The regulation of filamentous growth in yeast

Abstract

Filamentous growth is a nutrient-regulated growth response that occurs in many fungal species. In pathogens, filamentous growth is critical for host-cell attachment, invasion into tissues, and virulence. The budding yeast Saccharomyces cerevisiae undergoes filamentous growth, which provides a genetically tractable system to study the molecular basis of the response. Filamentous growth is regulated by evolutionarily conserved signaling pathways. One of these pathways is a mitogen activated protein kinase (MAPK) pathway. A remarkable feature of the filamentous growth MAPK pathway is that it is composed of factors that also function in other pathways. An intriguing challenge therefore has been to understand how pathways that share components establish and maintain their identity. Other canonical signaling pathways-rat sarcoma/protein kinase A (RAS/PKA), sucrose nonfermentable (SNF), and target of rapamycin (TOR)-also regulate filamentous growth, which raises the question of how signals from multiple pathways become integrated into a coordinated response. Together, these pathways regulate cell differentiation to the filamentous type, which is characterized by changes in cell adhesion, cell polarity, and cell shape. How these changes are accomplished is also discussed. High-throughput genomics approaches have recently uncovered new connections to filamentous growth regulation. These connections suggest that filamentous growth is a more complex and globally regulated behavior than is currently appreciated, which may help to pave the way for future investigations into this eukaryotic cell differentiation behavior.

Figure 1

The life cycle of the budding yeast Saccharomyces cerevisiae. The diagram shows yeast-form cells, which can be induced to undergo different growth responses depending on ploidy and growth condition. Haploid and diploid cells interconvert between the two types by mating and sporulation, respectively. Both haploid and diploid cells can undergo filamentous growth, form biofilms, or enter stationary phase (quiescence) in response to nutrient (glucose or nitrogen) limitation. Diploid cells also sporulate in response to the limitation of carbon and nitrogen sources. Secreted alcohols act as autoinducers to stimulate filamentous growth.

Figure 2

The filamentous growth response. Several biological assays permit the evaluation of the filamentous growth response in yeast, using the Σ1278b strain background. (A) Haploid wild-type (left) and flo11 mutant (right) colonies grown on YEPD + 4% agar medium for 7 days show the Flo11-dependent colony ruffling. Bar, 0.5 cm. (B) The plate-washing assay (Roberts and Fink 1994). Haploid wild-type (left) and MAPK pathway mutant (right) cells were spotted onto YEPD medium (2% agar). After 3 days the plate was photographed (top), washed in a stream of water, and photographed again (bottom) to reveal invaded cells. Bar, 1 cm. (C) The single cell invasive growth assay (Cullen and Sprague 2000). Cells as in B were spread onto SC medium lacking glucose as a carbon source for 1 day. Bar, 10 µM. (D) Diploid pseudohyphal growth assay (Gimeno et al. 1992). Homozygous diploid versions of the strains described in B were examined on SLAHD (low nitrogen) medium. Bar, 50 µM.

Figure 3

The RAS/PKA pathway. The G-protein coupled receptor (GPCR) Gpr1 and its associated heterotrimeric G protein regulate the Ras2 GTPase activating proteins (GAPs), Ira1 and Ira2. Ras2 regulates adenylate cyclase, which produces cAMP. cAMP binds to Bcy1, inactivating the protein, and releasing Tpk1, Tpk2, and Tpk3 to activate Flo8 and other targets that contribute to nutrient-regulated filamentous growth. Filled hexagons represent sucrose and other sugars.

Figure 4

Three MAPK pathways in yeast share common components and also contain pathway-specific factors. (A) Three MAPK pathways are shown. Colored proteins represent pathway-specific factors; protein shown in black function in multiple pathways. Scaffold-mediated interactions are shown by colored, dashed lines. Not all protein interactions are shown. Hot1 is one of a number of transcription factors for the HOG pathway. The red question mark indicates that how nutritional signals feed into filamentous growth pathway regulation is not well understood. (B) Examples of MAPK morphogenesis in yeast. The pheromone response (Mating) pathway induces distinctive polarized structures called shmoos to promote cell fusion and diploid formation. The Filamentous Growth pathway induces filamentous growth, branched chains of elongated and connected cells. Activation of the HOG pathway does not induce polarized growth. Bar, 5 μm.

Figure 5

Model of the filamentous growth MAPK pathway. Upon nutrient limitation, expression of the gene encoding the aspartyl protease Yps1 is induced. Yps1 processes the signaling mucin Msb2 in its extracellular domain, which is required for MAPK activation. Processed Msb2 (Msb2*) associates with and functions through Sho1 to activate cytosolic signaling modules. Msb2 associates with the Rho GTPase Cdc42, and Sho1 functions in a complex with the GEF Cdc24. A straightforward possibility is that the association of activated Msb2 with Sho1 brings the GEF into close proximity with its GTPase. Activated Cdc42 binds effector proteins including the PAK Ste20, which when activated, phosphorylates the MAPKKK Ste11, thereby activating the MAPK cascade.

Figure 6

Mechanisms of signal integration among regulatory proteins and pathways that control filamentous growth. (A) Multiple signaling pathways converge on the FLO11 promoter to modulate gene expression (Rupp et al. 1999). Both Snf1 and Rim101 are thought to function through the transcriptional repressors Nrg1 and Nrg2 (Kuchin et al. 2002; Lamb and Mitchell 2003). (B) Multiple signaling pathways regulate the activity of the filamentation MAPK pathway, adapted from (Chavel et al. 2010). Rtg refers to the retrograde mitochondrial signaling pathway (Liu and Butow 2006). Other pathways also converge on FLO11 that are not shown here (Bruckner and Mosch 2011).

Figure 7

Patterns of budding and bud-site–selection proteins required during yeast-form and filamentous growth. In nutrient-rich conditions, haploid cells bud axially, using cortical landmarks Bud3, Bud4, Bud10, and Axl1 that are localized to the mother-bud neck. Diploid cells grow at both poles in a bipolar pattern using Bud8, Rax1, and Rax2 at the distal pole and Bud9 at the proximal pole. Under nutrient-limiting conditions, both cell types switch to a distal-unipolar pattern and bud more or less exclusively at the distal pole.