Candida albicans Kinesin Kar3 Depends on a Cik1-Like Regulatory Partner Protein for Its Roles in Mating, Cell Morphogenesis, and Bipolar Spindle Formation

Abstract

b Candida albicans is a major fungal pathogen whose virulence is associated with its ability to transition from a budding yeast form to invasive hyphal filaments. The kinesin-14 family member CaKar3 is required for transition between these morphological states, as well as for mitotic progression and karyogamy. While kinesin-14 proteins are ubiquitous, CaKar3 homologs in hemiascomycete fungi are unique because they form heterodimers with noncatalytic kinesin-like proteins. Thus, CaKar3-based motors may represent a novel antifungal drug target. We have identified and examined the roles of a kinesin-like regulator of CaKar3. We show that orf19.306 (dubbed CaCIK1) encodes a protein that forms a heterodimer with CaKar3, localizes CaKar3 to spindle pole bodies, and can bind microtubules and influence CaKar3 mechanochemistry despite lacking an ATPase activity of its own. Similar to CaKar3 depletion, loss of CaCik1 results in cell cycle arrest, filamentation defects, and an inability to undergo karyogamy. Furthermore, an examination of the spindle structure in cells lacking either of these proteins shows that a large proportion have a monopolar spindle or two dissociated half-spindles, a phenotype unique to the C. albicans kinesin-14 homolog. These findings provide new insights into mitotic spindle structure and kinesin motor function in C. albicans and identify a potentially vulnerable target for antifungal drug development. Candida albicans is a common commensal fungus that typically causes no harm to the host (1). However, in immunocompromised individuals, neonates, and patients under intensive care, it can cause serious skin and mucosal infections as well as life-threatening systemic infections (1, 2). It is also able to form tenacious biofilms on implanted medical devices (3). A major factor facilitating C. albicans virulence is its ability to readily switch between growth as a yeast, pseudohyphae, and hyphae (1, 4). In the yeast mode, daughter cells bud off and dissociate from the mother cell, while pseudohyphal cells remain connected at constricted septation sites (4). Hyphae are the invasive form and are important for virulence during systemic infections as well as for biofilm formation (1,5,6). In all of these morphological states, microtubuleassociated motor proteins play major roles in microtubule cytoskeleton remodeling, nuclear movements, chromosome segregation, and cargo transport (7-9). This provides a rationale for their use as novel targets for antifungal drugs (10).C. albicans Kar3 (CaKar3) is a kinesin-14 family member that has been shown to function in mitotic division and is critical for hypha formation and nuclear fusion during mating (9, 11). Its homolog in Saccharomyces cerevisiae has similar functions in mitosis and mating that are enabled by interactions with two kinesinlike proteins: S. cerevisiae Cik1 (ScCik1) and ScVik1 (12)(13)(14)(15)(16)(17)(18)(19)(20). ScKar3 forms complexes with each of these proteins via a central coiled-coil domain to form parallel heterodimers that are structurally ...

Aurora kinase Ipl1 facilitates bilobed distribution of clustered kinetochores to ensure error-free chromosome segregation in Candida albicans

“…The controlled disassembly and reassembly of MT fibers regulate the architecture or the shape of a cell. It has been shown previously MT motor mutants of kip1 , kar3, or cik1 result in altered cellular morphologies in C. albicans (Frazer et al , 2015, Chua et al , 2007, Sherwood & Bennett, 2008). To test whether the aberrant morphologies in the Ipl1-depleted cells are due to altered MT dynamics, we first depleted Ipl1 in two independent mutant strains, ipl1::FRT/PCK1 p -IPL1 (CNV6 and CNV7) for 4 h and spotted them on the plates containing thiabendazole (40 µg/ml), a MT-depolymerizing agent.…”

“…The virulence of the organism has been shown to be associated with the morphological transition between yeast and hyphal cells. In these morphological forms, the MT-associated motor proteins have been shown to regulate nuclear movements, chromosome segregation, and cytoskeleton remodeling (Finley et al , 2008, Martin et al , 2004, Sherwood & Bennett, 2008, Frazer et al , 2015). Yeast and filamentous hypha, both exhibit variations in the length of the mitotic spindle, the former having shorter mitotic spindles of maximum of 8 µm and the latter with a longer spindle MTs of maximum of 20 µm, indicating that the cell morphology influences the length of MTs in the same organism (Barton & Gull, 1988, McCoy et al , 2015).…”

“…Yeast and filamentous hypha, both exhibit variations in the length of the mitotic spindle, the former having shorter mitotic spindles of maximum of 8 µm and the latter with a longer spindle MTs of maximum of 20 µm, indicating that the cell morphology influences the length of MTs in the same organism (Barton & Gull, 1988, McCoy et al , 2015). C. albicans genome encodes only one form of the kinesin-5 (Kip1) and kinesin-14 (Kar3Cik1) family motors in contrast to the two kinesin-5 (Kip1 and Cin8) and kinesin-14 (Kar3Cik1 and Kar3Vik1) family motors in S. cerevisiae (Chua et al , 2007, Frazer et al , 2015). C. albicans exhibits various cellular morphologies and provides an ideal system to explore various features of MT dynamics during mitosis because of its extraordinary genomic plasticity to tolerate chromosomal alterations and ploidy changes under unfavorable conditions such as the presence of anti-fungal drugs (Sanyal, 2012).…”

Aurora kinase Ipl1 facilitates bilobed distribution of clustered kinetochores to ensure error‐free chromosome segregation in Candida albicans

“…The virulence of the organism has been shown to be associated with the morphological transition between yeast and hyphal cells. In these morphological forms, MT‐associated motor proteins have been shown to regulate nuclear movements, chromosome segregation and cytoskeleton remodeling (Martin et al , ; Finley et al , ; Sherwood and Bennett, ; Frazer et al , ). Both yeast and filamentous hyphal cells exhibit variations in the length of the mitotic spindle, the former having shorter mitotic spindles of maximum of 8 µm and the latter with a longer spindle MTs of maximum of 20 µm, indicating that the cell morphology influences the length of MTs in the same organism (Barton and Gull, ; McCoy et al , ).…”

“…Both yeast and filamentous hyphal cells exhibit variations in the length of the mitotic spindle, the former having shorter mitotic spindles of maximum of 8 µm and the latter with a longer spindle MTs of maximum of 20 µm, indicating that the cell morphology influences the length of MTs in the same organism (Barton and Gull, ; McCoy et al , ). The C. albicans genome encodes only one form each of the kinesin‐5 (Kip1) and kinesin‐14 (Kar3Cik1) family of motor proteins in contrast to the two kinesin‐5 (Kip1 and Cin8) and kinesin‐14 (Kar3Cik1 and Kar3Vik1) family motors in S. cerevisiae (Chua et al , ; Frazer et al , ). In addition, C. albicans exhibits extraordinary genome plasticity to tolerate chromosomal alterations and thus provides an ideal system to explore MT dynamics during mitosis.…”