TATA-binding associated factors have distinct roles during early mammalian development.

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.

[Clinical features and genetic analysis of 17 Chinese pedigrees affected with X-linked intellectual disability].

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_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.

Insights into Molecular Diversity within the FUS/EWS/TAF15 Protein Family: Unraveling Phase Separation of the N-Terminal Low-Complexity Domain from RNA-Binding Protein EWS.

The FET protein family, comprising FUS, EWS, and TAF15, plays crucial roles in mRNA maturation, transcriptional regulation, and DNA damage response. Clinically, they are linked to Ewing family tumors and neurodegenerative diseases such as amyotrophic lateral sclerosis. The fusion protein EWS::FLI1, the causative mutation of Ewing sarcoma, arises from a genomic translocation that fuses a portion of the low-complexity domain (LCD) of EWS (EWSLCD) with the DNA binding domain of the ETS transcription factor FLI1. This fusion protein modifies transcriptional programs and disrupts native EWS functions, such as splicing. The exact role of the intrinsically disordered EWSLCD remains a topic of active investigation, but its ability to phase separate and form biomolecular condensates is believed to be central to EWS::FLI1's oncogenic properties. Here, we used paramagnetic relaxation enhancement NMR, microscopy, and all-atom molecular dynamics (MD) simulations to better understand the self-association and phase separation tendencies of the EWSLCD. Our NMR data and mutational analysis suggest that a higher density and proximity of tyrosine residues amplify the likelihood of condensate formation. MD simulations revealed that the tyrosine-rich termini exhibit compact conformations with unique contact networks and provided critical input on the relationship between contacts formed within a single molecule (intramolecular) and inside the condensed phase (intermolecular). These findings enhance our understanding of FET proteins' condensate-forming capabilities and underline differences between EWS, FUS, and TAF15.

TAF7L regulates early stages of male germ cell development in the rat.

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.

Participation of the TBP-associated factors (TAFs) in cell differentiation.

The understanding of the mechanisms that regulate gene expression to establish differentiation programs and determine cell lineages, is one of the major challenges in Developmental Biology. Besides the participation of tissue-specific transcription factors and epigenetic processes, the role of general transcription factors has been ignored. Only in recent years, there have been scarce studies that address this issue. Here, we review the studies on the biological activity of some TATA-box binding protein (TBP)-associated factors (TAFs) during the proliferation of stem/progenitor cells and their involvement in cell differentiation. Particularly, the accumulated evidence suggests that TAF4, TAF4b, TAF7L, TAF8, TAF9, and TAF10, among others, participate in nervous system development, adipogenesis, myogenesis, and epidermal differentiation; while TAF1, TAF7, TAF15 may be involved in the regulation of stem cell proliferative abilities and cell cycle progression. On the other hand, evidence suggests that TBP variants such as TBPL1 and TBPL2 might be regulating some developmental processes such as germ cell maturation and differentiation, myogenesis, or ventral specification during development. Our analysis shows that it is necessary to study in greater depth the biological function of these factors and its participation in the assembly of specific transcription complexes that contribute to the differential gene expression that gives rise to the great diversity of cell types existing in an organism. The understanding of TAFs' regulation might lead to the development of new therapies for patients which suffer from mutations, alterations, and dysregulation of these essential elements of the transcriptional machinery.

Deletion of taf1 and taf5 in zebrafish capitulate cardiac and craniofacial abnormalities associated with TAFopathies through perturbations in metabolism.

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.

m6A-enriched lncRNA LINC00839 promotes tumor progression by enhancing TAF15-mediated transcription of amine oxidase AOC1 in nasopharyngeal carcinoma.

