Abstract
The human pathogenic fungus Candida albicans can switch between yeast and hyphal morphologies as a function of environmental conditions and cellular physiology. The yeast-to-hyphae morphogenetic switch is activated by well-established, kinase-based signal transduction pathways that are induced by extracellular stimuli. In order to identify possible inhibitory pathways of the yeast-to-hyphae transition, we interrogated a collection of C. albicans protein kinases and phosphatases ectopically expressed under the regulation of the TETon promoter. Proportionately more phosphatases than kinases were identified that inhibited hyphal morphogenesis, consistent with the known role of protein phosphorylation in hyphal induction. Among the kinases, we identified AKL1 as a gene that significantly suppressed hyphal morphogenesis in serum. Akl1 specifically affected hyphal elongation rather than initiation: overexpression of AKL1 repressed hyphal growth, and deletion of AKL1 resulted in acceleration of the rate of hyphal elongation. Akl1 suppressed fluid-phase endocytosis, probably via Pan1, a putative clathrin-mediated endocytosis scaffolding protein. In the absence of Akl1, the Pan1 patches were delocalized from the sub-apical region, and fluid-phase endocytosis was intensified. These results underscore the requirement of an active endocytic pathway for hyphal morphogenesis. Furthermore, these results suggest that under standard conditions, endocytosis is rate-limiting for hyphal elongation.
Highlights
Phosphorylation, the most prevalent post-translational modification of cellular proteins, can modify protein conformation or protein-protein interactions, and as a consequence can alter the enzymatic activity of proteins, their localization, or their stability
The results shown here further underscore the relation between hyphal morphogenesis and the endocytic pathway, and suggest that hyphal elongation can be regulated via modulation of endocytosis
While this collection likely contains the vast majority of the kinase and phosphatase gene complement of C. albicans, it should be stressed that ectopic expression of a gene does not necessarily cause activation
Summary
Phosphorylation, the most prevalent post-translational modification of cellular proteins, can modify protein conformation or protein-protein interactions, and as a consequence can alter the enzymatic activity of proteins, their localization, or their stability. The signal transduction pathways that mediate hyphal induction have been thoroughly explored, and found to include transmission of extracellular signals via Ras to both protein kinase A and the MAP kinase cascade (Leberer et al, 1996; Feng et al, 1999; Biswas and Morschhäuser, 2005; Maidan et al, 2005) These signaling pathways result in the activation of a number of transcription factors such as Efg (Stoldt et al, 1997) and Ume (Banerjee et al, 2008; Zeidler et al, 2009) and inactivation of the transcriptional repressor Nrg, which needs to be removed from hyphalspecific gene (HSG) promoters in order to allow induction of hyphal morphogenesis (Braun et al, 2001; Murad et al, 2001; Lu et al, 2011). Effective hyphal development requires alterations in many cellular functions; for example, growth of hyphae requires an active endocytic pathway, possibly in order to counterbalance the loss of membrane and proteins by exocytosis at the hyphal tip (Shaw et al, 2011)
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