Essential role for gamma -tubulin in the acentriolar female meiotic spindle of Drosophila

Abstract

Microtubule nucleation in vivo requires γ‐tubulin, a highly conserved component of microtubule‐organizing centers. In Drosophila melanogaster there are two γ‐tubulin genes, γTUB23C and γTUB37C. Here we report the cytological and molecular characterization of the 37C isoform. By Western blotting, this protein can only be detected in ovaries and embryos. Antibodies against this isoform predominantly label the centrosomes in embryos from early cleavage divisions until cycle 15, but fail to reveal any particular localization of γ‐tubulin in the developing egg chambers. The loss of function of this gene results in female sterility and has no effect on viability or male fertility. Early stages of oogenesis are unaffected by mutations in this gene, as judged both by morphological criteria and by localization of reporter genes, but the female meiotic spindle is extremely disrupted. Nuclear proliferation within the eggs laid by mutant females is also impaired. We conclude that the expression of the 37C γ‐tubulin isoform of D.melanogaster is under strict developmental regulation and that the organization of the female meiotic spindle requires γ‐tubulin.

Assembly pathway of the anastralDrosophilaoocyte meiosis I spindle

“…and S.A.E., unpublished). Mutants of γ-Tub37C have been reported previously to disrupt oocyte meiosis I spindles (Tavosanis et al, 1997;Wilson and Borisy, 1998), although the abnormal meiotic figures in one of these studies may have been caused by inadvertent activation of the oocytes, based on their subsequently reported absence in ovulated oocytes or embryos (Llamazares et al, 1999). The role of γ-tubulin37C in meiosis I spindle assembly in Drosophila oocytes remains an open question that will probably require expression and careful analysis of a fluorescently labeled γ-tubulin protein to answer definitively.…”

“…To examine this idea, we analyzed oocytes of a γ-Tub37C mutant, APL10, produced by APL10/+ heterozygous or APL10 homozygous females carrying ca nd and three or four copies of ncdgfp*. APL10 affects the maternally expressed γ-tubulin at cytological map position 37C, which is present in oocytes and early embryos of Drosophila (Tavosanis et al, 1997). Nine of 16 APL10/+ oocytes (Fig.…”

TheDrosophilaγ-Tubulin Small Complex Subunit Dgrip84 Is Required for Structural and Functional Integrity of the Spindle Apparatus

“…The function of ␥-tubulin has been studied extensively in several organisms and in cultured cells using antibody microinjection, mutants or gene silencing (Oakley et al, 1990;Horio et al, 1991;Joshi et al, 1992;Sunkel et al, 1995;Marschall et al, 1996;Spang et al, 1996 Ruiz et al, 1999;Bobinnec et al, 2000;Sampaio et al, 2001;Raynaud-Messina et al, 2004). For example, in Drosophila where two ␥-tubulin isotypes are expressed, homozygous ␥-tub 23C mutants die during late larval stage, exhibiting atypical mitotic spindles and abnormal centrosomal structures (Sunkel et al, 1995), whereas disruption of the ␥-tub 37CD gene results in abnormal female meiotic spindles (Tavosanis et al, 1997). However, our knowledge about the function of the ␥-tubulin-interacting proteins is limited.…”

HorkaD, a Chromosome Instability-Causing Mutation in Drosophila, Is a Dominant-Negative Allele of lodestar

“…) Characterization of mutant phenotypes: /-females and males were dissected and fixed according to González and Glover (1993). The stage 14 oocytes were immunostained according to Tavosanis et al (1997). The eggs and the embryos were prepared as follows: the chorion was removed by Clorox, the dechorionated embryos were fixed in a 1:1 mixture of 4% paraformaldehyde:heptane or in a 1:1 mixture of methanol:heptane, and the vitelline membrane was removed subsequently by agitation in a mixture of heptane and methanol.…”