Neofunctionalization of H1 linker histones drives divergent gene expression and histone methylation in Cryptococcus neoformans.

The opportunistic pathogen Cryptococcus neoformans adapts to diverse host microenvironments during infection, yet the contribution of epigenetic mechanisms remains unclear. Comparative genomics across 50 basidiomycete species revealed that one chromatin protein, the linker histone H1, is uniquely duplicated across the entire Cryptococcus genus, generating paralogs H1.51 and H1.52 with divergent evolutionary trajectories. Molecular evolution analyses revealed that H1.51 experienced intensified selection with episodic positive selection in clinical isolates, while H1.52 evolved under sustained purifying selection, indicating adaptive neofunctionalization. H1.52 deletion triggered transcriptional reprogramming of 1561 genes (23% of the transcriptome), with downregulation of nucleolar function, ribosome biogenesis, and translation machinery, whereas H1.51 loss produced minimal transcriptional effects. We found that H1.52 governs chromatin architecture. Its deletion expanded H3K4me2 euchromatin coverage 1.54-fold (from 18.4% to 28.4% of the genome) into broader constitutive domains with loss of H3K9-marked heterochromatin islands. Following prolonged culture under host mimicking conditions (3-, 5-, and 7-day periods), H1.52 deletion resulted in enhanced metabolic activity and accelerated growth recovery compared to wild-type. These findings establish that Cryptococcus neoformans H1.52 coordinates chromatin compaction, maintains H3K9-marked heterochromatin islands and is a regulator of metabolic responses to host stress.