elsevier Inc.)

Early embryonic development is a finely orchestrated process that requires precise regulation of gene expression coordinated with morphogenetic events. TATA-box binding protein-associated factors (TAFs), integral components of transcription initiation coactivators like TFIID and SAGA, play a crucial role in this intricate process. Here we show that disruptions in TAF5, TAF12 and TAF13 individually lead to embryonic lethality in the mouse, resulting in overlapping yet distinct phenotypes. Taf5 and Taf12 mutant embryos exhibited a failure to implant post-blastocyst formation, and Taf5 mutants have aberrant lineage specification within the inner cell mass. In contrast, Taf13 mutant embryos successfully implant and form egg-cylinder stages but fail to initiate gastrulation. Strikingly, we observed a depletion of pluripotency factors in TAF13-deficient embryos, including OCT4, NANOG and SOX2, highlighting an indispensable role of TAF13 in maintaining pluripotency. Transcriptomic analysis revealed distinct gene targets affected by the loss of TAF5, TAF12 and TAF13. Thus, we propose that TAF5, TAF12 and TAF13 convey locus specificity to the TFIID complex throughout the mouse genome., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: X-Linked dystonia-parkinsonism (XDP) is an adult-onset neurodegenerative disorder characterized by rapidly progressive dystonia and parkinsonism. Mosaic Divergent Repeat Interruptions affecting motif Length and Sequence (mDRILS) were recently found within the TAF1 SVA repeat tract and were shown to associate with repeat stability and age at onset in XDP, specifically the AGGG [5'-SINE-VNTR-Alu(AGAGGG) 2 AGGG(AGAGGG) n ] mDRILS., Objective: This study aimed to investigate the stability of mDRILS frequencies and stability of (AGAGGG) n repeat length during transmission in parent-offspring pairs., Methods: Fifty-six families (n = 130) were investigated for generational transmission of repeat length and mDRILS. The mDRILS stability of 16 individuals was assessed at two sampling points 1 year apart. DNA was sequenced with long-read technologies after long-range polymerase chain reaction amplification of the TAF1 SVA. Repeat number and mDRILS were detected with Noise-Cancelling Repeat Finder (NCRF)., Results: When comparing the repeat domain, 51 of 65 children had either contractions or expansions of the repeat length. The AGGG frequency remained stable across generations at 0.074 (IQR: 0.069-0.078) (z = -0.526; P = 0.599). However, the median AGGG frequency in children with an expansion (0.072 [IQR: 0.066-0.076]) was lower compared with children with retention or contraction (0.080 [IQR: 0.073-0.083]) (z = -0.007; P = 0.003). In a logistic regression model, the AGGG frequency predicted the outcome of either expansion or retention/contraction when including repeat number and sex as covariates (β = 80.7; z-score = 2.63; P = 0.0085). The AGGG frequency varied slightly over 1 year (0.070 [IQR: 0.063-0.080] to 0.073 [IQR: 0.069-0.078])., Conclusions: Our results show that a higher AGGG frequency may stabilize repeats across generations. This highlights the importance of further investigating mDRILS as a disease-modifying factor with generational differences. © 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society., (© 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.)

The TATA box-binding protein-associated factor 1 (TAF1) is a ubiquitously expressed protein and the largest subunit of the basal transcription factor TFIID, which plays a key role in initiation of RNA polymerase II-dependent transcription. TAF1 missense variants in human males cause X-linked intellectual disability, a neurodevelopmental disorder, and TAF1 is dysregulated in X-linked dystonia-parkinsonism, a neurodegenerative disorder. However, this field has lacked a genetic mouse model of TAF1 disease to explore its mechanism in mammals and treatments. Here, we generated and validated a conditional cre-lox allele and the first ubiquitous Taf1 knockout mouse. We discovered that Taf1 deletion in male mice was embryonically lethal, which may explain why no null variants have been identified in humans. In the brains of Taf1 heterozygous female mice, no differences were found in gross structure, overall expression and protein localisation, suggesting extreme skewed X inactivation towards the non-mutant chromosome. Nevertheless, these female mice exhibited a significant increase in weight, weight with age, and reduced movement, suggesting that a small subset of neurons was negatively impacted by Taf1 loss. Finally, this new mouse model may be a future platform for the development of TAF1 disease therapeutics., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

The NuA3 complex is a major regulator of gene transcription and the cell cycle in yeast. Five core subunits are required for complex assembly and function, but it remains unclear how these subunits interact to form the complex. Here, we report that the Taf14 subunit of the NuA3 complex binds to two other subunits of the complex, Yng1 and Sas3, and describe the molecular mechanism by which the extra-terminal domain of Taf14 recognizes the conserved motif present in Yng1 and Sas3. Structural, biochemical, and mutational analyses show that two motifs are sandwiched between the two extra-terminal domains of Taf14. The head-to-toe dimeric complex enhances the DNA binding activity of Taf14, and the formation of the hetero-dimer involving the motifs of Yng1 and Sas3 is driven by sequence complementarity. In vivo assays in yeast demonstrate that the interactions of Taf14 with both Sas3 and Yng1 are required for proper function of the NuA3 complex in gene transcription and DNA repair. Our findings suggest a potential basis for the assembly of three core subunits of the NuA3 complex, Taf14, Yng1 and Sas3., (© 2024. The Author(s).)

