Louder, Robert K. | Search NASA

DOE OSTI2021

Structure of the human SAGA coactivator complex

The SAGA complex is a regulatory hub involved in gene regulation, chromatin modification, DNA damage repair and signaling. While structures of yeast SAGA (ySAGA) have been reported, there are noteworthy functional and compositional differences for this complex in metazoans. Here we present the cryogenic-electron microscopy (cryo-EM) structure of human SAGA (hSAGA) and show how the arrangement of distinct structural elements results in a globally divergent organization from that of yeast, with a different interface tethering the core module to the TRRAP subunit, resulting in a dramatically altered geometry of functional elements and with the integration of a metazoan-specific splicing module. Our hSAGA structure reveals the presence of an inositol hexakisphosphate (InsP 6 ) binding site in TRRAP and an unusual property of its pseudo-(Ψ)PIKK. Finally, we map human disease mutations, thus providing the needed framework for structure-guided drug design of this important therapeutic target for human developmental diseases and cancer.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2020

The gammaherpesviral TATA-box-binding protein directly interacts with the CTD of host RNA Pol II to direct late gene transcription

β- and γ-herpesviruses include the oncogenic human viruses Kaposi’s sarcoma-associated virus (KSHV) and Epstein-Barr virus (EBV), and human cytomegalovirus (HCMV), which is a significant cause of congenital disease. Near the end of their replication cycle, these viruses transcribe their late genes in a manner distinct from host transcription. Late gene transcription requires six virally encoded proteins, one of which is a functional mimic of host TATA-box-binding protein (TBP) that is also involved in recruitment of RNA polymerase II (Pol II) via unknown mechanisms. Here, we applied biochemical protein interaction studies together with electron microscopy-based imaging of a reconstituted human preinitiation complex to define the mechanism underlying Pol II recruitment. These data revealed that the herpesviral TBP, encoded by ORF24 in KSHV, makes a direct protein-protein contact with the C-terminal domain of host RNA polymerase II (Pol II), which is a unique feature that functionally distinguishes viral from cellular TBP. The interaction is mediated by the N-terminal domain (NTD) of ORF24 through a conserved motif that is shared in its β- and γ-herpesvirus homologs. Thus, these herpesviruses employ an unprecedented strategy in eukaryotic transcription, wherein promoter recognition and polymerase recruitment are facilitated by a single transcriptional activator with functionally distinct domains.

59 BASIC BIOLOGICAL SCIENCES↗