# Prime editing links the split integrated stress response to pathogenic eIF2B mutations and white matter degeneration

## TL;DR

This study uses prime editing to model a rare brain disorder and finds a new stress response pathway linked to disease progression.

## Contribution

First use of prime editing to model Vanishing White Matter Disease and demonstrates the split ISR's role in human cells.

## Key findings

- All modeled eIF2B mutations activate the split ISR in human cells.
- Split ISR impairs protein synthesis needed for proper cell differentiation.
- ISRIB can suppress the split ISR and its harmful effects.

## Abstract

Vanishing White Matter Disease (VWMD) is a devastating, currently incurable neurodevelopmental disorder primarily affecting white matter. The prevailing view attributes VWMD to the activation of the canonical integrated stress response (c-ISR). However, recent studies have identified a novel, distinct pathway called the split ISR (s-ISR), though its activation has so far only been documented in mouse stem cells harboring a single eIF2B mutation, leaving uncertainty about whether it occurs in human cells, whether other mutations can trigger it, and what role it plays in the disease. Here, we used prime editing (PE) to engineer multiple eIF2B pathogenic mutations into HEK293T and induced pluripotent stem cells (iPSCs), generating human models. We demonstrated PE’s effectiveness and safety, marking the first successful application of PE for modeling VWMD. We found that all modeled mutations activate the s-ISR, indicating that this response is a common feature across VWMD mutations, and that it can be further amplified by stress-induced c-ISR and effectively suppressed by ISRIB. Mechanistically, we show that s-ISR hinders mutant iPSCs from achieving the high protein synthesis levels necessary for proper differentiation, expecially into astrocytes. This impairment disrupts their maturation process, directly linking s-ISR activation to the white matter abnormalities of VWMD.