We have characterized the CDC14 gene, which encodes a dual-specificity protein phosphatase in Candida albicans, and demonstrated that its deletion results in defects in cell separation, mitotic exit and morphogenesis. The C. albicans cdc14Δ mutants formed large aggregates of cells that resembled those found in ace2-null strains. In cdc14Δ cells, expression of Ace2p target genes was reduced and Ace2p did not accumulate specifically in daughter nuclei. Taken together, these results imply that Cdc14p is required for the activation and daughter-specific nuclear accumulation of Ace2p. Consistent with a role in cell separation, Cdc14p was targeted to the septum region during the M-G1 transition in yeast-form cells. Interestingly, hypha-inducing signals abolished the translocation of Cdc14p to the division plate, and this regulation depended on the cyclin Hgc1p, since hgc1Δ mutants were able to accumulate Cdc14p in the septum region of the germ tubes. In addition to its role in cytokinesis, Cdc14p regulated mitotic exit, since synchronous cultures of cdc14Δ cells exhibited a severe delay in the destruction of the mitotic cyclin Clb2p. Finally, deletion of CDC14 resulted in decreased invasion of solid agar medium and impaired true hyphal growth.

Filamentous hyphae of the human pathogen, Candida albicans, invade mucosal layers and medical silicones. In vitro, hyphal tips reorient thigmotropically on contact with small obstacles. It is not known how surface topography is sensed but hyphae lacking the cortical marker, Rsr1/Bud1, are unresponsive. We show that, on surfaces, the morphology of hyphal tips and the position of internal polarity protein complexes are asymmetrically skewed towards the substratum and biased towards the softer of two surfaces. In nano-fabricated chambers, the Spitzenkörper (Spk) responded to touch by translocating across the apex towards the point of contact, where its stable maintenance correlated with contour-following growth. In the rsr1Δ mutant, the position of the Spk meandered and these responses were attenuated. Perpendicular collision caused lateral Spk oscillation within the tip until after establishment of a new growth axis, suggesting Spk position does not predict the direction of growth in C. albicans. Acute tip reorientation occurred only in cells where forward growth was countered by hyphal friction sufficient to generate a tip force of ∼ 8.7 μN (1.2 MPa), more than that required to penetrate host cell membranes. These findings suggest mechanisms through which the organization of hyphal tip growth in C. albicans facilitates the probing, penetration and invasion of host tissue.

In the hyphal tip of Candida albicans we have made detailed quantitative measurements of (i) exocyst components, (ii) Rho1, the regulatory subunit of (1,3)-β-glucan synthase, (iii) Rom2, the specialized guanine-nucleotide exchange factor (GEF) of Rho1, and (iv) actin cortical patches, the sites of endocytosis. We use the resulting data to construct and test a quantitative 3-dimensional model of fungal hyphal growth based on the proposition that vesicles fuse with the hyphal tip at a rate determined by the local density of exocyst components. Enzymes such as (1,3)-β-glucan synthase thus embedded in the plasma membrane continue to synthesize the cell wall until they are removed by endocytosis. The model successfully predicts the shape and dimensions of the hyphae, provided that endocytosis acts to remove cell wall-synthesizing enzymes at the subapical bands of actin patches. Moreover, a key prediction of the model is that the distribution of the synthase is substantially broader than the area occupied by the exocyst. This prediction is borne out by our quantitative measurements. Thus, although the model highlights detailed issues that require further investigation, in general terms the pattern of tip growth of fungal hyphae can be satisfactorily explained by a simple but quantitative model rooted within the known molecular processes of polarized growth. Moreover, the methodology can be readily adapted to model other forms of polarized growth, such as that which occurs in plant pollen tubes.

Candida albicans hyphae continue to grow throughout mitosis, and phosphorylation of Exo84 by Cdk is necessary for efficient hyphal growth. In contrast, phosphorylation of Exo84 by Cdk halts cell growth in Saccharomyces cerevisiae. The location of Cdk1 target sites in Exo84 explains how phosphoregulation mediates these different patterns of growth.

Candida albicans hyphae grow in a highly polarized fashion from their tips. This polarized growth requires the continuous delivery of secretory vesicles to the tip region. Vesicle delivery depends on Sec2p, the Guanine Exchange Factor (GEF) for the Rab GTPase Sec4p. GTP bound Sec4p is required for the transit of secretory vesicles from the trans-Golgi to sites of polarized growth. We previously showed that phosphorylation of Sec2p at residue S584 was necessary for Sec2p to support hyphal, but not yeast growth. Here we show that on secretory vesicles SEC2 mRNA is physically associated with Sec2p. Moreover, we show that the phosphorylation of S584 allows SEC2 mRNA to dissociate from Sec2p and we speculate that this is necessary for Sec2p function and/or translation. During hyphal extension, the growing tip may be separated from the nucleus by up to 15 μm. Transport of SEC2 mRNA on secretory vesicles to the tip localizes SEC2 translation to tip allowing a sufficient accumulation of this key protein at the site of polarized growth.