Reduced histone gene copy number disrupts Polycomb function.

The chromatin of animal cells contains two types of histones: canonical histones that are expressed during S phase of the cell cycle to package the newly replicated genome, and variant histones with specialized functions that are expressed throughout the cell cycle and in non-proliferating cells. Determining whether and how canonical and variant histones cooperate to regulate genome function is integral to understanding how chromatin-based processes affect normal and pathological development. Here, we demonstrate that variant histone H3.3 is essential for development only when canonical histone gene copy number is reduced, suggesting that coordination between canonical and variant expression is necessary to provide sufficient H3 protein for normal genome function. To identify genes that depend upon, or are involved in, this coordinate regulation we screened for heterozygous chromosome 3 deficiencies that impair development of flies bearing reduced and gene copy number. We identified two regions of chromosome 3 that conferred this phenotype, one of which contains the gene, which is necessary for establishing domains of facultative chromatin that repress master regulator genes during development. We further found that reduction in dosage decreases viability of animals with no gene copies. Moreover, heterozygous mutations result in de-repression of the Polycomb target gene and cause ectopic sex combs when either canonical or variant gene copy number is also reduced. We conclude that Polycomb-mediated facultative heterochromatin function is compromised when canonical and variant gene copy number falls below a critical threshold.