Spatially concentrated adenine base editors efficiently correct <i>PLP1</i> mutations in oligodendrocytes

Oligodendrocytes (OLs), the myelinating cells of the central nervous system, are particularly prone to pathogenic G-to-A mutations, such as PLP1A243V, which causes Pelizaeus-Merzbacher disease (PMD), a lethal hypomyelinating disorder lacking effective therapy. Although adenine base editors (ABEs) can in principle correct such mutations, their application in OLs is limited by inefficient on-target editing. Here, we develop a spatially concentrated ABE (cABE) strategy that enhances editing by promoting nuclear translocation of tRNA adenosine deaminase (TadA*) from the cytoplasm. Using a SunTag-based multivalent recruitment system, TadA* is locally enriched at genomic targets (cABE-1.0), achieving robust editing in vitro. To enable in vivo delivery while preserving high efficiency and fidelity, SpCas9 is replaced with compact eNme2-C Cas9, generating an AAV-compatible variant (cABE-2.0). Notably, cABE-2.0 forms dynamic nuclear puncta with properties of liquid-liquid phase separation, enhancing on-target editing while substantially reducing transcriptome-wide RNA off-target effects. Functionally, cABE-2.0 efficiently corrects the PLP1A243V mutation in OLs, restores Plp subcellular localization, and rescues myelination-related phenotypes. These findings demonstrate that spatial reorganization, rather than increasing intrinsic catalytic activity of TadA*, provides a distinct principle for improving base editing in difficult-to-edit cell types, such as OLs, offering a mechanistic and technical framework for gene therapy of PMD and related myelin disorders.