# Molecular mechanisms of memory formation and tolerance in CRISPR-Cas systems

> **NIH GM R00** · SLOAN-KETTERING INST CAN RESEARCH · 2026 · $249,000

## Abstract

Project Summary
CRISPR-Cas are prokaryotic adaptive immune systems that protect bacteria and archaea from invading mobile
genetic elements, such as phages and plasmids. CRISPR-Cas systems acquire immunological memories during
infection by integrating short fragments from the invader’s genome into the CRISPR locus of the host. These
fragments, called “spacers”, are later transcribed into CRISPR RNAs that are loaded on Cas nucleases and
guide them to recognize and cleave infecting nucleic acids. Depending on their genetic composition, CRISPR-
Cas systems are classified into six types (I-VI). While spacer acquisition has been extensively studied in type I
and II systems, type III systems are just now starting to be explored. The overall goal of this application is to
define the molecular mechanisms that govern spacer acquisition by the prevalent, yet less studied, type III-A
CRISPR-Cas system, and understand its implications during CRISPR-Cas defense and tolerance. Preliminary
work on the type III-A system of Staphylococcus epidermidis revealed that this system preferentially acquires
new spacers by two independent modes. The first mode acquires spacers from some, but not all, highly
transcribed genes, and spans their entire transcribed region. The first aim of this proposal is to elucidate how
the acquisition machinery recognizes specific genes as substrates for preferential acquisition. This will be
achieved by dissecting the DNA sequences that recruit the spacer-integrase complex to specific genes, finding
host factors that mediate gene-specific spacer acquisition, and test for the physiological relevance of this process
during the CRISPR-Cas immune response. The second mode of acquisition by the type III-A system is similar
to the previously studied type I and II systems, where spacers are acquired from free dsDNA ends at the bacterial
chromosomal terminus, in a manner that is dependent on the cell’s DNA-repair machinery. Such self-targeting
spacers are expected to

## Key facts

- **NIH application ID:** 11308749
- **Project number:** 5R00GM148720-04
- **Recipient organization:** SLOAN-KETTERING INST CAN RESEARCH
- **Principal Investigator:** Naama  Aviram
- **Activity code:** R00 (R01, R21, SBIR, etc.)
- **Funding institute:** GM
- **Fiscal year:** 2026
- **Award amount:** $249,000
- **Award type:** 5
- **Project period:** 2023-09-01T00:00:00 → 2028-03-31T00:00:00

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/11308749

## Citation

> US National Institutes of Health, RePORTER application 11308749, Molecular mechanisms of memory formation and tolerance in CRISPR-Cas systems (5R00GM148720-04). Retrieved via AI Analytics 2026-07-01 from https://api.ai-analytics.org/grant/nih/11308749. Licensed CC0.

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