Dysregulation of long noncoding RNAs (lncRNAs) contributes to tumorigenesis by modulating specific cancer-related pathways, but the roles of N6-methyladenosine (m6A)-enriched lncRNAs and underlying mechanisms remain elusive in nasopharyngeal carcinoma (NPC). Here, we reanalyzed the previous genome-wide analysis of lncRNA profiles in 18 pairs of NPC and normal tissues as well as in ten paired samples from NPC with or without post-treatment metastases. We discerned that an oncogenic m6A-enriched lncRNA, LINC00839, which was substantially upregulated in NPC and correlated with poor clinical prognosis, promoted NPC growth and metastasis both in vitro and in vivo. Mechanistically, by using RNA pull-down assay combined with mass spectrometry, we found that LINC00839 interacted directly with the transcription factor, TATA-box binding protein associated factor (TAF15). Besides, chromatin immunoprecipitation and dual-luciferase report assays demonstrated that LINC00839 coordinated the recruitment of TAF15 to the promoter region of amine oxidase copper-containing 1 (AOC1), which encodes a secreted glycoprotein playing vital roles in various cancers, thereby activating AOC1 transcription in trans. In this study, potential effects of AOC1 in NPC progression were first proposed. Moreover, ectopic expression of AOC1 partially rescued the inhibitory effect of downregulation of LINC00839 in NPC. Furthermore, we showed that silencing vir-like m6A methyltransferase-associated (VIRMA) and insulin-like growth factor 2 mRNA-binding proteins 1 (IGF2BP1) attenuated the expression level and RNA stability of LINC00839 in an m6A-dependent manner. Taken together, our study unveils a novel oncogenic VIRMA/IGF2BP1-LINC00839-TAF15-AOC1 axis and highlights the significance and prognostic value of LINC00839 expression in NPC carcinogenesis.

BcTaf14 regulates growth and development, virulence, and stress responses in the phytopathogenic fungus Botrytis cinerea.

TATA box-binding protein (TBP)-associated factor 14 (Taf14), a transcription-associated factor containing a conserved YEATS domain and an extra-terminal (ET) domain, is a multifunctional protein in Saccharomyces cerevisiae. However, the role of Taf14 in filamentous phytopathogenic fungi is not well understood. In this study, the homologue of ScTaf14 in Botrytis cinerea (named BcTaf14), a destructive phytopathogen causing grey mould, was investigated. The BcTaf14 deletion strain (ΔBcTaf14) showed pleiotropic defects, including slow growth, abnormal colony morphology, reduced conidiation, abnormal conidial morphology, reduced virulence, and altered responses to various stresses. The ΔBcTaf14 strain also exhibited differential expression of numerous genes compared to the wild-type strain. BcTaf14 could interact with the crotonylated H3K9 peptide, and mutation of two key sites (G80 and W81) in the YEATS domain disrupted this interaction. The mutation of G80 and W81 affected the regulatory effect of BcTaf14 on mycelial growth and virulence but did not affect the production and morphology of conidia. The absence of the ET domain at the C-terminus rendered BcTaf14 unable to localize to the nucleus, and the defects of ΔBcTaf14 were not recovered to wild-type levels when BcTaf14 without the ET domain was expressed. Our results provide insight into the regulatory roles of BcTaf14 and its two conserved domains in B. cinerea and will be helpful for understanding the function of the Taf14 protein in plant-pathogenic fungi.

An increase in polyadenylation of histone isoforms, Hist1h2ah and Hist2h3c2, is governed by 3'-UTR in de-differentiated and undifferentiated hepatocyte.

