# Systems biological interrogation of bacterial persistence

> **NIH NIH R01** · COLUMBIA UNIVERSITY HEALTH SCIENCES · 2021 · $530,830

## Abstract

Summary
Antibiotic persistence remains one of the most challenging barriers to effective clearance of chronic bacterial
infections. Despite the intervening decades since its original discovery, we still lack a basic understanding of
how molecular networks implement the defining features of persistence: phenotypic heterogeneity and tolerance
to lethal levels of antibiotic exposure. Much of the work on persistence has been phenomenological and the
mechanistic studies which, so far, have only focused on a few candidate pathways, have failed to provide an
adequate understanding of this complex phenotype. Here, we propose an unbiased systems biology approach
to characterize the genetic and regulatory underpinnings of persistence. A primary aim is to precisely define the
cellular state of persister cells which typically make up only a small (< 10-4) fraction of the population. We will
utilize our previously developed genetic and chemically induced models of E. coli hyper-persistence to trigger
persister cells, and to follow the trajectory of their phenotypic diversification by using our recently developed
single-cell RNA sequencing technology (PETRI-seq). This will enable us to define the dynamics of the process
at single-cell resolution, and identify highly specific persister markers, which we can use to purify them to near-
homogeneity. We will then profile the global gene regulatory state of persisters by using our recently optimized
technologies for in vivo Protein-DNA and RNA-RNA interactions. Combining the single-cell RNA atlas and global
regulatory interactions will enable us to generate causal graphical models of pivotal regulatory events that
underlie persister formation. We will utilize our recently developed CRISPR-interference technology (CALM) to
systematically determine how knock-down perturbations to all essential and non-essential genes affect
quantitative parameters of persistence, including persister-fraction and kill-rates. Finally, by combining the
regulatory and genetic maps of persistence, we will identify and validate the most critical vulnerability nodes
whose targeting will eliminate persister formation and survival. The massive scale of these observations will
reveal the most comprehensive and unbiased global view of persister generation, gene regulation, and
physiology, to date. This is a critical foundation upon which we can devise rational strategies for reducing the
clinical burden of persisters. Finally, the unique conceptual and technological approaches here will serve as a
blueprint for exploring the genetic and regulatory basis of other complex bacterial phenomena, such as biofilms,
where phenotypic heterogeneity is a defining hallmark.

## Key facts

- **NIH application ID:** 10264904
- **Project number:** 5R01AI077562-12
- **Recipient organization:** COLUMBIA UNIVERSITY HEALTH SCIENCES
- **Principal Investigator:** Saeed F Tavazoie
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $530,830
- **Award type:** 5
- **Project period:** 2009-06-01 → 2025-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10264904

## Citation

> US National Institutes of Health, RePORTER application 10264904, Systems biological interrogation of bacterial persistence (5R01AI077562-12). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10264904. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*