Development of cancer therapies targeting chromatin modifiers and transcriptional regulatory factors is rapidly expanding to include new targets and novel targeting strategies. At the same time, basic molecular research continues to refine our understanding of the epigenetic mechanisms regulating transcription, gene expression, and oncogenesis. This mini-review focuses on cancer therapies targeting the chromatin-associated factors that recognize histone lysine acetylation. Recently reported safety and efficacy are discussed for inhibitors targeting the bromodomains of bromodomain and extraterminal domain (BET) family proteins. In light of recent results indicating that the transcriptional regulator BRD4-PTEFb can function independently of BRD4's bromodomains, the clinical trial performance of these BET inhibitors is placed in a broader context of existing and potential strategies for targeting BRD4-PTEFb. Recently developed therapies targeting bromodomain-containing factors within the SWI/SNF (BAF) family of chromatin remodeling complexes are discussed, as is the potential for targeting the bromodomain-containing transcription factor TAF1 and the YEATS acetylrecognition domain-containing factor GAS41. Recent findings regarding the selectivity and combinatorial specificity of acetylrecognition are highlighted. In conclusion, the potential for further development is discussed with a focus on proximity-based therapies targeting this class of epigenetic factors., 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 © 2024 Elsevier Ltd. All rights reserved.)

Thyroid dysgenesis (TD) is the common pathogenic mechanism of congenital hypothyroidism (CH). In addition, known pathogenic genes are limited to those that are directly involved in thyroid development. To identify additional candidate pathogenetic genes, we performed forward genetic screening for TD in zebrafish, followed by positional cloning. The candidate gene was confirmed in vitro using the Nthy-ori 3.1 cell line and in vivo using a zebrafish model organism. We obtained a novel zebrafish line with thyroid dysgenesis and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 ( taf1 ) by positional cloning. Further molecular studies revealed that taf1 was needed for the proliferation of thyroid follicular cells by binding to the NOTCH1 promoter region. Knockdown of TAF1 impaired the proliferation and maturation of thyroid cells, thereby leading to thyroid dysplasia. This study showed that TAF1 promoted Notch signaling and that this association played a pivotal role in thyroid development. NEW & NOTEWORTHY In our study, we obtained a novel zebrafish line with thyroid dysgenesis (TD) and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 ( taf1 ). Further researches revealed that taf1 was required for thyroid follicular cells by binding to the NOTCH1 promoter region. Our findings revealed a novel role of TAF1 in thyroid morphogenesis.

SET domain containing 7(SETD7), a member of histone methyltransferases, is abnormally expressed in multiple tumor types. However, the biological function and underlying molecular mechanism of SETD7 in clear cell renal cell carcinoma (ccRCC) remain unclear. Here, we explored the biological effects of SETD7-TAF7-CCNA2 axis on proliferation and metastasis in ccRCC. We identified both SETD7 and TAF7 were up-regulated and significantly promoted the proliferation and migration of ccRCC cells. Concurrently, there was a significant positive correlation between the expression of SETD7 and TAF7, and the two were colocalized in the nucleus. Mechanistically, SETD7 methylates TAF7 at K5 and K300 sites, resulting in the deubiquitination and stabilization of TAF7. Furthermore, re-expression of TAF7 could partially restore SETD7 knockdown inhibited ccRCC cells proliferation and migration. In addition, TAF7 transcriptionally activated to drive the expression of cyclin A2 (CCNA2). And more importantly, the methylation of TAF7 at K5 and K300 sites exhibited higher transcriptional activity of CCNA2, which promotes formation and progression of ccRCC. Our findings reveal a unique mechanism that SETD7 mediated TAF7 methylation in regulating transcriptional activation of CCNA2 in ccRCC progression and provide a basis for developing effective therapeutic strategies by targeting members of SETD7-TAF7-CCNA2 axis., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Mechanisms of functional cross-talk between global transcriptional repression and efficient DNA damage repair during genotoxic stress are poorly known. In this study, using human AF9 as representative of Super Elongation Complex (SEC) components, we delineate detailed mechanisms of these processes. Mechanistically, we describe that Poly-Serine domain-mediated oligomerization is pre-requisite for AF9 YEATS domain-mediated TFIID interaction-dependent SEC recruitment at the promoter-proximal region for release of paused RNA polymerase II. Interestingly, during genotoxic stress, CaMKII-mediated phosphorylation-dependent nuclear export of AF9-specific deacetylase HDAC5 enhances concomitant PCAF-mediated acetylation of K339 residue. This causes monomerization of AF9 and reduces TFIID interaction for transcriptional downregulation. Furthermore, the K339 acetylation-dependent enhanced AF9-DNA-PKc interaction leads to phosphorylation at S395 residue which reduces AF9-SEC interaction resulting in transcriptional downregulation and efficient repair of DNA damage. After repair, nuclear re-entry of HDAC5 reduces AF9 acetylation and restores its TFIID and SEC interaction to restart transcription., (© 2024. The Author(s).)

Objective: To analyze the clinical features and genetic etiology of 17 Chinese pedigrees affected with X-linked intellectual disability (XLID)., Methods: Seventeen pedigrees affected with unexplained intellectual disability which had presented at Henan Provincial People's Hospital from May 2021 to May 2023 were selected as the study subjects. Clinical data of the probands and their pedigree members were collected. Trio-whole exome sequencing (Trio-WES), Sanger sequencing and X chromosome inactivation (XCI) analysis were carried out. Pathogenicity of candidate variants was predicted based on the guidelines from the American College of Medical Genetics and Genomics and co-segregation analysis., Results: The 17 probands, including 9 males and 8 females with an age ranging from 0.6 to 8 years old, had all shown mental retardation and developmental delay. Fourteen variants were detected by genetic testing, which included 4 pathogenic variants (MECP2: c.502C>T, MECP2: c.916C>T/c.806delG, IQSEC2: c.1417G>T), 4 likely pathogenic variants (MECP2: c.1157&#95;1197del/c.925C>T, KDM5C: c.2128A>T, SLC6A8: c.1631C>T) and 6 variants of uncertain significance (KLHL15: c.26G>C, PAK3: c.970A>G/c.1520G>A, GRIA3: c.2153C>G, TAF1: c.2233T>G, HUWE1: c.10301T>A). The PAK3: c.970A>G, GRIA3: c.2153C>G and TAF1: c.2233T>G variants were considered as the genetic etiology for pedigrees 12, 14 and 15 by co-segregation analysis, respectively. The proband of pedigree 13 was found to have non-random XCI (81:19). Therefore, the PAK3: c.1520G>A variant may underlie its pathogenesis., Conclusion: Trio-WES has attained genetic diagnosis for the 17 XLID pedigrees. Sanger sequencing and XCI assay can provide auxiliary tests for the diagnosis of XLID.

