Cdc42 Is Required for Proper Growth and Development in the Fungal Pathogen Colletotrichum trifolii

Abstract

Cdc42 is a highly conserved small GTP-binding protein that is involved in regulating morphogenesis in eukaryotes. In this study, we isolated and characterized a highly conserved Cdc42 gene from Colletotrichum trifolii (CtCdc42), a fungal pathogen of alfalfa. CtCdc42 is, at least in part, functionally equivalent to Saccharomyces cerevisiae Cdc42p, since it restores the temperature-sensitive phenotype of a yeast Cdc42p mutant. Inhibition of CtCdc42 by expression of an antisense CtCdc42 or a dominant negative form of CtCdc42 (DN Cdc42) resulted in appressorium differentiation under noninductive conditions, suggesting that CtCdc42 negatively regulates pathogenic development in this fungus. We also examined the possible linkage between CtCdc42 and Ras signaling. Expression of a dominant active Cdc42 (DA Cdc42) in C. trifolii leads to aberrant hyphal growth under nutrient-limiting conditions. This phenotype was similar to that of our previously reported dominant active Ras (DA Ras) mutant. Also consistent with our observations of the DA Ras mutant, high levels of reactive oxygen species (ROS) were observed in the DA Cdc42 mutant, and proline restored the wild-type phenotype. Moreover, overexpression of DN Cdc42 resulted in a significant decrease in spore germination, virtually no hyphal branching, and earlier sporulation, again similar to what we observed in a dominant negative Ras (DN Ras) mutant strain. Interestingly, coexpression of DA Cdc42 with DN Ras resulted in germination rates close to wild-type levels, while coexpression of DN Cdc42 with the DA Ras mutant restored the wild-type phenotype. These data suggest that CtCdc42 is positioned as a downstream effector of CtRas to regulate spore germination and pathogenic development.

Control of T Lymphocyte Signaling by Ly108, a Signaling Lymphocytic Activation Molecule Family Receptor Implicated in Autoimmunity

“…There was also a reduction of the ability of SAP R78A to interact with Ly108 (third panel, lanes 3 and 4), in comparison to wild-type SAP (lanes 1 and 2). It is unlikely that this reduction was due to a lower affinity of Ly108 toward SAP R78A, since mutation of Arg-78 does not affect the phosphotyrosine-binding fold of the SAP SH2 domain (5,7,8). This reduced association was probably caused by the attenuated tyrosine phosphorylation of Ly108.…”

“…These adaptors are composed primarily of a Src homology 2 (SH2) domain and link SRRs to intracellular signals. In the case of SAP, a second binding surface in the SH2 domain enables SAP to couple SRRs to the Src-related protein-tyrosine kinase FynT, thereby triggering protein tyrosine phosphorylation signals (5)(6)(7)(8). EAT-2 and ERT possess one or two carboxyl-terminal tyrosines that undergo phosphorylation and link SRRs to alternative signals (9).…”

Structural Basis for the Interaction between the Growth Factor-binding Protein GRB10 and the E3 Ubiquitin Ligase NEDD4

“…Some instances have been reported in which SH2 domains use binding sites different from and not overlapping with the classical Tyr(P)-binding pocket to colocalize a kinase and substrate (45,46). As shown in Fig.…”

“…There are a few other cases in which an SH2 domain binds proteins using binding sites other than the classical Tyr(P)-binding pocket (45,46). For example, the Itk kinase domain docking site on the PLC␥1 SH2C domain surface, which includes residues Glu 709 , Arg 748 , Met 750 , Lys 751 , and Arg 753 , is far from and does not overlap with the classical Tyr(P)-binding pocket (47).…”

Parallels in Fungal Pathogenesis on Plant and Animal Hosts

“…Specific nutritional mechanisms are activated via complex regulatory networks that include genes involved in germination, such as those encoding cAMP-dependent kinases, G ␣ proteins, and MAP kinases, as well as Ras proteins, which are a family of small monomeric GTPases involved in a variety of cellular signaling pathways (26,29,59). In the alfalfa pathogen Colletotrichum trifolii, Ras regulates spore germination and pathogenic development via genes including Cdc42, which is required for growth and development (20). Cdc42 is involved in germ tube formation and invasive hyphal growth in C. albicans, and ectopic expression leads to loss of virulence in mice (6).…”