No abstract

In this paper we compared the protein synthesis patterns of Paracoccidioides brasiliensis isolates. The protein profiles were compared for both yeast and mycelial forms and similarity analysis among them was performed by calculating similarity matrices and grouping the isolates in dendrograms. The examined isolates exhibited highly variable cellular morphology at 36 °C, when typical yeast cells were expected. On the other hand, at 26 °C all the isolates showed mycelial morphology. The analysis of protein synthesis profiles made it possible to cluster the P. brasiliensis isolates into groups that correlated with the morphological data. Interestingly, growth at 36 °C strongly decreased the heterogeneity of protein synthesis patterns seen in mycelial isolates. It was possible to cluster the isolates grown at 36 °C in three groups based on their two-dimensional protein synthesis analysis. The similarity index observed among the mycelial isolates was lower than that obtained with yeast cells, suggesting a more homogeneous gene expression pattern in the host-adapted form than in the saprobic phase.

A critical step in the lifecycle of many fungal pathogens is the ability to switch between filamentous and yeast growth, a process known as dimorphism. cAMP-dependent protein kinase (PKA) controls morphological changes and the pathogenicity of several animal and plant pathogenic fungi. In this work, we report the analysis of PKA activity during the mycelium to yeast transition in the pathogenic fungus Paracoccidioides lutzii. This fungus, as well as the closely related species Paracoccidioides brasiliensis, causes paracoccidioidomycosis, a systemic mycosis that affects thousands of people in Latin America. Infection occurs when hypha fragments or spores released from mycelium are inhaled by the host, an event that triggers the morphological switch. We show here that PKA activity is regulated in the fungus phase, increasing during the mycelium to yeast transition. Also, morphological transition from mycelium to yeast is blocked by the compound H89, a specific PKA inhibitor. Nevertheless, the fungus recovers its ability to change morphology when H89 is removed from the culture media. This recovery is accompanied by a significant increase in PKA activity. Our results strongly indicate that PKA directly affects phase transition in P. lutzii.

Expression of the 43 kDa glycoprotein (gp43) was analysed in several Paracoccidioides brasiliensis isolates. Using one-and two-dimensional analysis of crude cellular extracts, it was shown that protein expression in yeast and mycelium was dependent on the isolate analysed. In two strains, in both yeast and mycelium cells, gp43 was present, whereas expression was restricted to the yeast phase of two other strains. The clinical implications of this phase-specific gp43 expression are uncertain.