GSE196438
Alkaline nucleoplasm facilitates contractile gene expression in the mammalian heart [ChIP-seq]
Cardiac contractile strength is recognised as highly pH-sensitive, but less is known about the influence of pH on cardiac gene expression, which may be relevant during changes in myocardial metabolism or vascularization in development or disease. We sought evidence for pH-responsive cardiac genes and a context in which this has physiological relevance. pHLIP, a peptide-based reporter of acidity, revealed a non-uniform pH landscape in early-postnatal myocardium, dissipating in later life. pH-responsive differentially-expressed genes (pH-DEGs) were identified by transcriptomics of neonatal cardiomyocytes cultured over a range of pH.  Enrichment analysis indicated “striated muscle contraction” as a pH-responsive biological process. Label-free proteomics verified fifty-four pH-responsive gene-products, including contractile elements and the adaptor protein CRIP2.  Using transcriptional assays, acidity was found to inhibit p300/CBP acetylase activity and, as its functional readout, negatively affect myocardin, a co-activator of cardiac gene expression. In cultured myocytes, acid-inhibition of p300/CBP reduced H3K27 acetylation, as demonstrated by chromatin immunoprecipitation.  H3K27ac levels were more strongly reduced at promoters of acid-downregulated DEGs, suggesting an epigenetic mechanism of pH-sensitive gene expression.  By tandem cytoplasmic/nuclear pH imaging, the cardiac nucleus was found to exercise a degree of control over its pH through Na+/H+ exchangers at the nuclear envelope. Thus, we describe how extracellular pH signals gain access to the nucleus and regulate the expression of a subset of cardiac genes, notably those coding for contractile proteins and Crip2. Acting as a proxy of a well-perfused myocardium, alkaline conditions are permissive for expressing genes related to the contractile apparatus. Overall design: ChIP-seq in NRVMs at various pHs or treated with A485