Functional Characterization of Myosin I Tail Regions in Candida albicans

Abstract

The molecular motor myosin I is required for hyphal growth in the pathogenic yeast Candida albicans. Specific myosin I functions were investigated by a deletion analysis of five neck and tail regions. Hyphal formation requires both the TH1 region and the IQ motifs. The TH2 region is important for optimal hyphal growth. All of the regions, except for the SH3 and acidic (A) regions that were examined individually, were required for the localization of myosin I at the hyphal tip. Similarly, all of the domains were required for the association of myosin I with pelletable actin-bound complexes. Moreover, the hyphal tip localization of cortical actin patches, identified by both rhodamine-phalloidin staining and Arp3-green fluorescent protein signals, was dependent on myosin I. Double deletion of the A and SH3 domains depolarized the distribution of the cortical actin patches without affecting the ability of the mutant to form hyphae, suggesting that myosin I has distinct functions in these processes. Among the six myosin I tail domain mutants, the ability to form hyphae was strictly correlated with endocytosis. We propose that the uptake of cell wall remodeling enzymes and excess plasma membrane is critical for hyphal formation.

Transcript Profiles of Candida albicans Cortical Actin Patch Mutants Reflect Their Cellular Defects: Contribution of the Hog1p and Mkc1p Signaling Pathways

“…In C. albicans, perturbations in the actin cytoskeleton are coupled to defects in hyphal development. For example, myosin I function is important for proper cortical actin patch distribution and for endocytosis and is critical for the formation of true hyphae (41,42). The sla2⌬ mutant also exhibits defects in both hyphal formation and the organization of cortical actin patches (5) (see Fig.…”

“…Total RNA was extracted with phenol and glass beads from wild-type and myosin I deletion strains grown to early log phase in YPD at an optical density at 600 nm of 0.8. Twenty micrograms of total RNA per sample was separated on a 7.5% formaldehyde, 1% agarose gel, blotted onto Zeta-Probe nylon membrane (Bio-Rad, Ontario, Canada), and probed with 32 P-labeled DNA specific for ACT1 (orf19.5007), orf19.5302, CRH1 (orf19.2706), AGP2 (orf19.4679), orf19.7296, SOD5 (orf19.2060), EBP1 (orf19.125), PHR1 (orf19.3829), and TOS2 (orf19.1911) as described previously (42). All Northern probes were PCR products subsequently labeled by random priming (Amersham Biosciences, NJ).…”

Hyphal Growth: a Tale of Motors, Lipids, and the Spitzenkörper

“…The first evidence for such a role of myosin-1 was found in Saccharomyces cerevisiae, where deletion of myosin-1 motors impaired endocytosis and almost abolished growth (38). In Aspergillus nidulans (73,88) and in Candida albicans (86,87), myosin-1 is essential for hyphal growth. In both fungal species, myosin-1 activity is required to mediate the endocytotic uptake of the endocytic marker dye FM4-64 into the vacuole (86,142).…”

“…Indeed, the myosin-1 tail is able to induce actin polymerization (37,65). It is therefore possible that the motor domain mediates localization of myosin-1 to sites of endocytosis, namely the hyphal tip and septa (86,143), whereas the tail supports internalization and motility of endocytic vesicles by triggering actin polymerization. Consequently, the effect of deletion of myosin-1 on hyphal growth and morphology can be explained by a role of the motor in endocytotic FIG.…”

“…In Aspergillus nidulans (73,88) and in Candida albicans (86,87), myosin-1 is essential for hyphal growth. In both fungal species, myosin-1 activity is required to mediate the endocytotic uptake of the endocytic marker dye FM4-64 into the vacuole (86,142). Surprisingly, a mutant allele in MYOA, the myosin-1 in A. nidulans, which has almost no ATPase activity, is still able to support hyphal growth (70).…”