TheCryptococcus neoformansTitan cell is an inducible and regulated morphotype underlying pathogenesis
Authors: Dambuza Ivy M. , Drake Thomas , Chapuis Ambre , Taylor-Smith Leanne , LeGrave Nathalie , Rasmussen Tim , Fisher Matthew C. , Bicanic Tihana , Harrison Thomas S. , Jaspars Marcel , May Robin C. , Brown Gordon D. , Yuecel Raif , MacCallum Donna M. , Ballou Elizabeth R. 
Published: September 2017

Fungi undergo changes in cell shape in response to environmental stimuli that drive pathogenesis and niche adaptation, such as the yeast-to-hyphal transition of dimorphic fungi in response to changing temperature. The basidiomyceteCryptococcus neoformansundergoes an unusual morphogenetic transition in the host lung from haploid yeast to large, highly polyploid cells termed Titan cells. Titan cells influence fungal interaction with host cells, including through increased drug resistance, altered cell size, and altered Pathogen Associated Molecular Pattern exposure. Despite the important role these cells play in pathogenesis, understanding the environmental stimuli that drive the morphological transition, and the molecular mechanisms underlying their unique biology, has been hampered by the lack of a reproduciblein vitroinduction system. Here we demonstrate reproduciblein vitroTitan cell induction in response to environmental stimuli consistent with the host lung.In vitroTitan cells exhibit all the properties ofin vivogenerated Titan cells, the current gold standard, including altered capsule, cell wall, size, high mother cell ploidy, and aneuploid progeny. We identify bacterial peptidoglycan as a serum compound associated with shift in cell size and ploidy, and demonstrate the capacity of bronchial lavage fluid andE. colico-culture to induce Titanisation. Additionally, we demonstrate the capacity of our assay to identify established and previously undescribed regulators of Titanisationin vitroand investigate the Titanisation capacity of clinical isolates and their impact on disease outcome. Together, these findings provide new insight into the environmental stimuli and molecular mechanisms underlying the yeast-to-titan transition and establish an essentialin vitromodel for the future characterization of this important morphotype.

Author Summary

Changes in cell shape underlie fungal pathogenesis by allowing immune evasion and dissemination.AspergillusandCandida albicanshyphae drive tissue penetration.Histoplasma capsulatumandC. albicansyeast growth allows evasion and dissemination. As major virulence determinates, morphogenic transitions are extensively studied in animal models andin vitro. The pathogenic fungusCryptococcus neoformansis a budding yeast that, in the host lung, switches to an unusual morphotype termed the Titan cell. Titans are large, polyploid, have altered cell wall and capsule, and produce haploid daughters. Their size prevents engulfment by phagocytes, yet they are linked to dissemination and altered immune response. Despite their important influence on disease, replicating the yeast-to-Titan switchin vitrohas proved challenging. Here we show that Titans are induced by host-relevant stimuli, including serum and bronchio-alveolar lavage fluid. We identify bacterial peptidoglycan as a relevant inducing compound and predict anin vivoTitan defect for a clinical isolate. Genes regulatingin vivoTitanisation also influencein vitroformation. Titanisation is a conserved morphogenic switch across theC. neoformansspecies complex. Together, we show that Titan cells are a regulated morphotype analogous to the yeast-to-hyphal transition and establish new ways to study Titans outside the host lung.