H2A.Z histone variant is encoded by two genes,
H2afz and
H2afv, that originate two distinct protein isoforms differing by 3 aa, H2A.Z-1, and H2A.Z-2, respectively
[22]. The H2A.Z histone variant is mostly enriched at TSS, promoters, enhancers, facultative heterochromatin, and centromeres, and was linked to both transcriptional activation and repression, in which H2A.Z-associated PTMs play a major role
[23]. H2A.Z is expressed in C2C12 myoblasts and silencing its expression does not interfere with myoblast differentiation or myotube formation, which shows that its continued expression is not required for differentiation
[24]. A non-acetylatable form of H2A.Z (where the 5 tail lysines are mutated into arginines) fused to GFP (H2A.Z-Ac-mut-GFP) can be incorporated into the genome in a similar fashion as the wild type H2A.Z fused to GFP (H2A.Z-GFP), including at the
Myod1 and
Myog loci (
Figure 2A). H2A.Z-Ac-mut-GFP-expressing myoblasts display reduced myogenic gene expression, such as
Myod1,
Myog, and
Myh3, when triggered to differentiate. Impaired myogenesis is associated with the lack of RNA polymerase II recruitment in the presence of H2A.Z-Ac-mut-GFP to myogenic gene loci upon differentiation
[24]. Consequently, H2A.Z-Ac-mut-GFP overexpression in C2C12 cells blocks myotube formation while H2A.Z-GFP overexpression does not significantly interfere with this process, which shows the H2A acetylation regulates differentiation
[24]. This shows that acetylation of the histone variant H2A.Z plays a role in transcription initiation of myogenic gene expression. The exchange of H2A for its variant H2A.Z is modulated by the chromatin remodeling complex SNF2-related CBP activator protein (SRCAP), a mechanism conserved in yeast and in mammals
[25][26][27]. ChIP-seq analysis of C2C12 cells confirmed the presence of p18
Hamlet (ZNHI1), a component of the SRCAP complex and a substrate of the p38 MAPK pathway, at the
Myog promoter in differentiating C2C12 cells
[25]. In addition, p18
Hamlet is required for the incorporation of H2A.Z at the
Myog promoter, and the enrichment of these two proteins at this genomic region increases during differentiation of C2C12 cells and murine primary myoblasts. Phosphorylation of p18
Hamlet by p38 is required for its recruitment, and for the incorporation of H2A.Z in the
Myog promoter, which suggests that p38 MAPK-dependent signals can impact on chromatin structural changes
[25]. Inhibition of the expression of components of the SRCAP complex leads to impaired myogenic gene expression and blocks myoblast differentiation
[25]. Myofibroblast differentiation that relies on TGF-β1 expression is also regulated by H2A.Z occupancy
[28]. In contrast, in this case, in order to facilitate TGF-β1 expression, H2A.Z must be depleted from the TGF-β1 promoter region through a mechanism that requires the ribosomal function regulator ELF6 (Yang et al., 2015).