Growth control and polarization

Medical Mycology Supplement 1 2005, 43, S23
/S25 Growth control and polarization M. MOMANY Department of Plant Biology, University of Georgia, Athens, GA, USA Keywords branching, polarity Introduction While the conidia of Aspergillus fumigatus are ubiquitous in the environment and frequently inhaled, a competent immune system in most individuals generally clears them before they cause disease. Studies have not addressed early growth of A. fumigatus inside the immunocompromised host, but in vitro investigations have established the landmarks of early development [1]. After a conidium breaks dormancy, the reactivated A. fumigatus cell undergoes a brief period of isotropic expansion before a germ tube emerges. As is true for all filamentous fungi, later growth is highly polar, occurring exclusively at the tips of hyphae and branches. In invasive aspergillosis this highly polar tip growth allows A. fumigatus to invade blood vessels and tissue where it continues to grow, eventually causing hemorrhage and necrosis. Polarity in Saccharomyces cerevisiae and aspergilli Much has been learned about polar growth in the budding yeast Saccharomyces cerevisiae. In this fungus, cortical markers specify the site of bud emergence and Correspondence: M. Momany, Department of Plant Biology, University of Georgia, Athens, GA 30605, USA. Tel: /1 706 542 2014; Fax: /1 706 542 1805; E-mail: – 2005 ISHAM the Cdc42 Rho GTPase relays this information to the morphogenetic machinery, including actin and components of the secretory system. Genome comparisons reveal differences and similarities in polarity between S. cerevisiae and A. fumigatus, A. nidulans and A. oryzae [2]. The suite of cortical markers that specify the site of bud emergence in yeast (including Bud3p, Bud4p, Bud8p, Bud9p, Axl12p and Rax2p) are either absent or very poorly conserved in the aspergilli. However the signal relay consisting of the Cdc42p Rho GTPase and its associated GEF (Cdc24p), GAPs (Rga1p, Bem2p and Bem3p) and downstream effectors (Ste20p and Cla4p) are highly conserved. Of particular interest is the fact that the Cdc42p orthologues are essential in Candida albicans and Ashbya gossyippi , both relatives of S. cerevisiae. However, they are nonessential in the more distantly related filamentous fungi A. nidulans, Penicillium marneffei and Magnaporthe grisea . These filamentous fungi also contain the Rac GTPase, which is absent in yeasts. Deletion of Rac in A. niger and in P. marneffei results in increased branching [2]. Though there is no experimental evidence yet, it seems possible that Rac might be at least partially redundant with Cdc42 and that this redundancy might explain why deletion of Cdc42 is not lethal in these fungi. The positional signal that is relayed through the Rho GTPase modules ultimately affects the morphogenetic machinery that remodels the cell surface. Many DOI: 10.1080/13693780400024263 Filamentous fungi and yeasts both undergo polar growth. In Saccharomyces cerevisiae, where the mechanisms for polar growth are well-understood, polarity requires three steps: establishment of cortical markers specifying the site of bud emergence; relaying the bud site information via the Cdc42 Rho GTPase module; and recruitment of the morphogenetic machinery needed to remodel the cell surface to the specified site. Comparison of the genomes of Aspergillus fumigatus, A. nidulans and A. oryzae with that of S. cerevisiae show that the cortical markers are absent or poorly conserved, while the RhoGTPase signaling module and the morphogenetic machinery are highly conserved in the aspergilli. Genetic approaches to polarity using A. nidulans polarity mutants with defects in germ tube emergence (swo mutants) or branching (ahb mutants) will also be discussed. S24 Momany components of the morphogenetic machinery are conserved between S. cerevisiae and the aspergilli. Not surprisingly, cytoskeletal elements such as actin and tubulins are highly conserved. Components of the polarisome, the complex responsible for organizing actin assembly in polar growth are also conserved, as are components of the exocyst, a protein complex important in secretion. It is increasingly clear that germ tube emergence in filamentous fungi shares certain key features with bud emergence in S. cerevisiae, but important differences exist [3,4]. In filamentous fungi polar growth is persistent, while in yeast it is sporadic. In filamentous fungi polarity signals must be coordinated over an extended multicellular hypha, while in yeast those signals need only reach a single small cell. In filamentous fungi multiple axes of polarity are established simultaneously through branching, while in yeast a single axis of polarity is established through budding. Furthermore, in filamentous fungi the cell cycles of multiple nuclei must be coordinated with each other and with germ tube and branch emergence. Some of these key differences suggest that the basic polarity machinery is regulated differently in filamentous fungi. Genetic approaches to polarity: swo mutants Classical genetics is a powerful tool for dissecting cellular processes. In A. nidulans we have taken a genetic approach to polarity, generating mutants with defects in germ tube emergence and branching. The temperature-sensitive swo (swollen) mutants do not properly extend a germ tube at the restrictive temperature of 428C [5]. Based on temperature-shift experiments, we have classified the eight swo mutants into groups with defects in polarity establishment (marking the spot for germ tube emergence) and those with defects in polarity maintenance (transducing signals or recruiting the morphogenetic machinery so that the germ tube extends). Here we will focus on two of these mutants, swoF and swoA . Previous work has shown that swoF is required for both polarity establishment and maintenance while swoA is required for polarity maintenance. At restrictive temperature swoF cells swell slightly but do not extend a germ tube, even though they continue nuclear division. Sequencing of the gene that complemented the swoF mutant showed that it encodes an N-myristoyl transferase (NMT) [6]. NMTs co-translationally add a fatty acid group to the N-terminus of target proteins. This myristoylation is Fig. 1 Aspergillus nidulans mutant phenotypes at restrictive temperature. (A) wild-type; (B) swoA; (C) swoF; (D) ahbA ; (E) ahbB. Panel A from Euk Cell 1:242; B from Fungal Genet and Biol 37:266; D, E from Fungal Genet and Biol 41:1001. – 2005 ISHAM, Medical Mycology, 43, S23
/S25 Filamentous fungi are not just tall yeasts thought to increase the affinity of the target protein for the plasma membrane. Our working hypothesis is that modification by swoF is needed for either proper localization or function of a protein or proteins required for appropriate polar growth. In S. cerevisiae, the targets of NMTs include proteins involved in vesicle assembly, which is consistent with a role in polari (...truncated)