The epigenome is sensitive to the availability of metabolites that serve as substrates of chromatin-modifying enzymes. Links between acetyl-CoA metabolism, histone acetylation, and gene regulation have been documented, although how specificity in gene regulation is achieved by a metabolite has been challenging to answer. Recent studies suggest that acetyl-CoA metabolism is tightly regulated both spatially and temporally to elicit responses to nutrient availability and signaling cues. Here we discuss evidence that acetyl-CoA production is differentially regulated in the nucleus and cytosol of mammalian cells. Recent findings indicate that acetyl-CoA availability for site-specific histone acetylation is influenced through post-translational modification of acetyl-CoA-producing enzymes, as well as through dynamic regulation of the nuclear localization and chromatin recruitment of these enzymes.