Instruct-ERIC Webinar Series: Structure Meets Function - Webinar #10

4 May 2021

Featuring expert speakers from Instruct Centres across Europe, Instruct-ERIC Webinar Series: Structure Meets Function highlights some of the latest developments in structural biology, demonstrating how integrative methods are enabling scientists to decipher the mechanisms that underpin health and disease.

Watch the previous webinars in the series here.

The tenth webinar in the series will be hosted by Instruct Centre France 1,  on 12 May 2021, 11:00 - 12:30 CEST.


Webinar moderator: Patrick Schultz, IGBMC Strasbourg

Introduction – Integrated Structural Biology at the Centre for Integrative Biology / IGBMC

Speaker: Bruno Klaholz


Talk 1: Structural basis for the allosteric regulation of Human Topoisomerase 2α

Speaker: Valérie Lamour

Affiliation: IGBMC Strasbourg

Abstract: The human type IIA topoisomerases (Top2) are essential enzymes that regulate DNA topology and chromosome organization. The Top2α isoform is a prime target for antineoplastic compounds used in cancer therapy that form ternary cleavage complexes with the DNA. Despite extensive studies, structural information on this large dimeric assembly is limited to the catalytic domains, hindering the exploration of allosteric mechanism governing the enzyme activities. We produced the entire human Top2α in mammalian cells and present cryo-EM structures of the entire human Top2α nucleoprotein complex in different conformations at subnanometer resolutions. Our data unveils the molecular determinants that fine tune the allosteric connections between the ATPase domain and the DNA binding/cleavage domain. The reconstruction of the DNA binding/cleavage domain at 4.2 Å uncovers a linker leading to the non conserved C-terminal domain, which plays a critical role in modulating the enzyme’s activities and opens perspective for the analysis of post-translational modifications.

Talk 2: Structure of the SAGA transcriptional co-activator

Speaker: Gabor Papai

Affiliation: IGBMC Strasbourg

Abstract: The Spt-Ada-Gcn5 Acetyltransferase (SAGA) transcriptional coactivator is a 19 subunit protein complex interacting with sequence dependent transcriptional activators, to modify chromatin through histone acetylation and deubiquitination. SAGA interacts with TBP and is generally required for optimal expression of the genetic information. We determined the structure of the full P. pastoris SAGA complex at 3.6 Å resolution by single particle cryo-electron microscopy. SAGA is organized into two lobes, one of which is fully occupied by the 430 KDa Tra1 protein, the major target of trans-activator binding. The organization of the second lobe is dictated by the TAF (TBP Associated Factor) module partially shared by TFIID and composed of Taf5 whose WD40 domain orchestrates the position of three pairs of subunits containing a histone fold (HF), namely Taf6-Taf9, Taf12-Ada1 and Taf10-Spt7. This module holds a fourth HF pair found in two copies in the Spt3 subunit and forms an octameric structure reminiscent of the histone octamer. A stable complex was formed with TBP and the cryo-EM structure shows an interaction of Spt3 with the C-terminal stirrup of TBP consistent with genetic and cross-linking data. The yeast-specific Spt8 subunit interacts both with the N-terminal half of TBP and with the C-terminal end of Spt7. The enzymatic histone acetyl transferase and histone H2B deubiquitination modules are anchored onto the TAF module through the Ada3 and the Sgf73 subunits, respectively, but retain outstanding conformational flexibility by virtue of flexible loops connecting the anchoring regions to the catalytic domains.

Talk 3: Structural basis of transcription translation coupling and collision in bacteria

Speaker: Albert Weixlbaumer

Affiliation: IGBMC Strasbourg

Abstract: Transcription and translation had first been proposed to be coupled processes in prokaryotes in 1964. Early electron micrographs (Miller spreads) confirmed this and suggested that messenger RNA (mRNA) is translated by the ribosome, while still being transcribed by RNA polymerase (RNAP). The pioneering ribosome may contact RNAP, forming a supramolecular complex known as the expressome. However, the structural basis of expressome assembly and its consequences for transcription and translation are poorly understood. We obtained a series of cryo-EM reconstructions representing uncoupled, coupled and collided expressome states for the first time at high resolution. The transcription factor NusG forms a physical connection between RNAP and the ribosome to stabilize an otherwise variable interaction interface. Shortening of the intervening mRNA rearranges the assembly and aligns the ribosome entrance-channel to the RNAP exit-channel. In this collided complex, NusG-linkage is no longer possible. These structures reveal mechanisms of coordination between transcription and translation and provide a framework for future study.

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