X-linked dystonia-parkinsonism (XDP) is a rare neurodegenerative disease endemic to the Philippines. The genetic cause for XDP is an insertion of a SINE-VNTR-Alu (SVA)-type retrotransposon within intron 32 of TATA-binding protein associated factor 1 (TAF1) that causes an alteration of TAF1 splicing, partial intron retention, and decreased transcription. Although TAF1 is expressed in all organs, medium spiny neurons (MSNs) within the striatum are one of the cell types most affected in XDP. To define how mutations in the TAF1 gene lead to MSN vulnerability, we carried out a proteomic analysis of human XDP patient-derived neural stem cells (NSCs) and MSNs derived from induced pluripotent stem cells. NSCs and MSNs were grown in parallel and subjected to quantitative proteomic analysis in data-independent acquisition mode on the Orbitrap Eclipse Tribrid mass spectrometer. Subsequent functional enrichment analysis demonstrated that neurodegenerative disease-related pathways, such as Huntington's disease, spinocerebellar ataxia, cellular senescence, mitochondrial function and RNA binding metabolism, were highly represented. We used weighted coexpression network analysis (WGCNA) of the NSC and MSN proteomic data set to uncover disease-driving network modules. Three of the modules significantly correlated with XDP genotype when compared to the non-affected control and were enriched for DNA helicase and nuclear chromatin assembly, mitochondrial disassembly, RNA location and mRNA processing. Consistent with aberrant mRNA processing, we found splicing and intron retention of TAF1 intron 32 in XDP MSN. We also identified TAF1 as one of the top enriched transcription factors, along with YY1, ATF2, USF1 and MYC. Notably, YY1 has been implicated in genetic forms of dystonia. Overall, our proteomic data set constitutes a valuable resource to understand mechanisms relevant to TAF1 dysregulation and to identify new therapeutic targets for XDP., Competing Interests: Declaration of Competing Interest The authors have no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Male germ cell development is dependent on the orchestrated regulation of gene networks. TATA-box binding protein associated factors (TAFs) facilitate interactions of TATA-binding protein with the TATA element, which is known to coordinate gene transcription during organogenesis. TAF7 like (Taf7l) is situated on the X chromosome and has been implicated in testis development. We examined the biology of TAF7L in testis development using the rat. Taf7l was prominently expressed in preleptotene to leptotene spermatocytes. To study the impact of TAF7L on the testis we generated a global loss-of-function rat model using CRISPR/Cas9 genome editing. Exon 3 of the Taf7l gene was targeted. A founder was generated possessing a 110 bp deletion within the Taf7l locus, which resulted in a frameshift and the premature appearance of a stop codon. The mutation was effectively transmitted through the germline. Deficits in TAF7L did not adversely affect pregnancy or postnatal survival. However, the Taf7l disruption resulted in male infertility due to compromised testis development and failed sperm production. Mutant germ cells suffer meiotic arrest at late zygotene/early pachynema stages, with defects in sex body formation. This testis phenotype was more pronounced than previously described for the subfertile Taf7l null mouse. We conclude that TAF7L is essential for male germ cell development in the rat., (© 2023 Federation of American Societies for Experimental Biology.)

In eukaryotes, TATA-binding protein (TBP) occupancy of the core promoter globally correlates with transcriptional activity of class II genes. Elucidating how TBP is delivered to the TATA box or TATA-like element is crucial to understand the mechanisms of transcriptional regulation. A previous study demonstrated that the inhibitory DNA binding (IDB) surface of human TBP plays an indispensable role during the two-step formation of the TBP-TATA complex, first assuming an unstable and unbent intermediate conformation, and subsequently converting slowly to a stable and bent conformation. The DNA binding property of TBP is altered by physical contact of this surface with TBP regulators. In the present study, we examined whether the interaction between Taf1 N-terminal domain 2 (TAND2) and the IDB surface affected DNA binding property of yeast TBP by exploiting TAND2-fused TBP derivatives. TAND2 promoted formation of two distinct types of TBP-TATA complexes, which we arbitrarily designated as complex I and II. While complex I was stable and similar to the well-characterized original TBP-TATA complex, complex II was unstable and moved along DNA. Removal of TAND2 from TBP after complex formation revealed that continuous contact of TAND2 with the IDB surface was required for formation of complex II but not complex I. Further, TFIIA could be incorporated into the complex of TAND2-fused TBP and the TATA box, which was dependent on the amino-terminal non-conserved region of TBP, implying that this region could facilitate the exchange between TAND2 and TFIIA on the IDB surface. Collectively, these findings provide novel insights into the mechanism by which TBP is relieved from the interaction with TAND to bind the TATA box or TATA-like element within promoter-bound TFIID., 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 Elsevier B.V. All rights reserved.)

