# Mechanistic studies of transcription initiation and elongation functions of an RNA polymerase II variant, Pol II(G), that is implicated in development and cancer

> **NIH NIH R01** · ROCKEFELLER UNIVERSITY · 2024 · $368,346

## Abstract

Eukaryotic RNA polymerase II (Pol II) plays a pivotal role in transcription. Normal physiological processes
depend upon precise transcriptional controls, whereas transcriptional dysregulation is the basis of numerous
pathologies that include cancer. Pol II recruitment to specific promoters is regulated by multiple cofactors that
include the multi-subunit Mediator, which directly binds both to enhancer/promoter-bound
transcriptional activators and to Pol II to facilitate gene activation. Following initiation and promoter escape,
Pol II remains subject to regulation by multiple elongation factors, acting either at Pol II pause-release or
productive elongation steps. Pol II(G) is a recently described form of Pol II that contains the tightly
associated, metazoan-specific Gdown1 polypeptide along with the normal 12 subunits. Our genetic-
based studies of Pol II(G) have demonstrated that Gdown1 is essential for early embryonic development
and for cell-specific transcription in quiescent hepatocytes, in which heavy localization to gene bodies of
highly expressed liver-specific genes (e.g., albumin) is indicative of elongation functions and in which ablation
leads to downregulation of both liver-specific and lipid metabolism genes, cell cycle re-entry and (in the
absence of p53) a premalignant type of transformation. Studies in hepatocarcinoma and breast cancer cells
have also indicated a key role for Gdown1 in cell growth and in expression of lipid metabolism genes, which
are generally important for maintenance of cancer cell growth. Our biochemical studies have revealed that
the Pol II-associated Gdown1 conditionally represses basal (activator- and Mediator-independent)
transcription initiation by preventing association of TFIIB and TFIIF with Pol II, thereby establishing a potential
checkpoint and eliciting a strong requirement for activator-bound Mediator to overcome repression. Our
structural studies have defined Gdown1 interaction sites on Pol II and provided clues regarding Mediator
interactions that might facilitate its reversal of the conditionally repressed initiation capacity of Pol II(G),
although the underlying mechanism remains unclear. With the general objective of understanding the
molecular mechanisms of action of Pol II(G) in conjunction with its roles in breast cancer and hepatocarcinoma
cells, especially on Gdown1-regulated cell-specific and lipid metabolism genes, as a potential basis for new
cancer therapeutics, our specific aims are: (i) to investigate the mechanisms underlying Mediator-dependent
transcription initiation and post-initiation events by Pol II(G), including concomitant, newly described
interactions with general transcription factors and elongation factor TFIIS, using powerful in vitro transcription
and immobilized template assays and CX-MS and cryo-EM structural analyses of interacting complexes and (ii)
to investigate Gdown1 functions in hepatocarcinoma cells in promoter-proximal pausing, pause release and
transcript...

## Key facts

- **NIH application ID:** 10926851
- **Project number:** 5R01CA273709-03
- **Recipient organization:** ROCKEFELLER UNIVERSITY
- **Principal Investigator:** ROBERT G ROEDER
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $368,346
- **Award type:** 5
- **Project period:** 2022-07-25 → 2027-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10926851

## Citation

> US National Institutes of Health, RePORTER application 10926851, Mechanistic studies of transcription initiation and elongation functions of an RNA polymerase II variant, Pol II(G), that is implicated in development and cancer (5R01CA273709-03). Retrieved via AI Analytics 2026-06-11 from https://api.ai-analytics.org/grant/nih/10926851. Licensed CC0.

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