Histone variant H2A.Z.2 primes MITF promoter to direct specification of melanocyte precursors

Stem cell establishment and fate assignment are critical for both early development, as well as regeneration. A committed cell state is characterized by its unique transcriptional landscape. In contrast, the upstream fate-biased state is often metastable and transitory, thereby mechanisms of specification are difficult to identify. In this study, we elucidate the role of a histone variant H2a.z.2 in specification of melanocyte lineage from multipotent neural crest cells. Silencing of H2a.z.2 reduces the number of melanocyte precursors in developing zebrafish embryos, and from mouse embryonic stem cells in vitro. We demonstrate that this histone variant occupies nucleosomes in the promoter of key melanocyte determinant Mitf, and enhances its induction. CRISPR-Cas9 based targeted mutagenesis of this gene in zebrafish drastically reduces adult melanocytes, as well as their regeneration. Thereby our study establishes a histone based specification code upstream to the core gene regulatory network in the neural crest lineage leading to melanocytes. This epigenetic code renders the promoter of key determinant in a poised state conducive for activation by external instructive signals and facilitates establishment of downstream cell fate. Total RNA was extracted from Control morpholino and h2a.z.2 morpholino injected sox10:GFP positive neural crest cells at 16-18hpf. The RNA was quality controlled and quantified using agilent bioanalyzer. The samples were analyzed by Agilent_zebrafish_GXP_8X60K array. Two biological replicates each of control and h2a.z.2 morpholino injected sox10:GFP positive cells were analyzed. We acknowledge Genotypic Technology Private Limited Bangalore for the microarray processing and data analysis reported in this publication.