Binding of general transcription factors TFIID and TFIIA to basal promoters is rate-limiting for transcriptional initiation of eukaryotic protein-coding genes. Consequently, activator proteins interacting with subunits of TFIID and/or TFIIA can drastically increase the rate of initiation events. Yeast transcriptional activator Ino2 interacts with several Taf subunits of TFIID, among them the multifunctional Taf1 protein. In contrast to mammalian Taf1, yeast Taf1 lacks bromodomains which are instead encoded by separate proteins Bdf1 and Bdf2. In this work, we show that Bdf1 not only binds to acetylated histone H4 but can also be recruited by Ino2 and unrelated activators such as Gal4, Rap1, Leu3 and Flo8. An activator-binding domain was mapped in the N-terminus of Bdf1. Subunits Toa1 and Toa2 of yeast TFIIA directly contact sequences of basal promoters and TFIID subunit TBP but may also mediate the influence of activators. Indeed, Ino2 efficiently binds to two separate structural domains of Toa1, specifically with its N-terminal four-helix bundle structure required for dimerization with Toa2 and its C-terminal β-barrel domain contacting TBP and sequences of the TATA element. These findings complete the functional analysis of yeast general transcription factors Bdf1 and Toa1 and identify them as targets of activator proteins., (© 2023. The Author(s).)

The RNA polymerase II (Pol II) pre-initiation complex (PIC) is a critical node in eukaryotic transcription regulation, and its formation is the major rate-limiting step in transcriptional activation. Diverse cellular signals borne by transcriptional activators converge on this large, multiprotein assembly and are transduced via intermediary factors termed coactivators. Cryogenic electron microscopy, multi-omics and single-molecule approaches have recently offered unprecedented insights into both the structure and cellular functions of the PIC and two key PIC-associated coactivators, Mediator and TFIID. Here, we review advances in our understanding of how Mediator and TFIID interact with activators and affect PIC formation and function. We also discuss how their functions are influenced by their chromatin environment and selected cofactors. We consider how, through its multifarious interactions and functionalities, a Mediator-containing and TFIID-containing PIC can yield an integrated signal processing system with the flexibility to determine the unique temporal and spatial expression pattern of a given gene., (© 2023. Springer Nature Limited.)

Core promoters are sites where transcriptional regulatory inputs of a gene are integrated to direct the assembly of the preinitiation complex (PIC) and RNA polymerase II (Pol II) transcription output. Until now, core promoter functions have been investigated by distinct methods, including Pol II transcription initiation site mappings and structural characterization of PICs on distinct promoters. Here, we bring together these previously unconnected observations and hypothesize how, on metazoan TATA promoters, the precisely structured building up of transcription factor (TF) IID-based PICs results in sharp transcription start site (TSS) selection; or, in contrast, how the less strictly controlled positioning of the TATA-less promoter DNA relative to TFIID-core PIC components results in alternative broad TSS selections by Pol II., Competing Interests: Declaration of interests No interests are declared., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Taf14 is a subunit of multiple fundamental complexes implicated in transcriptional regulation and DNA damage repair in yeast cells. Here, we investigate the association of Taf14 with the consensus sequence present in other subunits of these complexes and describe the mechanistic features that affect this association. We demonstrate that the precise molecular mechanisms and biological outcomes underlying the Taf14 interactions depend on the accessibility of binding interfaces, the ability to recognize other ligands, and a degree of sensitivity to temperature and chemical and osmotic stresses. Our findings aid in a better understanding of how the distribution of Taf14 among the complexes is mediated., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Tatiana G Kutateladze reports financial support was provided by National Institutes of Health. Tatiana G Kutateladze reports a relationship with NIH that includes: funding grants., (Copyright © 2023. Published by Elsevier B.V.)

Intellectual disability is a neurodevelopmental disorder that affects 2-3% of the general population. Syndromic forms of intellectual disability frequently have a genetic basis and are often accompanied by additional developmental anomalies. Pathogenic variants in components of TATA-binding protein associated factors (TAFs) have recently been identified in a subset of patients with intellectual disability, craniofacial hypoplasia, and congenital heart disease. This syndrome has been termed as a TAFopathy and includes mutations in TATA binding protein (TBP), TAF1, TAF2, and TAF6. The underlying mechanism by which TAFopathies give rise to neurodevelopmental, craniofacial, and cardiac abnormalities remains to be defined. Through a forward genetic screen in zebrafish, we have recovered a recessive mutant phenotype characterized by craniofacial hypoplasia, ventricular hypoplasia, heart failure at 96 h post-fertilization and lethality, and show it is caused by a nonsense mutation in taf5. CRISPR/CAS9 mediated gene editing revealed that these defects where phenocopied by mutations in taf1 and taf5. Mechanistically, taf5-/- zebrafish displayed misregulation in metabolic gene expression and metabolism as evidenced by RNA sequencing, respiration assays, and metabolite studies. Collectively, these findings suggest that the TAF complex may contribute to neurologic, craniofacial, and cardiac development through regulation of metabolism., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

X-linked dystonia-parkinsonism (XDP) is a neurodegenerative disease caused by a retrotransposon insertion in intron 32 of the TAF1 gene. This insertion causes mis-splicing of intron 32 (TAF1-32i) and reduced TAF1 levels. TAF1-32i transcript is unique to XDP patient cells and can be detected in their extracellular vesicles (EVs). We engrafted patient and control iPSC-derived neural progenitor cells (hNPCs) into the striatum of mice. To track TAF1-32i transcript spread by EVs, we transduced the brain-implanted hNPCs with a lentiviral construct called ENoMi, which consists of a re-engineered tetraspanin scaffold tagged with bioluminescent and fluorescent reporter proteins under an EF-1α promoter. Alongside this improved detection in ENoMi-hNPCs-derived EVs, their surface allows specific immunocapture purification, thereby facilitating TAF1-32i analysis. Using this ENoMi-labeling method, TAF1-32i was demonstrated in EVs released from XDP hNPCs implanted in mouse brains. Post-implantation of ENoMi-XDP hNPCs, TAF1-32i transcript was retrieved in EVs isolated from mouse brain and blood, and levels increased over time in plasma. We compared and combined our EV isolation technique to analyze XDP-derived TAF1-32i with other techniques, including size exclusion chromatography and Exodisc. Overall, our study demonstrates the successful engraftment of XDP patient-derived hNPCs in mice as a tool for monitoring disease markers with EVs., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

