Similar Datasets
Project description:RNA polymerase II (Pol II) transcription initiation requires assembly of a preinitiation complex (PIC) comprising the general transcription factors amongst which the multiprotein complex TFIID. TFIID comprises the TATA-binding protein (TBP) and 13 TBP-associated factors (TAFs). Recent cryo-electron microscopy studies reported topological reorganization of TFIID upon promoter binding with the TAF11-TAF13 heterodimer as the last TFIID subunits directly contacting TBP prior to deposition on promoter DNA upstream of the transcription start site. To challenge the role of the TAF11-TAF13 heterodimer, we inactivated the gene encoding Taf13 both in mice and embryonic stem cells (ESCs). Taf13 inactivation in mice is embryonic lethal with null embryos implanting but surviving only until E6.5. Taf13-null ESCs are viable, but show slower proliferation and fail to form embryoid bodies and differentiate. Biochemical analyses from these ESCs show that Taf13 loss had minimal effect on TFIID integrity, and genomic profiling shows only a mild reduction of promoter recruitment of TBP and Taf4 while that of Taf1 is increased. Moreover, while TFIIH recruitment is unaffected indicating normal PIC formation, that of Pol II is globally diminished. These data indicate that the Taf11-Taf13 heterodimer is not essential for TBP/TFIID recruitment and PIC formation, but is specifically required for normal Pol II recruitment.
2025-06-18 | GSE282035 | GEO
Project description:RNA polymerase II transcription initiation requires assembly of a preinitiation complex (PIC). According to the stepwise model of PIC assembly, TFIID is the first GTF to be recruited. TFIID comprises TBP and 13 TBP-associated factors (TAFs). We focused on TAF13, the smallest subunit of TFIID. Taf13 has a histone-like domain that heterodimerizes with its partner Taf11. Recently published structures of TFIID place the Taf13-Taf11 heterodimer in lobe A where it forms a “bridge” between TBP and TFIID. According to this structure, the Taf11-Taf13 heterodimer has been suggested to be critical for TBP deposition at promoters. To better address this question, we generated mouse Embryonic Stem Cells (ESCs) cell lines and genetically modified mice where the gene coding for Taf13 was inactivated. Taf13-null mice died at E6.5-E7.5, whereas ESCs survived up to 15 days after inactivation of Taf13, but showed slower growth as compared to controls. In order to understand how inactivation of Taf13 impact the genes expressions we performed bulk mRNA-seq from the Taf13 expressing and null lines. This analysis showed that loss of TAF13 resulted in a greater than two-fold upregulation of 562 genes but only 41 of them were up-regulated more than four-fold. On the other hand, 382 genes were down-regulated, 34 of which showed more than a four-fold repression. Ontology analyses showed that the affected genes did not belong to any specific pathway, but we noticed that many primitive endoderm markers were up-regulated by TAF13 deficiency (Sox7, Sox17, Gata4, Gata6) and some ESC markers were downregulated (Nanog, Klf4, Zfp42) while the pluripotency markers Sox2 and Oct3/4 were unchanged.
2025-06-18 | GSE254024 | GEO
Project description:RNA polymerase II transcription initiation requires assembly of a preinitiation complex (PIC). According to the stepwise model of PIC assembly, TFIID is the first GTF to be recruited. TFIID comprises TBP and 13 TBP-associated factors (TAFs). We focused on TAF13, the smallest subunit of TFIID. Taf13 has a histone-like domain that heterodimerizes with its partner Taf11. Recently published structures of TFIID place the Taf13-Taf11 heterodimer in lobe A where it forms a “bridge” between TBP and TFIID. According to this structure, the Taf11-Taf13 heterodimer has been suggested to be critical for TBP deposition at promoters. To better address this question, we generated mouse Embryonic Stem Cells (ESCs) cell lines and genetically modified mice where the gene coding for Taf13 was inactivated. Taf13-null mice died at E6.5-E7.5, whereas ESCs survived up to 15 days after inactivation of Taf13, but showed slower growth as compared to controls. To understand how Taf13 inactivation impacts TFIID and RNA polymerase II (Pol II) recruitment, we performed ChIP seq for the TBP and Taf1 subunits of TFIID along with PolII on ESC lines expressing or not Taf13. ChIP seq for histone marks H3K4Me3 and H3K27ac were also performed. Taf13 inactivation does not impair TBP recruitment at promoters, while that of Pol II was reduced both at the promoter and gene body. Surprisingly, recruitment of Taf1 was increased and while the H3K4Me3 mark was not affected, H3K27ac was notably increased in the absence of Taf13. Taken together, these data showed that loss of the Taf11-Taf13 heterodimer did not strongly impact TBP recruitment to promoters, but led to increased Taf1 and H3K27ac levels, but reduced paused and elongating Pol II.
