How host-like signals drive gene expression and gene expression drives capsule expansion in Cryptococcus neoformans

Cryptococcus neoformans is an opportunistic fungal pathogen with a polysaccharide capsule that becomes greatly enlarged in the mammalian host and during in vitro growth in response to host-like conditions. To understand how individual host-like signals affect capsule size and gene expression, we grew cells with and without all combinations of 5 signals suspected of affecting capsule size and systematically measured cell and capsule sizes of 47,458 cells. We also collected samples for RNA-Seq at 30, 90, 180, and 1440 minutes and carried out RNA-Seq in quadruplicate, yielding 881 RNA-Seq samples. This massive, uniformly collected dataset will be a significant resource for the research community. Analysis revealed that capsule induction requires both tissue culture medium and either CO(2) or exogenous cyclic AMP, a second messenger. Rich medium (YPD) blocks capsule growth completely, DMEM permits it, and RPMI yields the largest capsules. Medium has the biggest impact on overall gene expression, followed by CO(2), mammalian body temperature (37° compared to 30°), and then cAMP. Surprisingly, adding CO(2) or cAMP pushes overall gene expression in the opposite direction from tissue culture media, even though both tissue culture medium and CO(2) or cAMP are required for capsule development. By modeling the relationship between gene expression and capsule size, we identified novel genes whose deletion affects capsule size.