A nucleosome is typically positioned with its proximal edge (NPE) ∼50 bp downstream from the transcription start site of metazoan RNA polymerase II promoters. This +1 nucleosome has distinctive characteristics, including the presence of variant histone types and trimethylation of histone H3 at lysine 4. To address the role of these features in transcription complex assembly, we generated templates with four different promoters and nucleosomes located at a variety of downstream positions, which were transcribed in vitro using HeLa nuclear extracts. Two promoters lacked TATA elements, but all supported strong initiation from a single transcription start site. In contrast to results with minimal in vitro systems based on the TATA-binding protein (TBP), TATA promoter templates with a +51 NPE were transcriptionally inhibited in extracts; activity continuously increased as the nucleosome was moved downstream to +100. Inhibition was much more pronounced for the TATA-less promoters: +51 NPE templates were inactive, and substantial activity was only seen with the +100 NPE templates. Substituting the histone variants H2A.Z, H3.3, or both did not eliminate the inhibition. However, addition of excess TBP restored activity on nucleosomal templates with TATA promoters, even with an NPE at +20. Remarkably, nucleosomal templates with histone H3 trimethylated at lysine 4 are active with an NPE at +51 for both TATA and TATA-less promoters. Our results strongly suggest that the +1 nucleosome interferes with promoter recognition by TFIID. This inhibition can be overcome with TBP alone at TATA promoters or through positive interactions with histone modifications and TFIID., 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.)

Heart formation requires transcriptional regulators that underlie congenital anomalies and the fetal gene program activated during heart failure. Attributing the effects of congenital heart disease (CHD) missense variants to disruption of specific protein domains allows for a mechanistic understanding of CHDs and improved diagnostics. A combined chemical and genetic approach was employed to identify novel CHD drivers, consisting of chemical screening during pluripotent stem cell (PSC) differentiation, gene expression analyses of native tissues and primary cell culture models, and the in vitro study of damaging missense variants from CHD patients. An epigenetic inhibitor of the TATA-Box Binding Protein Associated Factor 1 (TAF1) bromodomain was uncovered in an unbiased chemical screen for activators of atrial and ventricular fetal myosins in differentiating PSCs, leading to the development of a high affinity inhibitor (5.1 nM) of the TAF1 bromodomain, a component of the TFIID complex. TAF1 bromodomain inhibitors were tested for their effects on stem cell viability and cardiomyocyte differentiation, implicating a role for TAF1 in cardiogenesis. Damaging TAF1 missense variants from CHD patients were studied by mutational analysis of the TAF1 bromodomain, demonstrating a repressive role of TAF1 that can be abrogated by the introduction of damaging bromodomain variants or chemical TAF1 bromodomain inhibition. These results indicate that targeting the TAF1/TFIID complex with chemical compounds modulates cardiac transcription and identify an epigenetically-driven CHD mechanism due to damaging variants within the TAF1 bromodomain., Competing Interests: Declaration of competing interest All authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Three classes of yeast protein-coding genes are distinguished by their dependence on the transcription cofactors TFIID, SAGA, and Mediator (MED) Tail, but whether this dependence is determined by the core promoter, upstream activating sequences (UASs), or other gene features is unclear. Also unclear is whether UASs can broadly activate transcription from the different promoter classes. Here, we measure transcription and cofactor specificity for thousands of UAS-core promoter combinations and find that most UASs broadly activate promoters regardless of regulatory class, while few display strong promoter specificity. However, matching UASs and promoters from the same gene class is generally important for optimal expression. We find that sensitivity to rapid depletion of MED Tail or SAGA is dependent on the identity of both UAS and core promoter, while dependence on TFIID localizes to only the promoter. Finally, our results suggest the role of TATA and TATA-like promoter sequences in MED Tail function., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Transcription by RNA Polymerase II (Pol II) is initiated by the hierarchical assembly of the pre-initiation complex onto promoter DNA. Decades of research have shown that the TATA-box binding protein (TBP) is essential for Pol II loading and initiation. Here, we report instead that acute depletion of TBP in mouse embryonic stem cells has no global effect on ongoing Pol II transcription. In contrast, acute TBP depletion severely impairs RNA Polymerase III initiation. Furthermore, Pol II transcriptional induction occurs normally upon TBP depletion. This TBP-independent transcription mechanism is not due to a functional redundancy with the TBP paralog TRF2, though TRF2 also binds to promoters of transcribed genes. Rather, we show that the TFIID complex can form and, despite having reduced TAF4 and TFIIA binding when TBP is depleted, the Pol II machinery is sufficiently robust in sustaining TBP-independent transcription., Competing Interests: JK, TN, AU, MB, JC, JL, FV, PL, ST No competing interests declared, (© 2023, Kwan, Nguyen et al.)

Taf4 (TATA-box binding protein-associated factor 4) is a subunit of the general transcription factor TFIID, a component of the RNA polymerase II pre-initiation complex that interacts with tissue-specific transcription factors to regulate gene expression. Properly regulated gene expression is particularly important in the intestinal epithelium that is constantly renewed from stem cells. Tissue-specific inactivation of Taf4 in murine intestinal epithelium during embryogenesis compromised gut morphogenesis and the emergence of adult-type stem cells. In adults, Taf4 loss impacted the stem cell compartment and associated Paneth cells in the stem cell niche, epithelial turnover and differentiation of mature cells, thus exacerbating the response to inflammatory challenge. Taf4 inactivation ex vivo in enteroids prevented budding formation and maintenance and caused broad chromatin remodeling and a strong reduction in the numbers of stem and progenitor cells with a concomitant increase in an undifferentiated cell population that displayed high activity of the Ezh2 and Suz12 components of Polycomb Repressive Complex 2 (PRC2). Treatment of Taf4-mutant enteroids with a specific Ezh2 inhibitor restored buddings, cell proliferation and the stem/progenitor compartment. Taf4 loss also led to increased PRC2 activity in cells of adult crypts associated with modification of the immune/inflammatory microenvironment that potentiated Apc-driven tumorigenesis. Our results reveal a novel function of Taf4 in antagonizing PRC2-mediated repression of the stem cell gene expression program to assure normal development, homeostasis, and immune-microenvironment of the intestinal epithelium., (© 2023. The Author(s).)