2025-06-18 | GSE254023 | GEO
Project description:The TFIID subunit Taf13 is dispensable for RNA polymerase II transcription, but essential for early embryogenesis
| PRJNA1068141 | ENA
Project description:The TFIID subunit Taf13 is dispensable for RNA polymerase II transcription, but essential for early embryogenesis [CUT&TAG]
| PRJNA1186697 | ENA
Project description:The TFIID subunit Taf13 is dispensable for RNA polymerase II transcription, but essential for early embryogenesis (RNA-Seq)
| PRJNA1068142 | ENA
Project description:The TFIID subunit Taf13 is dispensable for RNA polymerase II transcription, but essential for early embryogenesis (ChIP-Seq)
| PRJNA1068143 | ENA
Project description:The evolutionarily conserved RNA polymerase II transcription factor D (TFIID) complex is composed of TATA box-binding protein (TBP) and 13 TBP-associated factors (Tafs). The mechanisms by which many Taf subunits contribute to the essential function of TFIID are only poorly understood. To address this gap in knowledge, we present the results of a molecular genetic dissection of the TFIID subunit Taf2. Through systematic site-directed mutagenesis, we have discovered 12 taf2 temperature-sensitive (ts) alleles. Two of these alleles display growth defects that can be strongly suppressed by overexpression of the yeast-specific TFIID subunit TAF14 but not by overexpression of any other TFIID subunit. In Saccharomyces cerevisiae, Taf14 is also a constituent of six other transcription-related complexes, making interpretation of its role in each of these complexes difficult. Although Taf14 is not conserved as a TFIID subunit in metazoans, it is conserved through its chromatin-binding YEATS domain. Based on the Taf2-Taf14 genetic interaction, we demonstrate that Taf2 and Taf14 directly interact and mapped the Taf2-Taf14 interaction domains. We used this information to identify a Taf2 separation-of-function variant (Taf2-ΔC). Although Taf2-ΔC no longer interacts with Taf14 in vivo or in vitro, it stably incorporates into the TFIID complex. In addition, purified Taf2-ΔC mutant TFIID is devoid of Taf14, making this variant a powerful reagent for determining the role of Taf14 in TFIID function. Furthermore, we characterized the mechanism through which Taf14 suppresses taf2ts alleles, shedding light on how Taf2-Taf14 interaction contributes to TFIID complex organization and identifying a potential role for Taf14 in mediating TFIID-chromatin interactions.
| S-EPMC5077207 | biostudies-literature
Project description:The recognition of core promoter sequences by the general transcription factor TFIID is the first step in the process of RNA polymerase II (Pol II) transcription initiation. Metazoan holo-TFIID is composed of the TATA binding protein (TBP) and of 13 TBP associated factors (TAFs). Inducible Taf7 knock out (KO) results in the formation of a Taf7-less TFIID complex, while Taf10 KO leads to serious defects within the TFIID assembly pathway. Either TAF7 or TAF10 depletions correlate with the detected TAF occupancy changes at promoters, and with the distinct phenotype severities observed in mouse embryonic stem cells or mouse embryos. Surprisingly however, under either Taf7 or Taf10 deletion conditions, TBP is still associated to the chromatin, and no major changes are observed in nascent Pol II transcription. Thus, partially assembled TFIID complexes can sustain Pol II transcription initiation, but cannot replace holo-TFIID over several cell divisions and/or development.
| S-EPMC10705246 | biostudies-literature
Project description: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.
| S-EPMC10404688 | biostudies-literature