# Cellular homeostasis pathways in bacteria

> **NIH NIH R35** · UNIVERSITY OF CALIFORNIA, SAN FRANCISCO · 2023 · $943,376

## Abstract

The overarching goal of my research is to uncover the fundamental principles that drive bacterial success,
enabling them to colonize and proliferate in every corner of this planet. To accomplish this goal, I have
developed a unified bacterial-centric research vision cutting across classically defined subfields. I meld my long
history of unraveling the intricacies of bacterial control mechanisms with an ability to develop and implement
novel global technologies to open up understudied areas and computational approaches to extend findings
beyond model organisms. The current grant explores three important and related areas.
 First, fueled by two novel CRISPRi strategies that we developed, we continue our quest to identify
cellular construction principles by exploring three understudied sets of genes: cell envelope genes, essential
genes, and genes that accelerate growth transitions. We tackle the redundancy of function that has prevented
genetic analysis of the envelope with double CRISPRi, a technology that allows simultaneous knockdown of
two genes via adjacently encoded sgRNAs. We unravel the tradeoffs underlying the expression of essential
genes with mismatched CRISPRi, which uses single mismatches in the base pairing region of sgRNAs to
predictably titrate their efficacy. By measuring the fitness impact of graded knockdown, we determine the
expression-fitness relationships of essential genes and how they are affected by environmental and genetic
changes. Finally, we identify essential and non-essential genes that accelerate growth transitions.
 Second, we continue our studies of the general principles controlling translational output both by
exploring the extent to which ribosomes themselves influence the upstream process of transcription
(transcription/translation coupling) and the downstream process of mRNA degradation, and by determining
whether alternative ribosomal proteins produced under stress conditions result in new translational properties.
These studies are enabled by new technologies we developed for genome-wide measurement of ribosome
spacing and mRNA degradation.
 Third, we have begun an exciting new study of gene regulatory networks throughout the bacterial
kingdom. This effort is fueled by our new statistically rigorous, phylogenetic foot-printing approach, which we
have validated to have a low false positive rate coupled with high recall and precision. We plan to leverage the
vast existing database of bacterial genomes to examine evolution of gene regulatory networks across bacteria.
 Our studies also address an overwhelming current challenge: to develop experimental and
computational approaches that enable researchers to comprehensively explore the regulatory wiring and
functional diversity of bacteria that thrive in a wide variety of rapidly changing environments. Such approaches
can synergize with and exploit metagenomic data to empower mechanistic interrogation of gene function in
understudied organisms.

## Key facts

- **NIH application ID:** 10661724
- **Project number:** 5R35GM118061-08
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
- **Principal Investigator:** CAROL Anne GROSS
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2023
- **Award amount:** $943,376
- **Award type:** 5
- **Project period:** 2016-06-07 → 2026-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10661724

## Citation

> US National Institutes of Health, RePORTER application 10661724, Cellular homeostasis pathways in bacteria (5R35GM118061-08). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10661724. Licensed CC0.

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