In Saccharomyces cerevisiae, class II gene promoters have been divided into two subclasses, TFIID- and SAGA-dominated promoters or TFIID-dependent and coactivator-redundant promoters, depending on the experimental methods used to measure mRNA levels. A prior study demonstrated that Spt3, a TBP-delivering subunit of SAGA, functionally regulates the PGK1 promoter via two mechanisms: by stimulating TATA box-dependent transcriptional activity and conferring Taf1/TFIID independence. However, only the former could be restored by plasmid-borne SPT3. In the present study, we sought to determine why ectopically expressed SPT3 is unable to restore Taf1/TFIID independence to the PGK1 promoter, identifying that this function was dependent on the construction protocol for the SPT3 taf1 strain. Specifically, simultaneous functional loss of Spt3 and Taf1 during strain construction was a prerequisite to render the PGK1 promoter Taf1/TFIID-dependent in this strain. Intriguingly, genetic approaches revealed that an as-yet unidentified trans-acting factor reprogrammed the transcriptional mode of the PGK1 promoter from the Taf1/TFIID-independent state to the Taf1/TFIID-dependent state. This factor was generated in the haploid SPT3 taf1 strain in an Hsp104-dependent manner and inherited meiotically in a non-Mendelian fashion. Furthermore, RNA-seq analyses demonstrated that this factor likely affects the transcription mode of not only the PGK1 promoter, but also of many other class II gene promoters. Collectively, these findings suggest that a prion or biomolecular condensate is generated in a Hsp104-dependent manner upon simultaneous functional loss of TFIID and SAGA, and could alter the roles of these transcription complexes on a wide variety of class II gene promoters without altering their primary sequences. Therefore, these findings could provide the first evidence that TFIID dependence of class II gene transcription can be altered epigenetically, at least in Saccharomyces cerevisiae., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Iwami et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Introduction: Oligoasthenoteratozoospermia (OAT) is a major cause of infertility in males. Only a few pathogenic genes of OAT have been clearly identified till now. A large number of OAT-affected cases remain largely unknown., Methods: Here, Whole-exome sequencing (WES) in 725 idiopathic OAT patients was performed. Ejaculated spermatozoa by OAT patients were microinjected into mouse oocytes to estimate fertilization potential. Diff-quick staining and transmission electron microscopy were performed to evaluate sperm morphology and ultrastructure. The protein expression level and localization In vitro were detected by Western Blotting and Immunocytochemistry., Results: We identified four X-linked hemizygous deleterious variants of TAF7L-namely, c.1301&#95;1302del;(p.V434Afs*5), c.699G>T;(p.R233S), c.508delA; (p. T170fs), c.719dupA;(p.K240fs) -in five probands. Intracytoplasmic sperm injection (ICSI) were carried out in M1, M2-1and M3 patient's wife. However only M1 patient's wife became pregnant after embryo transfer. In vitro study demonstrated significantly reduced fertilization ability in patient with TAF7L mutation. The TAF7L mutation let to abnormal sperm head and impaired histone-to protamine exchange. Variant 719dupA (p. K240fs) resulted in producing a truncated TAF7L protein and localized massively within the nucleus. In addition, TAF7L expression were not able to be detected due to variants c.1301&#95;1302del (p. V434Afs*5) and c.508delA (p. T170fs) In vitro., Conclusion: Our findings support that TAF7L is one of pathogenic genes of OAT and deleterious mutations in TAF7L may cause impaired histone-to-protamine affected the chromatin compaction of sperm head., 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 © 2023 Bai, Sha, Tan, Li, Zhang, Xu, Xu, Ji, Wang, Chen, Zhang, Yao, Li and Zhi.)

Objective: X-linked Dystonia Parkinsonism (XDP) is associated with a SINE-VNTR- Alu (SVA) retrotransposon insertion in an intron of the TAF1 gene that alters gene transcription and splicing. In this study, we determined if the SVA insertion introduces glucocorticoid (GC)-responsive cis -regulatory elements that may contribute to dysregulated TAF1 transcription and XDP disease progression., Methodology: We performed in silico analysis to identify potential GC receptor (GR) binding sites within the XDP-SVA. We also conducted promoter-reporter assays on HeLa and HEK293T cells to assess the intrinsic promoter activity of three XDP-SVA variants representing different hexameric repeat lengths associated with differences in disease onset. We treated XDP fibroblast cell models with GR agonist (CORT) or antagonist (RU486), then subjected TAF1 and the XDP-associated aberrant transcript, TAF1-32i to gene expression analysis., Results: A transcription factor binding site search revealed three binding sites for GR within the XDP-SVA-two within the SINE region and one in the Alu region. Promoter-reporter assays showed induction of XDP-SVA promoter activity upon CORT treatment that was dependent on the cell line and XDP-SVA hexamer repeat length. Gene expression analysis showed that baseline TAF1 levels differed between control and patient fibroblast cell lines, and treatment with CORT led to an increasing trend in the expression of the aberrant TAF1-32i transcript but did not reach statistical significance. Treatment with RU486 increased TAF1 mRNA expression only in the control cell lines., Conclusion: Using reporter assays, the XDP-SVA was shown to exhibit CORT-dependent transcriptional activation. Gene expression analysis also showed that GC signaling may influence TAF1 and TAF1-32i expression, possibly through interaction with the XDP-SVA. Our data provide a potential link between stress and XDP progression., Competing Interests: Both authors declared no conflict of interest., (© 2022 Journal of the ASEAN Federation of Endocrine Societies.)

