# Systematic discovery of critical oligodendrocyte transcription factors by a CRISPRi screen

> **NIH NIH R21** · STATE UNIVERSITY OF NEW YORK AT BUFFALO · 2020 · $438,625

## Abstract

Project Summary
Myelination of the central nervous system (CNS) by oligodendrocytes (OLs) is essential for the development
and function of the CNS. Myelin develops in the CNS as OLs differentiate into the myelin-forming mature
phenotype. The transcriptional regulatory network governing OL differentiation remains incompletely
understood. To address this fundamental issue, an innovative CRISPRi (CRISPR interference) screen has
been developed that can identify novel transcription factors that are important for OL differentiation. In this
screen, the expression of a candidate transcription factor is knocked down by CRISPRi, a potent gene
knockdown technique, and its impact on the differentiation of primary mouse OPCs is measured by the
reporter activity of Rffl, a well-characterized Myrf luciferase reporter. Myrf is a key OL transcription factor that is
indispensable for OL differentiation and CNS myelination. Myrf expression marks the onset of OL
differentiation. Rffl is an OL enhancer in the Rffl locus that is bound and activated by Myrf in differentiating OLs.
The activation of Rffl by Myrf is highly specific and sensitive. These features make Rffl an ideal luciferase
reporter for OL differentiation, making it possible to read out OL differentiation by a simple luciferase assay.
Capitalizing on CRISPRi and Rffl as well as in-house optimized protocols for primary mouse OPCs, the
CRISPRi screen has so far tested 87 transcription factors that are highly expressed in OL lineage cells. Well-
characterized OL transcription factors such as YY1, Sox10, and Myrf came out positive. The screen has also
uncovered 8 novel transcription factors that are required for OL differentiation, including Rnf10. These results
demonstrate the validity and effectiveness of the CRISPRi screen, justifying its further application (Aim I).
Rnf10 is a transcriptional regulator that plays a pivotal role in neuronal differentiation and synaptonuclear
communication. However, nothing is known about its role in OL lineage cells. Since the preliminary data
strongly suggest a crucial role of Rnf10 in OL differentiation, its role in CNS myelination will be determined
(Aim II).

## Key facts

- **NIH application ID:** 9874221
- **Project number:** 1R21NS114476-01
- **Recipient organization:** STATE UNIVERSITY OF NEW YORK AT BUFFALO
- **Principal Investigator:** YUNGKI PARK
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $438,625
- **Award type:** 1
- **Project period:** 2019-12-01 → 2022-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9874221

## Citation

> US National Institutes of Health, RePORTER application 9874221, Systematic discovery of critical oligodendrocyte transcription factors by a CRISPRi screen (1R21NS114476-01). Retrieved via AI Analytics 2026-06-11 from https://api.ai-analytics.org/grant/nih/9874221. Licensed CC0.

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