Replication-dependent histones have a stem-loop structure at the 3' end of messenger RNA (mRNA) and are stabilized by stem-loop binding protein (SLBP). Moreover, loss of SLBP and imbalance in the level of ARE (adenylate-uridylate-rich elements)-binding proteins, HuR, and BRF1 are associated with the polyadenylation of canonical histone mRNAs under different physiological conditions. Previous studies from the lab have shown increased protein levels of H2A1H and H3.2 in N-nitrosodiethylamine (NDEA)-induced hepatocellular carcinoma (HCC). In this study, we report that increase in the polyadenylation of histone mRNA contributes to increased levels of H2A1H and H3.2 in NDEA-induced HCC. The persistent exposure to carcinogen with polyadenylation of histone mRNA increases the total histone pool resulting in aneuploidy. The embryonic liver has also shown increased polyadenylated histone isoforms, Hist1h2ah and Hist2h3c2, primarily contributing to their increased protein levels. The increase in polyadenylation of histone mRNA in HCC and e15 are in coherence with the decrease in SLBP and BRF1 with an increase in HuR. Our studies in neoplastic CL38 cell line showed that direct stress on the cells induces downregulation of SLBP with enhanced histone isoform polyadenylation. Moreover, the polyadenylation is related to increase in activated MAP kinases, p38, ERK, and JNK in HCC liver tumor tissues and CL38 cells treated with arsenic. Our data suggest that SLBP degrades under stress, destabilizing the stem-loop, elongating histone isoforms mRNA with 3' polyadenylated tail with increase of HuR and decrease of BRF1. Overall, our results indicate that SLBP may play an essential part in cell proliferation, at least in persistent exposure to stress, by mediating the stabilization of histone isoforms throughout the cell cycle.

CircDNAJC11 interacts with TAF15 to promote breast cancer progression via enhancing MAPK6 expression and activating the MAPK signaling pathway.

BACKGROUND: Breast cancer (BC) is a common malignant tumor in women worldwide. Circular RNA (circRNA) has been proven to play a critical role in BC progression. However, the exact biological functions and underlying mechanisms of circRNAs in BC remain largely unknown. METHODS: Here, we first screened for differentially expressed circRNAs in 4 pairs of BC tissues and adjacent non-tumor tissues using a circRNA microarray. Functionally, gain- and loss-of-function experiments in vitro and in vivo showed that circDNAJC11 promoted BC cell proliferation, migration, invasion, and tumor growth. Mechanistically, RNA pull-down, mass spectrum, RNA immunoprecipitation, fluorescence in situ hybridization assays, and rescue experiments were executed. RESULTS: We found that circDNAJC11 was significantly upregulated in triple-negative breast cancer tissues and cells. Clinical data revealed that the high expression of circDNAJC11 was closely correlated with a poor prognosis of BC patients and could be an independent risk factor for BC prognosis. Functionally, gain- and loss-of-function experiments in vitro and in vivo showed that circDNAJC11 promoted BC cell proliferation, migration, invasion, and tumor growth. Mechanistically, RNA pull-down, mass spectrum, RNA immunoprecipitation, fluorescence in situ hybridization assays, and rescue experiments were executed. We demonstrated that circDNAJC11 combined with TAF15 to promote BC progression via stabilizing MAPK6 mRNA and activating the MAPK signaling pathway. CONCLUSIONS: The circDNAJC11/TAF15/MAPK6 axis played a crucial role in the progression and development of BC, suggesting that circDNAJC11 might be a novel biomarker and therapeutical target for BC.

Extraskeletal Myxoid Chondrosarcomas: The Uncommon Clinicopathologic Manifestations and Significance of TAF15::NR4A3 Fusion.

