Biology subjects

Rengachari, S.

Publications and source records attributed to Rengachari, S..

4 recordsLinked to original sources

bioRxiv2025

Structure of human cytoplasmic Pol II complex explains global transcription repression by Gdown1

RNA polymerase II (Pol II) is a 12-subunit enzyme crucial for gene transcription in the nucleus. However, its assembly in the cytoplasm, nuclear import, and nuclear function of assembly factors remain poorly understood. Here, we isolated Pol II from the cytoplasmic fraction of human cells (cfPol II) and determined its cryo-EM structure. The structure reveals that Pol II is fully assembled in the cytoplasm before nuclear import. We also found that Gdown1 binds Pol II through three distinct regions, indicating it may stabilize Pol II assembly intermediates. Notably, Gdown1 binding precludes the association of essential transcription factors IIB and IIF, rendering cfPol II inactive in promoter-dependent transcription in vitro. Our results provide a basis for Gdown1-dependent global transcription repression and suggest a model for the role of Gdown1 in Pol II assembly, import, and transcription regulation.

biochemistry↗

bioRxiv2022

Structural basis of SNAPc-dependent snRNA transcription initiation by RNA polymerase II

RNA Polymerase II (Pol II) carries out transcription of both protein-coding and non-coding genes. Whereas Pol II initiation at protein-coding genes has been studied in detail, Pol II initiation at non-coding genes such as small nuclear RNA (snRNA) genes is not understood at the structural level. Here we study Pol II initiation at snRNA gene promoters and show that the snRNA-activating protein complex (SNAPc) enables DNA opening and transcription initiation independent of TFIIE and TFIIH in vitro. We then resolve cryo-EM structures of the SNAPc-containing Pol II preinitiation complex (PIC) assembled on U1 and U5 snRNA promoters. The core of SNAPc binds two turns of DNA and recognizes the snRNA promoter-specific proximal sequence element (PSE) located upstream of the TATA box-binding protein TBP. Two extensions of SNAPc called wing-1 and wing-2 bind TFIIA and TFIIB, respectively, explaining how SNAPc directs Pol II to snRNA promoters. Comparison of structures of closed and open promoter complexes elucidates TFIIH-independent DNA opening. These results provide the structural basis of Pol II initiation at non-coding RNA gene promoters.

molecular biology↗

bioRxiv2021

Structure of human Mediator-RNA polymerase II transcription pre-initiation complex

Mediator is a conserved coactivator that enables regulated transcription initiation from eukaryotic protein-coding genes1-3. Mediator is recruited by transcriptional activators and binds the pre-initiation complex (PIC) to stimulate RNA polymerase II (Pol II) phosphorylation and promoter escape1-6. Here we prepare a 20-subunit recombinant human Mediator, reconstitute a 50-subunit Mediator-PIC complex, and resolve the complex structure by cryo-EM at an overall resolution of 4.5 [A]. Mediator binds with its head module to the Pol II stalk and the general transcription factors TFIIB and TFIIE, resembling the Mediator-PIC interactions in the corresponding yeast complex7-9. One end of Mediator contains the metazoan-specific subunits MED27-MED30, which associate with exposed regions in MED14 and MED17 to form the proximal part of the tail module that binds activators. The opposite end of Mediator positions the flexibly linked CDK-activating kinase (CAK) of the general transcription factor TFIIH near the C-terminal repeat domain (CTD) of Pol II. The Mediator shoulder domain holds the CAK subunit CDK7, whereas the hook domain contacts a CDK7 element that flanks the kinase active site. The shoulder and hook reside in the Mediator head and middle modules, respectively, which can move relative to each other and may induce an active conformation of CDK7 to allosterically stimulate CTD phosphorylation and Pol II escape from the promoter.

molecular biology↗

bioRxiv2017

Structural Basis Of STAT2 Recognition By IRF9 Reveals Molecular Insights Into ISGF3 Function

Cytokine signalling is mediated by the activation of distinct sets of structurally homologous JAK and STAT signalling molecules, which control nuclear gene expression and cell fate. A significant expansion in the gene regulatory repertoire controlled by JAK/STAT signalling has arisen by the selective interaction of STATs with IRF transcription factors. Type I interferons (IFN), the major antiviral cytokines, trigger the formation of the ISGF3 complex containing STAT1, STAT2 and IRF9. ISGF3 regulates the expression of IFN-stimulated genes (ISGs). ISGF3 assembly depends on selective interaction between IRF9, through its IRF-association domain (IAD), with the coiled-coil domain (CCD) of STAT2. Here, we report the crystal structures of the IRF9-IAD alone and in a complex with STAT2-CCD. Despite similarity in the overall structure among respective paralogs, the surface features of the IRF9-IAD and STAT2- CCD have diverged to enable specific interaction between these family members, thus enabling ISGF3 formation and expression of ISGs.

biophysics↗