Tandem affinity purification of the <i>Candida albicans</i> septin protein complex

Kaneko, Aki; Umeyama, Takashi; Hanaoka, Nozomu; Monk, Brian C.; Uehara, Yoshimasa; Niimi, Masakazu

doi:10.1002/yea.1147

Cited by 56 publications

(36 citation statements)

References 23 publications

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“…Quantification of the proteins levels indicated that the Cdc10/Sep7 ratios were similar during yeast and hyphal growth ( Figure 3E). These results are in good agreement with previous findings indicating that the stoichiometry of the septin complex is not affected by cell morphology (Kaneko et al, 2004). Thus, the composition of the septin ring does not substantially change during the switch from the yeast to hyphal form.…”

supporting

confidence: 93%

Sep7 Is Essential to Modify Septin Ring Dynamics and Inhibit Cell Separation duringCandida albicansHyphal Growth

González-Novo

Correa‐Bordes

Labrador

et al. 2008

MBoC

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When Candida albicans yeast cells receive the appropriate stimulus, they switch to hyphal growth, characterized by continuous apical elongation and the inhibition of cell separation. The molecular basis of this inhibition is poorly known, despite its crucial importance for hyphal development. In C. albicans, septins are important for hypha formation and virulence. Here, we used fluorescence recovery after photobleaching analysis to characterize the dynamics of septin rings during yeast and hyphal growth. On hyphal induction, septin rings are converted to a hyphal-specific state, characterized by the presence of a frozen core formed by Sep7/Shs1, Cdc3 and Cdc12, whereas Cdc10 is highly dynamic and oscillates between the ring and the cytoplasm. Conversion of septin rings to the hyphal-specific state inhibits the translocation of Cdc14 phosphatase, which controls cell separation, to the hyphal septum. Modification of septin ring dynamics during hyphal growth is dependent on Sep7 and the hyphal-specific cyclin Hgc1, which partially controls Sep7 phosphorylation status and protein levels. Our results reveal a link between the cell cycle machinery and septin cytoskeleton dynamics, which inhibits cell separation in the filaments and is essential for hyphal morphogenesis. INTRODUCTIONCandida albicans is an opportunistic human fungal pathogen that is used as a model to study morphogenetic changes in single-cell organisms. C. albicans is capable of growing with different morphologies ranging from the yeast form to elongated filaments in response to environmental signals such as pH, temperature, or the presence of serum (Odds, 1988;Brown and Gow, 1999;Sudbery, 2001). The ability to switch between the different morphologies is of key importance for the pathogenicity of this organism (Lo et al., 1997;Zheng et al., 2004).Septins are GTP-binding proteins that assemble into homo-and hetero-oligomers and filaments, and they are an important element in morphogenesis in animals and fungi. They assemble a ring-like structure at the site of cytokinesis (for recent reviews, see Gladfelter et al., 2001;Faty et al., 2002;Douglas et al., 2005;Versele and Thorner, 2005). The structure, dynamics and regulation of septin rings is well known in the budding yeast Saccharomyces cerevisiae, where the filaments form a collar at the bud neck that is composed of Cdc3, Cdc10, Cdc11, Cdc12, and Shs1/Sep7. Initially, these septins form a ring at the bud site before bud emergence, which then rearranges into an hourglass-shaped collar that spans both the mother and the daughter sides of the neck (Gladfelter et al., 2005;Kozubowski et al., 2005). This collar persists until cytokinesis, when it splits into two rings (Cid et al., 2001;Lippincott et al., 2001). The functions of the septins are of at least two types: first, they act as scaffold for anchoring other regulatory proteins at the bud neck. Second, they form a diffusion barrier to control molecules trafficking between mother and daughter cells (for recent reviews, see Longtine and Bi, 2003;Dougla...

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supporting

confidence: 93%

Sep7 Is Essential to Modify Septin Ring Dynamics and Inhibit Cell Separation duringCandida albicansHyphal Growth

González-Novo

Correa‐Bordes

Labrador

et al. 2008

MBoC

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“…The data indicate evolutionary conservation of this protein complex in fungi. An unexpected finding is Hof1, which is known to localize to the bud neck and has a role in cytokinesis, but has not been found in purified septin complexes previously (Altman and Kellogg, 1997;Carroll et al, 1998;Kaneko et al, 2004). The interaction was further confirmed by co-immunoprecipitation (data not shown).…”

Section: Gin4 Phosphorylates Sep7mentioning

confidence: 87%

CDK Regulates Septin organization through Cell-cycle-dependent Phosphorylation of the Nim1-related Kinase Gin4

Yong

Wang

et al. 2012

Journal of Cell Science

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SummaryCyclin-dependent kinases (CDKs) regulate septin organization in a cell-cycle-dependent manner in yeast. However, the mechanism remains unclear. Here, we show that the Candida albicans CDK Cdc28 phosphorylates the Nim1-related kinase Gin4, a known septin regulator, activating its kinase activity, which in turn phosphorylates the Sep7 septin. Gin4 contains a cluster of CDK phosphorylation sites near the kinase domain. Replacing serine/threonine with alanine in these sites prevents Gin4 activation, weakens its association with Sep7, alters Sep7 dynamics and causes morphological and cytokinetic defects. By contrast, phosphomimetic mutation enhances the kinase activity with only moderate deteriorating effects. We also found that Gin4 has both kinase-independent and -dependent functions, acting during G1 phase and mitosis, respectively, with the former being essential for septin ring assembly. Thus, we have identified a previously unknown signaling pathway linking CDKs and the septins that provides new insights into the mechanisms controlling septin organization and function in coordination with cell-cycle phases.

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“…At present, we consider CaBig1p to be an adaptor protein rather than a functional enzyme because neither CaBig1p nor ScBig1p contains any amino acid motifs associated with carbohydrate-active enzymes. Using a tandem affinity purification technique, we are currently investigating whether CaBig1p and ScBig1p have binding partner proteins (15).…”

Section: Vol 74 2006mentioning

confidence: 99%

Deletion of the Ca BIG1 Gene Reduces β-1,6-Glucan Synthesis, Filamentation, Adhesion, and Virulence in Candida albicans

et al. 2006

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The human fungal pathogen Candida albicans is able to change its shape in response to various environmental signals. We analyzed the C. albicans BIG1 homolog, which might be involved in ␤-1,6-glucan biosynthesis in Saccharomyces cerevisiae. C. albicans BIG1 is a functional homolog of an S. cerevisiae BIG1 gene, because the slow growth of an S. cerevisiae big1 mutant was restored by introduction of C. albicans BIG1.

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Tandem affinity purification of the Candida albicans septin protein complex

Cited by 56 publications

References 23 publications

Sep7 Is Essential to Modify Septin Ring Dynamics and Inhibit Cell Separation duringCandida albicansHyphal Growth

Sep7 Is Essential to Modify Septin Ring Dynamics and Inhibit Cell Separation duringCandida albicansHyphal Growth

CDK Regulates Septin organization through Cell-cycle-dependent Phosphorylation of the Nim1-related Kinase Gin4

Deletion of the Ca BIG1 Gene Reduces β-1,6-Glucan Synthesis, Filamentation, Adhesion, and Virulence in Candida albicans

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