Extraskeletal myxoid chondrosarcoma (EMC) is an ultrarare sarcoma typically exhibiting myxoid/reticular histology and NR4A3 translocation. However, morphologic variants and the relevance of non-EWSR1::NR4A3 fusions remain underexplored. Three challenging pan-Trk-expressing cases, featuring cellular to solid histology, were subjected to RNA exome sequencing (RES), unveiling different NR4A3-associated fusions. Alongside RES-analyzed cases, fluorescence in situ hybridization was performed to confirm 58 EMCs, with 48 available for pan-Trk immunostaining and KIT sequencing. Except for 1 (2%) NR4A3-rearranged EMC without identifiable partners, 46 (79%), 9 (16%), and 2 (3%) cases harbored EWSR1::NR4A3, TAF15::NR4A3, and TCF12::NR4A3 fusions, respectively. Five EWSR1::NR4A3-positive EMCs occurred in the subcutis (3) and bone (2). Besides 43 classical cases, there were 8 cellular, 4 rhabdoid/anaplastic, 2 solid, and 1 mixed tumor-like variants. Tumor cells were oval/spindle to pleomorphic and formed loose myxoid/reticular to compact sheet-like or fascicular patterns, imparting broad diagnostic considerations. RES showed upregulation of NTRK2/3, KIT, and INSM1. Moderate-to-strong immunoreactivities of pan-Trk, CD117, and INSM1 were present in 35.4%, 52.6%, and 54.6% of EMCs, respectively. KIT p. E554K mutation was detected in 2/48 cases. TAF15::NR4A3 was significantly associated with size >10 cm (78%, P = .025). Size >10 cm, moderate-to-severe nuclear pleomorphism, metastasis at presentation, TAF15::NR4A3 fusion, and the administration of chemotherapy portended shorter univariate disease-specific survival, whereas only size >10 cm (P = .004) and metastasis at presentation (P = .032) remained prognostically independent. Conclusively, EMC may manifest superficial or osseous lesions harboring EWSR1::NR4A3, underrecognized solid or anaplastic histology, and pan-Trk expression, posing tremendous challenges. Most TAF15::NR4A3-positive cases were >10 cm in size, ie, a crucial independent prognosticator, whereas pathogenic KIT mutation rarely occurred.

Utility of whole-exome sequencing for patients with multiple congenital anomalies with or without intellectual disability/developmental delay in East Asia population.

BACKGROUND: Congenital anomalies (CAs) with or without intellectual disability (ID)/developmental delay (DD) comprise a heterogeneous spectrum of diseases that affect approximately 3% of live births worldwide. Recently, whole-exome sequencing (WES) demonstrated the highly heterogeneous genetic causes of CAs. The purpose of this study was to evaluate a referral system to increase the yield of WES for CAs. METHODS: From August 2018 to July 2019, patients with CAs, with or without ID/DD, after excluding gross chromosomal aberrations, were referred to geneticists in two medical centers. Variant prioritization was conducted with an AI-assisted tool for whole exomes or a CA-related gene panel. RESULTS: Forty patients (27 males and 13 females) with CAs were enrolled in the study with a mean age of 4.71 years (range, 0.01-18.2). Pathogenic variants in 14 genes were discovered in 16 patients (three patients with CHD7 and 13 patients with one gene each of ATP6V1B2, TAF6, COL4A3BP, ANKH, BMP2, SMARCA4, CUL4B, PGAP3, SOX11, FBN2, PTPN11, SOS1, or PROKR2), with a positive diagnostic rate of 40%. Among the 16 positive cases, 13 (81%) also had ID/DD. The inheritance was autosomal dominant in 13 (81%), autosomal recessive in two (13%), and X-linked in one (6%). Only five patients received a correct clinical diagnosis before WES. The analyses of patients with a negative genetic diagnosis revealed a phenotype and gene mutation load similar to those of the positive-finding patients but with a lower percentage of ID/DD. CONCLUSIONS: The careful selection of patients by experienced geneticists and the exclusion of chromosomal aberrations raises the positive rate of the molecular diagnosis for CAs to 40%. However, more than half of the patients with CAs still do not have a genetic diagnosis by current technologies.

TFIID dependency of steady-state mRNA transcription altered epigenetically by simultaneous functional loss of Taf1 and Spt3 is Hsp104-dependent.

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.

Small molecule Z363 co-regulates TAF10 and MYC via the E3 ligase TRIP12 to suppress tumour growth.

