ABSTRACT: The 26S proteasome consists of 20S proteolytic core particle (CP) and a 19S regulatory particle
(RP). It is engaged in degradation of a variety of proteins, and thus, regulates many important cellular
processes. Importantly, our results reveal that the 19S RP is recruited to the upstream activating sequence to
facilitate transcription complex assembly at the core promoter to stimulate transcription initiation by enhancing
the targeting of co-activators, SAGA (Spt-Ada-Gcn5-acetyltransferase) or TFIID (Transcription factor IID) to the
activator at the SAGA or TFIID-regulated genes, respectively, in a proteolysis-independent manner.
Intriguingly, transcription initiation has also been shown to be promoted by the proteolytic activity of the
proteasome. However, it is not clearly understood how the 26S proteasome promotes transcription initiation.
We hypothesize that proteasome controls transcription initiation by regulating co-activator via ubiquitylation
and proteasomal degradation. Indeed, our preliminary results revealed that the Sgf73 component of the co-
activator, SAGA, undergoes ubiquitylation and proteasomal degradation, thus supporting our hypothesis.
However, the E3 ubiquitin ligase involved in such regulation of Sgf73 is yet unknown. Further, how this ligase
interacts with and is targeted to Sgf73 for ubiquitylation and proteasomal degradation, and the physiological
relevance of such regulation of Sgf73 on SAGA’s integrity (and hence its functions in regulation of chromatin
modification and transcription initiation) remain largely elusive. Moreover, other factors such as ubiquitin
protease and conjugase among others involved in Sgf73 ubiquitylation and proteasomal degradation are not
known. Answer to these important questions would fundamentally develop novel ubiquitin-proteasome system
regulation of SAGA in orchestrating chromatin modification and transcription, thus greatly advancing the field of
gene regulation. In addition, these results would have significant impact on disease pathogenesis and future
therapeutic development, since SAGA as well as Sgf73 are evolutionarily conserved from yeast to humans,
and associated with various diseases. Therefore, we propose to address above questions in this application.
Specifically, we will (i) identify E3 ubiquitin ligase involved in Sgf73 ubiquitylation and proteasomal degradation,
(ii) determine how Sgf73 recognizes E3 ligase, (iii) determine ubiquitylation site(s) on Sgf73, (iv) determine the
physiological relevance/role of Sgf73 ubiquitylation and proteasomal degradation in regulation of SAGA’s
integrity, chromatin modification and transcription, and (v) identify and characterize ubiquitin conjugase and
ubiquitin protease involved in regulation of Sgf73 ubiquitylation and proteasomal degradation, and hence
SAGA and its functions. Collective results would identify ubiquitin ligase, conjugase and protease in regulation
of ubiquitylation and proteasomal degradation of SAGA component, Sgf73, with roles in SAGA’s integrity and
functions in gene expression, thus advancing our understanding of gene regulation by ubiquitin-proteasome
system with implications in human health.
