Abstract
SUPT16H encodes a subunit of the FACT (FAcilitates Chromatin Transcription) complex, a histone chaperone essential for maintaining chromatin integrity during transcription, replication, and DNA repair. Pathogenic de novo SUPT16H missense variants have previously been linked to neurodevelopmental disorders in eight individuals. Here, we expand the genotypic and phenotypic spectrum by identifying 24 additional individuals harboring ultrarare heterozygous missense or truncating variants, who share overlapping clinical features including intellectual disability, autism spectrum disorder, hypotonia, and characteristic craniofacial dysmorphism. To elucidate the underlying mechanisms, we generated a supt16h knockout zebrafish model using CRISPR/Cas9. The supt16h loss-of-function (LOF) model recapitulated key patient phenotypes such as developmental delay, craniofacial anomalies, and hypotonia. Structural and functional analyses of selected SUPT16H variants demonstrated differential rescue of developmental defects in supt16h-deficient embryos, indicating variant-specific LOF effects in vivo. The presence of non-neural manifestations, including facial and ear anomalies, suggested a role for SUPT16H in neural crest development. Consistently, supt16h loss impaired neural crest cell migration and differentiation and triggered p53-dependent apoptosis in the central nervous system (CNS) and neural crest-derived pharyngeal arches. Notably, supt16h deficiency impaired oligodendrocyte specification in the CNS and perturbed differentiation of neural crest-derived Schwann cells in the peripheral nervous system, providing a plausible basis for hypotonia. These findings uncover a previously unrecognized role of SUPT16H in neural crest development, linking chromatin regulation to neural crest-derived lineage specification and differentiation, thereby defining SUPT16H deficiency as a neurocristopathy that broadens the clinical and mechanistic landscape of SUPT16H-associated disorders.
Oliveira DV, Kato A, Nakamura K, Ikura T, Okada M, Kobayashi J, Yanagihara H, Saito Y, Tauchi H, Komatsu K. Histone chaperone FACT regulates homologous recombination by chromatin remodeling through interaction with RNF20. J Cell Sci. 2014 Feb 15;127(Pt 4):763-72. doi: 10.1242/jcs.135855. Epub 2013 Dec 19. PMID: 24357716.
Abstract
The E3 ubiquitin ligase RNF20 regulates chromatin structure through ubiquitylation of histone H2B, so that early homologous recombination repair (HRR) proteins can access the DNA in eukaryotes during repair. However, it remains unresolved how RNF20 itself approaches the DNA in the presence of chromatin structure. Here, we identified the histone chaperone FACT as a key protein in the early steps of HRR. Depletion of SUPT16H, a component of FACT, caused pronounced defects in accumulations of repair proteins and, consequently, decreased HRR activity. This led to enhanced sensitivity to ionizing radiation (IR) and mitomycin-C in a fashion similar to RNF20-deficient cells, indicating that SUPT16H is essential for RNF20-mediated pathway. Indeed, SUPT16H directly bound to RNF20 in vivo, and mutation at the RING-finger domain in RNF20 abolished its interaction and accumulation, as well as that of RAD51 and BRCA1, at sites of DNA double-strand breaks (DSBs), whereas the localization of SUPT16H remained intact. Interestingly, PAF1, which has been implicated in transcription as a mediator of FACT and RNF20 association, was dispensable for DNA-damage-induced interaction of RNF20 with SUPT16H. Furthermore, depletion of SUPT16H caused pronounced defects in RNF20-mediated H2B ubiquitylation and thereby, impaired accumulation of the chromatin remodeling factor SNF2h. Consistent with this observation, the defective phenotypes of SUPT16H were effectively counteracted by enforced nucleosome relaxation. Taken together, our results indicate a primary role of FACT in RNF20 recruitment and the resulting chromatin remodeling for initiation of HRR.
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