Browsing by Subject "Septin"
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Item Open Access Regulation of Morphogenetic Events in Saccharomyces cerevisiae(2018) Lai, Hung-HsuehTip growth in fungi involves highly polarized secretion and modification of the cell wall at the growing tip. The genetic requirements for initiating polarized growth are perhaps best understood for the model budding yeast, Saccharomyces cerevisiae. Once the cell is committed to enter the cell cycle by activation of G1 cyclin/cyclin-dependent kinase (CDK) complexes, the polarity regulator Cdc42 becomes concentrated at the presumptive bud site, actin cables are oriented towards that site, and septin filaments assemble into a ring around the polarity site. Several minutes later, the bud emerges. Here, we investigated the mechanisms that regulate the timing of these events at the single cell level and the role of polarisome during pheromone-induced polarized growth. We employed genetics and live cell microscopy to characterize cellular events. Septin recruitment was delayed relative to polarity establishment, and our findings suggest that a CDK-dependent septin “priming” facilitates septin recruitment by Cdc42. Bud emergence was delayed relative to the initiation of polarized secretion, and our findings suggest that the delay reflects the time needed to weaken the cell wall sufficiently to bud. Rho1 activation by Rom2 occurred at around the time of bud emergence, perhaps in response to local cell wall weakening. This report reveals regulatory mechanisms underlying the morphogenetic events in the budding yeast.
Item Open Access Septins’ Role in Morphogenesis, Development, and Pathogenesis of the Human Fungal Pathogen Aspergillus fumigatus(2017) VargasMuniz, Jose MAspergillus fumigatus, the main etiology of invasive aspergillosis, is a leading cause of fungal mortality in immunocompromised patients. Although the incidence of invasive aspergillosis has increased in the last two decades due to a rise in the immunocompromised patient population, there is a lack of effective treatments and basic understanding of growth and disease. Septins are conserved GTPases that are involved in a myriad of cellular processes, ranging from cytokinesis to cell morphology. Here we describe the role of septins in A. fumigatus growth, development, and pathogenesis. Through gene deletion, we revealed that all 5 septin genes are dispensable for growth under basal conditions. Nonetheless, AspA, AspB, AspC, and AspE are required for regular septation and the core septins are required for conidia production. The ΔaspB strain was hypervirulent in the Galleria mellonella model of infection, while virulence was similar to the wild-type strain in our murine model of invasive aspergillosis. Deletion of aspB also lead to an increase in susceptibility to anti-cell wall agents. AspB, which under basal conditions interacts with 334 proteins, alters its protein interaction after exposure to a clinically relevant concentration of the antifungal caspofungin. A total of 69 of the basal interactants do not interact with AspB, and 54 new interactants were identified following caspofungin exposure. Of the interactants studied, only PpoA was implicated in the response to anti-cell wall agents.
Due to the pleiotropic role of AspB, we characterized how posttranslational modifications regulate AspB function. Gene deletion analyses revealed that Cla4 and ParA are indispensable for hyphal extension. Gin4, Cla4, and ParA contributes to both septation and conidiation of A. fumigatus. Deletion of cla4 and parA lead to hyperseptation, while deletion of gin4 leads to larger apical compartments that were similar in length to the ΔaspB strain. Cla4, Gin4, and ParA contribute to proper localization of AspB. Phosphoproteomic analyses show that AspB is phosphorylated at 7 residues: 5 in the GTPase domain, and 2 at C-terminus. Deletion of gin4 and cla4 did not alter the phosphorylated residues in AspB. Deletion of parA results in 2 new phosphorylation sites identified, one in the N-terminal polybasic region (T68) and one in the coil-coiled domain (S447). Mutation of T68 to glutamic acid leads to a slight increase in interseptal distances. These results demonstrate the key role of septins in response to anti-cell wall agents, as well as how phosphorylation regulates septin function in A. fumigatus.