BACKGROUND: The MYC oncoprotein, also known as the master regulator of genes, is a transcription factor that regulates numerous physiological processes, including cell cycle control, apoptosis, protein synthesis and cell adhesion, among others. MYC is overexpressed in approximately 70% of human cancers. Given its pervasive role in cancer biology, MYC down-regulation has become an attractive cancer treatment strategy. METHODS: The CRISPR/Cas9 method was used to produce KO cell models. Western blot was used to analyzed the expressions of MYC and TATA-binding proteinassociated factors 10 (TAF10) in cancer cells (MCF7, A549, HepG2 cells) Cell culture studies were performed to determine the mechanisms by which small molecules (Z363119456, Z363) affects MYC and TAF10 expressions and functions. Mouse studies were carried out to investigate the impact of Z363 regulation on tumor growth. RESULTS: Z363 activate Thyroid hormone Receptor-interacting Protein 12 (TRIP12), which phosphorylates MYC at Thr58, resulting in MYC ubiquitination and degradation and thereby regulating MYC target genes. Importantly, TRIP12 also induces TAF10 degradation, which reduces MYC protein levels. TRIP12, an E3 ligase, controls MYC levels both directly and indirectly by inhibiting MYC or TAF10 activity. CONCLUSIONS: In summary,these results demonstrate the anti-cancer properties of Z363, a small molecule that is co-regulated by TAF10 and MYC.

BDP1 as a biomarker in serous ovarian cancer.

BACKGROUND: TFIIIB, an RNA polymerase III specific transcription factor has been found to be deregulated in human cancers with much of the research focused on the TBP, BRF1, and BRF2 subunits. To date, the TFIIIB specific subunit BDP1 has not been investigated in ovarian cancer but has previously been shown to be deregulated in neuroblastoma, breast cancer, and Non-Hodgkins lymphoma. RESULTS: Using in silico analysis of clinically derived platforms, we report a decreased BDP1 expression as a result of deletion in serous ovarian cancer and a correlation with higher and advanced ovarian stages. Further analysis in the context of TP53 mutations, a major contributor to ovarian tumorigenesis, suggests that high BDP1 expression is unfavorable for overall survival and high BDP1 expression occurs in stages 2, 3 and 4 serous ovarian cancer. Additionally, high BDP1 expression is disadvantageous and unfavorable for progression-free survival. Lastly, BDP1 expression significantly decreased in patients treated with first-line chemotherapy, platin and taxane, at twelve-month relapse-free survival. CONCLUSIONS: Taken together with a ROC analysis, the data suggest BDP1 could be of clinical relevance as a predictive biomarker in serous ovarian cancer. Lastly, this study further demonstrates that both the over- and under expression of BDP1 warrants further investigation and suggests BDP1 may exhibit dual function in the context of tumorigenesis.

Long non-coding RNA SNHG4 enhances RNF14 mRNA stability to promote the progression of colorectal cancer by recruiting TAF15 protein.

SNHG4 is a lncRNA that was previously reported to promote colorectal cancer (CRC) progression via molecular sponge mechanism. Bioinformatic analysis suggested SNHG4 might scaffold TAF15 protein-RNF14 mRNA interaction. We aimed to investigate the mechanisms of potential SNHG4/TAF15/RNF14 axis in promoting CRC malignant phenotypes. Protein-RNA interaction was determined using RNA immunoprecipitation, pull-down and fluorescence in situ hybridization (FISH) combined immunofluorescence assays. Cell apoptosis rates were quantified using flow cytometry. CCK-8 and colony formation were adopted to determine cell proliferation. Wound healing and transwell assays were employed to assess cell migration and invasion, respectively. Xenograft tumor model was applied to assess the effects of SNHG4 on CRC tumorigenesis in vivo. SNHG4, TAF15 and RNF14 were up-regulated in CRC tissues. SNHG4 overexpression promoted cell proliferation, migration, invasion, and Wnt/ß-catenin pathway activation in vitro, as well as tumor growth in vivo. The inhibited malignant phenotypes caused by SNHG4 knockdown were impeded by TAF15 or RNF14 overexpression. Mechanistically, SNHG4 recruited TAF15 protein and thus promoted the interaction between TAF15 protein and RNF14 mRNA, leading to the increased RNF14 mRNA stability. This in turn facilitated the Wnt/ß-catenin signal transduction. SNHG4 enhanced RNF14 mRNA stability and activated the Wnt/ß-catenin pathway to promote the progression of colorectal cancer by recruiting TAF15 protein.