Spermatogenesis is regulated by complex tissue specific gene expression in the testis to achieve normal male fertility. X-chromosome specific TATA binding protein (TBP)-associated factor 7L (hTAF7L) is one of the transcriptional regulator genes considered essential for spermatogenesis. The aim of this study was to evaluate the role of variants/mutations in the testis-specific hTAF7L gene in non-obstructive azoospermia and severe oligozoospermia male infertility. We studied 156 idiopathic non-obstructive azoospermic, severe oligozoospermic infertile males and 50 fertile proven controls. Infertile males and control subjects were genotyped for variants of the hTAF7L gene using polymerase chain reaction and a direct Sanger sequencing approach. The odds ratio evaluated the association of hTAF7L gene variants with idiopathic male infertility. The variants found in the hTAF7L gene were subjected to an in-silico analysis study. In infertile study subjects, we observed 11 single base pair nucleotide changes at various exons and three frameshift variants at exon 10 in the hTAF7L gene. We also found more than one variant in some non-obstructive azoospermia and severe oligozoospermia infertile males along with control subjects. All these variants changed the amino acid sequences in the hTAF7L gene. However, similar changes were also seen in fertile subjects, and the differences were not statistically significant. In-silico tools also predicted that the variants were likely to be benign. The variants in cDNA of the hTAF7L gene were typical SNPs. It is found that the hTAF7L gene is highly polymorphic and these missense variants are not directly associated with male infertility. However, we feel that more studies are needed to elucidate the role of multiple variants of the hTAF7L gene in the process of normal spermatogenesis., (© 2022 Wiley-VCH GmbH.)

Regulation of RNA polymerase II transcription requires the concerted efforts of several multisubunit coactivator complexes, which interact with the RNA polymerase II preinitiation complex to stimulate transcription. We previously showed that separation of the Mediator core from Mediator's tail module results in modest overactivation of genes annotated as highly dependent on TFIID for expression. However, it is unclear if other coactivators are involved in this phenomenon. Here, we show that the overactivation of certain genes by Mediator core/tail separation is blunted by disruption of the Spt-Ada-Gcn5-Acetyl transferase complex through the removal of its structural Spt20 subunit, though this downregulation does not appear to completely depend on reduced Spt-Ada-Gcn5-Acetyl transferase association with the genome. Consistent with the enrichment of TFIID-dependent genes among genes overactivated by Mediator core/tail separation, depletion of the essential TFIID subunit Taf13 suppressed the overactivation of these genes when Med16 was simultaneously removed. As with Spt-Ada-Gcn5-Acetyl transferase, this effect did not appear to be fully dependent on the reduced genomic association of TFIID. Given that the observed changes in gene expression could not be clearly linked to alterations in Spt-Ada-Gcn5-Acetyl transferase or TFIID occupancy, our data may suggest that the Mediator core/tail connection is important for the modulation of Spt-Ada-Gcn5-Acetyl transferase and/or TFIID conformation and/or function at target genes., (© The Author(s) 2022. Published by Oxford University Press on behalf of Genetics Society of America.)

TATA-binding protein associated factor 4 (TAF4) is a subunit of the Transcription Factor IID (TFIID) complex, a central player in transcription initiation. Other members of this multimeric complex have been implicated previously as monogenic disease genes in human developmental disorders. TAF4 has not been described to date as a monogenic disease gene. We here present a cohort of eight individuals, each carrying de novo putative loss-of-function (pLoF) variants in TAF4 and expressing phenotypes consistent with a neuro-developmental disorder (NDD). Common features include intellectual disability, abnormal behavior, and facial dysmorphisms. We propose TAF4 as a novel dominant disease gene for NDD, and coin this novel disorder "TAF4-related NDD" (T4NDD). We place T4NDD in the context of other disorders related to TFIID subunits, revealing shared features of T4NDD with other TAF-opathies., (© 2022 The Authors. Human Mutation published by Wiley Periodicals LLC.)

Mediator (MED) is a conserved factor with important roles in basal and activated transcription. Here, we investigate the genome-wide roles of yeast MED by rapid depletion of its activator-binding domain (Tail) and monitoring changes in nascent transcription. Rapid Tail depletion surprisingly reduces transcription from only a small subset of genes. At most of these Tail-dependent genes, in unperturbed conditions, MED is detected at both the UASs and promoters. In contrast, at most Tail-independent genes, we find MED primarily at promoters but not at the UASs. These results suggest that MED Tail and activator-mediated MED recruitment regulates only a small subset of genes. Furthermore, we define three classes of genes that differ in PIC assembly pathways and the requirements for MED Tail, SAGA, TFIID, and BET factors Bdf1/2. Our combined results have broad implications for the roles of MED, other coactivators, and mechanisms of transcriptional regulation at different gene classes., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Viroids, a fascinating group of plant pathogens, are subviral agents composed of single-stranded circular noncoding RNAs. It is well-known that nuclear-replicating viroids exploit host DNA-dependent RNA polymerase II (Pol II) activity for transcription from circular RNA genome to minus-strand intermediates, a classic example illustrating the intrinsic RNA-dependent RNA polymerase activity of Pol II. The mechanism for Pol II to accept single-stranded RNAs as templates remains poorly understood. Here, we reconstituted a robust in vitro transcription system and demonstrated that Pol II also accepts minus-strand viroid RNA template to generate plus-strand RNAs. Further, we purified the Pol II complex on RNA templates for nano-liquid chromatography-tandem mass spectrometry analysis and identified a remodeled Pol II missing Rpb4, Rpb5, Rpb6, Rpb7, and Rpb9, contrasting to the canonical 12-subunit Pol II or the 10-subunit Pol II core on DNA templates. Interestingly, the absence of Rpb9, which is responsible for Pol II fidelity, explains the higher mutation rate of viroids in comparison to cellular transcripts. This remodeled Pol II is active for transcription with the aid of TFIIIA-7ZF and appears not to require other canonical general transcription factors (such as TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, and TFIIS), suggesting a distinct mechanism/machinery for viroid RNA-templated transcription. Transcription elongation factors, such as FACT complex, PAF1 complex, and SPT6, were also absent in the reconstituted transcription complex. Further analyses of the critical zinc finger domains in TFIIIA-7ZF revealed the first three zinc finger domains pivotal for RNA template binding. Collectively, our data illustrated a distinct organization of Pol II complex on viroid RNA templates, providing new insights into viroid replication, the evolution of transcription machinery, as well as the mechanism of RNA-templated transcription., Competing Interests: The authors have declared that no competing interests exist.

