Essential role of tubulin-folding cofactor D in microtubule assembly and its association with microtubules in fission yeast

Abstract

The main structural components of microtubules are alpha- and beta-tubulins. A group of proteins called cofactors are crucial in the formation of assembly-competent tubulin molecules in vitro. Whilst an in vitro role is emerging for these cofactors, their biological functions in vivo remain to be established. In order to understand the fundamental mechanisms that determine cell polarity, we have screened for fission yeast mutants with altered polarity. Here we show that alp1+ encodes a homologue of cofactor D and executes a function essential for cell viability. A temperature-sensitive alp1 mutant shows a variety of defects including abnormal mitoses, loss of microtubule structures, displacement of the nucleus, altered growth polarity and asymmetrical cell division. Overexpression of Alp1 is lethal in wild-type cells, resulting in altered cell shape, but is rescued by co-overexpression of beta-tubulin. Alp1 co-localizes with microtubules, both interphase arrays and mitotic spindles. Furthermore, Alp1 binds to and co-sediments with taxol (paclitaxel)-stabilized porcine microtubules. Our results suggest that, in addition to a function in the folding of beta-tubulin, cofactor D may play a vital role in microtubule-dependent processes as a microtubule-associated protein.

Revisiting the tubulin folding pathway: new roles in centrosomes and cilia

“…This is supported by observations in Schizosaccharomyces pombe where mutations in the alp1 gene, the TBCD homolog, cause G1/S arrest (82)(83)(84). Furthermore, TBCD depletion by RNAi in mammalian cells resulted in the formation of mono-and multipolar spindles, disorganized/short spindles, long midbodies and failure in cytokinesis (78,79).…”

Overproduction of elongation factor 1α, an essential translational component, causes aberrant cell morphology by affecting the control of growth polarity in fission yeast

“…Curled microtubules were frequently observed in the fission yeast alp (Radcliffe et al 1998;Hirata & Toda, unpublished observations) or tea1 mutant cells (Mata & Nurse 1997). Some alp genes encode microtubule-associated proteins that are important for the expression of the function of the microtubule (Hirata et al 1998a;Radcliffe et al 1998). Tea1 also functions in the microtubule cytoskeleton and is important for generating global spatial order (Mata & Nurse 1997).…”

“…It was shown earlier that altered polarity mutants may have defects in microtubular organization (Toda et al 1983;Yaffe et al 1996;Mata & Nurse 1997;Hirata et al 1998a). To investigate whether the over-expression of efa1 þ affects the organization of the microtubule cytoskeleton, we examined tubulin organization in efa1 þ -over-expressing cells using TAT-1 anti-tubulin antibody.…”

A Trimer Consisting of the Tubulin-specific Chaperone D (TBCD), Regulatory GTPase ARL2, and β-Tubulin Is Required for Maintaining the Microtubule Network

“…In Saccharomyces cerevisiae, screens for altered sensitivity to drugs acting on microtubules identified mutations in the yeast TBCD ortholog CIN1 (8,9). Mutations in TBCD orthologs were later identified in similar screens performed in Schizosaccharomyces pombe (Alp1) (10,11) and Arabidopsis thaliana (TTN1). Along with these compelling genetic data, the identification of TBCD as one of the five tubulin-specific cofactors required for in vitro folding of tubulin (5) confirmed a central role of TBCD in tubulin biosynthesis.…”