TFIIB-related factor 1 is a nucleolar protein that promotes RNA polymerase I-directed transcription and tumour cell growth.

Eukaryotic RNA polymerase I (Pol I) products play fundamental roles in ribosomal assembly, protein synthesis, metabolism and cell growth. Abnormal expression of both Pol I transcription-related factors and Pol I products causes a range of diseases, including ribosomopathies and cancers. However, the factors and mechanisms governing Pol I-dependent transcription remain to be elucidated. Here, we report that transcription factor IIB-related factor 1 (BRF1), a subunit of transcription factor IIIB required for RNA polymerase III (Pol III)-mediated transcription, is a nucleolar protein and modulates Pol I-mediated transcription. We showed that BRF1 can be localized to the nucleolus in several human cell types. BRF1 expression correlates positively with Pol I product levels and tumour cell growth in vitro and in vivo. Pol III transcription inhibition assays confirmed that BRF1 modulates Pol I-directed transcription in an independent manner rather than through a Pol III product-to-45S pre-rRNA feedback mode. Mechanistically, BRF1 binds to the Pol I transcription machinery components and can be recruited to the rDNA promoter along with them. Additionally, alteration of BRF1 expression affects the recruitment of Pol I transcription machinery components to the rDNA promoter and the expression of TBP and TAF1A. These findings indicate that BRF1 modulates Pol I-directed transcription by controlling the expression of selective factor 1 subunits. In summary, we identified a novel role of BRF1 in Pol I-directed transcription, suggesting that BRF1 can independently regulate both Pol I- and Pol III-mediated transcription and act as a key coordinator of Pol I and Pol III.

Reduced oxidative stress suppresses neurotoxicity in the Drosophila model of TAF15-associated proteinopathies.

TATA-binding protein associated factor 15 (TAF15) is involved in the pathology of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Although TAF15 accumulates as cytoplasmic aggregates in neurons and the clearance of aggregates may be a therapeutic strategy for ALS, the underlying mechanisms of TAF15 in ALS remain poorly understood. Recently, we found that glutathione transferase omega 2 (GstO2) expression level is significantly reduced in the brain tissue of TAF15-expressing flies. In this study, we demonstrated that GstO2 overexpression in TAF15-induced flies rescues the locomotive activity and neuromuscular junctional defects. Furthermore, TAF15 levels in both cytoplasm and nuclear fractions significantly decreased in the heads of GstO2 co-expressing flies. GstO2-co-overexpression in neurons caused a marked decrease in intracellular reactive oxygen species generation in TAF15-induced flies. Our findings demonstrated that GstO2 was a pathogenic regulator of TAF15-associated proteinopathies. They help expand our understanding of TAF15-associated ALS pathogenesis.

TIF1γ inhibits lung adenocarcinoma EMT and metastasis by interacting with the TAF15/TBP complex.

The molecular underpinnings of lung adenocarcinoma (LUAD) metastasis remain poorly defined. Here, using human LUAD cell lines, we find that transcriptional intermediary factor 1 γ (TIF1γ) binds to TATA box binding protein (TBP) in competition with TBP-associated factor 15 (TAF15) and impedes TAF15/TBP-mediated interleukin 6 (IL-6) transactivation. TIF1γ modifies TAF15 through multi-mono-ubiquitylation and drives nuclear export of TAF15. Functionally, TAF15 accelerates epithelial-mesenchymal transition (EMT) and metastasis of LUAD cells, acting in just the opposite way as TIF1γ. Low TIF1γ or high TAF15 expression levels are shown in metastatic LUAD specimens and correlate with poor survival of individuals with LUAD. Our findings suggest that the TAF15/TBP complex is required for IL-6 activation-induced EMT and invasion, which are inhibited by TIF1γ. This study highlights the crucial interaction between TIF1γ and the TAF15/TBP complex for regulating EMT and metastasis in LUAD.

The association of testis-specific hTAF7L gene variants with idiopathic azoospermic and severe oligozoospermic male infertility.

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.