A p21-Activated Kinase Is Required for Conidial Germination in Penicillium marneffei

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Abstract
Asexual spores (conidia) are the infectious propagules of many pathogenic fungi, and the capacity to sense the host environment and trigger conidial germination is a key pathogenicity determinant. Germination of conidia requires the de novo establishment of a polarised growth axis and consequent germ tube extension. The molecular mechanisms that control polarisation during germination are poorly understood. In the dimorphic human pathogenic fungus Penicillium marneffei, conidia germinate to produce one of two cell types that have very different fates in response to an environmental cue. At 25 °C, conidia germinate to produce the saprophytic cell type, septate, multinucleate hyphae that have the capacity to undergo asexual development. At 37 °C, conidia germinate to produce the pathogenic cell type, arthroconidiating hyphae that liberate uninucleate yeast cells. This study shows that the p21-activated kinase pakA is an essential component of the polarity establishment machinery during conidial germination and polarised growth of yeast cells at 37 °C but is not required for germination or polarised growth at 25 °C. Analysis shows that the heterotrimeric G protein α subunit GasC and the CDC42 orthologue CflA lie upstream of PakA for germination at both temperatures, while the Ras orthologue RasA only functions at 25 °C. These findings suggest that although some proteins that regulate the establishment of polarised growth in budding yeast are conserved in filamentous fungi, the circuitry and downstream effectors are differentially regulated to give rise to distinct cell types. Many fungal infections are initiated by the entry of dormant fungal spores into their host. Once inside the host these dormant spores must reactivate (germinate) for the infection to proceed. Productive infections necessitate that the fungus grow and divide within the host, which makes understanding the mechanisms that control germination crucial to developing preventative or prophylactic treatments for fungal infections. The molecular mechanisms that control spore germination are poorly understood and studies of the opportunistic fungal pathogen Penicillium marneffei have shown that a group of highly conserved signalling and cell polarity factors, known as small GTPases, play important roles in germination and other aspects of morphogenesis. In this study we have shown that a downstream target of these small GTPases, a p21-activated kinase plays a crucial role in germination at the host temperature of 37 °C but not at 25 °C. This is the first component of germination, which shows temperature-dependent effects and provides insights into the different mechanisms used by fungal pathogens to infect their host or to grow saprophytically in non-host environments.