# Regulation of transcription elongation

> **NIH NIH R01** · PENNSYLVANIA STATE UNIVERSITY, THE · 2020 · $306,540

## Abstract

Project Summary
 RNA polymerase (RNAP) pausing and termination are important components of gene expression in all
organisms. NusA and NusG are two general transcription elongation factors that are capable of stimulating
pausing and termination in bacteria. Pausing allows synchronization RNAP position with RNA folding and/or
regulatory factor binding. Intrinsic and Rho-dependent termination are two transcription termination
mechanisms identified in bacteria. Canonical intrinsic terminators consist of an uninterrupted RNA hairpin
followed by a U-tract. Although NusA was known to stimulate intrinsic termination in vitro, since NusA is
essential for viability its role on termination in vivo was not known until recently. RNA-seq studies with a B.
subtilis NusA depletion strain identified a class of intrinsic terminator that requires NusA. NusA-dependent
terminators have weak RNA hairpins and/or poor U-tracts. There is also evidence that NusG stimulates
termination of mycobacterial RNAP at suboptimal intrinsic terminators in vitro. In Rho-dependent termination,
Rho promotes transcript release when it catches up to paused RNAP. E. coli NusG participates in some Rho-
dependent termination events by serving as a bridge between RNAP and Rho.
 NusA and NusG cooperatively stimulate pausing at two sites in the 5'UTR of the B. subtilis trp operon.
NusG makes sequence-specific contacts with the non-template DNA (ntDNA) strand within the paused
transcription bubble. As RNAP and template DNA must move with respect to one another for elongation to
resume, interaction of NusG with both components inhibits elongation. The T-rich ntDNA sequence at the two
pause sites constitutes a conserved NusG recognition motif. NET-seq will be used to identify pause sites
throughout the B. subtilis genome that respond to NusA and/or NusG. The ability to deplete NusA and delete
nusG without growth defects makes B. subtilis the ideal organism for these studies. By combining RNase
footprinting with NET-seq (RNET-seq), the effect of NusA and NusG on the translocation state of RNAP will be
determined at each pause site. Similarly, a comprehensive genomic analysis of the effects of NusA, NusG and
Rho on termination in B. subtilis will be performed using strains containing all combinations of NusA depletion,
nusG and rho alleles. A subset of regulatory pause sites and terminators will then be characterized in vitro.
 A hallmark of transcription attenuation mechanisms is the presence of overlapping antiterminator and
terminator structures that form in the 5'UTR. The 5'UTR of B. subtilis yxjB contains two such sets of
overlapping structures. A model will be tested in which YxjB autoregulates its expression by binding to its
5'UTR and promoting termination at both terminators by preventing formation of the two antiterminators. The
model also posits that the downstream terminator hairpin sequesters the yxjB ribosome binding site. Thus, this
hairpin would repress translation of transcripts ...

## Key facts

- **NIH application ID:** 9856442
- **Project number:** 5R01GM098399-08
- **Recipient organization:** PENNSYLVANIA STATE UNIVERSITY, THE
- **Principal Investigator:** PAUL L BABITZKE
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $306,540
- **Award type:** 5
- **Project period:** 2012-08-01 → 2021-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9856442

## Citation

> US National Institutes of Health, RePORTER application 9856442, Regulation of transcription elongation (5R01GM098399-08). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9856442. Licensed CC0.

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