TAF8 is part of the transcription factor II D complex, composed of the TATA-binding protein and 13 TATA-binding protein-associated factors (TAFs). Transcription factor II D is the first general transcription factor recruited at promoters to assemble the RNA polymerase II preinitiation complex. So far disorders related to variants in 5 of the 13 subunits of human transcription factor II D have been described. Recently, a child with a homozygous c.781-1G>A mutation in TAF8 has been reported. Here we describe seven further patients with mutations in TAF8 and thereby confirm the TAF8 related disorder. In two sibling patients, we identified two novel compound heterozygous TAF8 splice site mutations, c.45+4A > G and c.489G>A, which cause aberrant splicing as well as reduced expression and mislocalization of TAF8. In five further patients, the previously described c.781-1G > A mutation was present on both alleles. The clinical phenotype associated with the different TAF8 mutations is characterized by severe psychomotor retardation with almost absent development, feeding problems, microcephaly, growth retardation, spasticity and epilepsy. Cerebral imaging showed hypomyelination, a thin corpus callosum and brain atrophy. Moreover, repeated imaging in the sibling pair demonstrated progressive cerebral and cerebellar atrophy. Consistently, reduced N-acetylaspartate, a marker of neuronal viability, was observed on magnetic resonance spectroscopy. Further review of the literature shows that mutations causing a reduced expression of transcription factor II D subunits have an overlapping phenotype of microcephaly, developmental delay and intellectual disability. Although transcription factor II D plays an important role in RNA polymerase II transcription in all cells and tissues, the symptoms associated with such defects are almost exclusively neurological. This might indicate a specific vulnerability of neuronal tissue to widespread deregulation of gene expression as also seen in Rett syndrome or Cornelia de Lange syndrome., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The effects of rapid acute depletion of components of RNA polymerase II (Pol II) general transcription factors (GTFs) that are thought to be critical for formation of preinitiation complexes (PICs) and initiation in vitro were quantified in HAP1 cells using precision nuclear run-on sequencing (PRO-Seq). The average dependencies for each factor across >70 000 promoters varied widely even though levels of depletions were similar. Some of the effects could be attributed to the presence or absence of core promoter elements such as the upstream TBP-specificity motif or downstream G-rich sequences, but some dependencies anti-correlated with such sequences. While depletion of TBP had a large effect on most Pol III promoters only a small fraction of Pol II promoters were similarly affected. TFIIB depletion had the largest general effect on Pol II and also correlated with apparent termination defects downstream of genes. Our results demonstrate that promoter activity is combinatorially influenced by recruitment of TFIID and sequence-specific transcription factors. They also suggest that interaction of the preinitiation complex (PIC) with nucleosomes can affect activity and that recruitment of TFIID containing TBP only plays a positive role at a subset of promoters., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Genome-wide, little is understood about how proteins organize at inducible promoters before and after induction and to what extent inducible and constitutive architectures depend on cofactors. We report that sequence-specific transcription factors and their tethered cofactors (e.g., SAGA [Spt-Ada-Gcn5-acetyltransferase], Mediator, TUP, NuA4, SWI/SNF, and RPD3-L) are generally bound to promoters prior to induction ("poised"), rather than recruited upon induction, whereas induction recruits the preinitiation complex (PIC) to DNA. Through depletion and/or deletion experiments, we show that SAGA does not function at constitutive promoters, although a SAGA-independent Gcn5 acetylates +1 nucleosomes there. When inducible promoters are poised, SAGA catalyzes +1 nucleosome acetylation but not PIC assembly. When induced, SAGA catalyzes acetylation, deubiquitylation, and PIC assembly. Surprisingly, SAGA mediates induction by creating a PIC that allows TFIID (transcription factor II-D) to stably associate, rather than creating a completely TFIID-independent PIC, as generally thought. These findings suggest that inducible systems, where present, are integrated with constitutive systems., (© 2022 Mittal et al.; Published by Cold Spring Harbor Laboratory Press.)

Enhancers in higher eukaryotes and upstream activating sequences (UASs) in yeast have been shown to recruit components of the RNA polymerase II (Pol II) transcription machinery. At least a fraction of Pol II recruited to enhancers in higher eukaryotes initiates transcription and generates enhancer RNA (eRNA). In contrast, UASs in yeast do not recruit transcription factor TFIIH, which is required for transcription initiation. For both yeast and mammalian systems, it was shown that Pol II is transferred from enhancers/UASs to promoters. We propose that there are two modes of Pol II recruitment to enhancers in higher eukaryotes. Pol II complexes that generate eRNAs are recruited via TFIID, similar to mechanisms operating at promoters. This may involve the binding of TFIID to acetylated nucleosomes flanking the enhancer. The resulting eRNA, together with enhancer-bound transcription factors and co-regulators, contributes to the second mode of Pol II recruitment through the formation of a transcription initiation domain. Transient contacts with target genes, governed by proteins and RNA, lead to the transfer of Pol II from enhancers to TFIID-bound promoters.