The gamma-tubulin ring complex (gammaTuRC) is a large multi-protein complex that is required for microtubule nucleation from the centrosome. Here, we show that the GCP-WD protein (originally named NEDD1) is the orthologue of the Drosophila Dgrip71WD protein, and is a subunit of the human gammaTuRC. GCP-WD has the properties of an attachment factor for the gammaTuRC: depletion or inhibition of GCP-WD results in loss of the gammaTuRC from the centrosome, abolishing centrosomal microtubule nucleation, although the gammaTuRC is intact and able to bind to microtubules. GCP-WD depletion also blocks mitotic chromatin-mediated microtubule nucleation, resulting in failure of spindle assembly. Mitotic phosphorylation of GCP-WD is required for association of gamma-tubulin with the spindle, separately from association with the centrosome. Our results indicate that GCP-WD broadly mediates targeting of the gammaTuRC to sites of microtubule nucleation and to the mitotic spindle, which is essential for spindle formation.

The gamma-tubulin complex is a large multiprotein complex that is required for microtubule nucleation at the centrosome. Here we report the purification and characterization of the human gamma-tubulin complex and the identification of its subunits. The human gamma-tubulin complex is a ring of ~25 nm, has a subunit structure similar to that reported for gamma-tubulin complexes from other species, and is able to nucleate microtubule polymerization in vitro. Mass spectrometry analysis of the human gamma-tubulin complex components confirmed the presence of four previously identified components (gamma-tubulin and gamma-tubulin complex proteins [GCPs] 2, 3, and 4) and led to the identification of two new components, GCP5 and GCP6. Sequence analysis revealed that the GCPs share five regions of sequence similarity and define a novel protein superfamily that is conserved in metazoans. GCP5 and GCP6, like other components of the gamma-tubulin complex, localize to the centrosome and associate with microtubules, suggesting that the entire gamma-tubulin complex takes part in both of these interactions. Stoichiometry experiments revealed that there is a single copy of GCP5 and multiple copies of gamma-tubulin, GCP2, GCP3, and GCP4 within the gamma-tubulin complex. Thus, the gamma-tubulin complex is conserved in structure and function, suggesting that the mechanism of microtubule nucleation is conserved.

Androgen has an important role in development of the prostate, and the actions of androgen are mediated, in part, by locally produced growth factors. These growth factors are postulated to mediate stromal-epithelial interaction in the prostate to maintain normal tissue physiology. Transforming growth factor-␣ (TGF-␣) is one of the growth factors that can stimulate prostatic growth. The expression of TGF-␣ is thought to be regulated by androgen. The expression of epidermal growth factor receptor (EGFR), which is the receptor of TGF-␣ and EGF, also may be regulated by androgen. The hormonal and developmental regulation of TGF-␣ and EGFR messenger RNA (mRNA) levels in isolated epithelial and stromal cells from rat ventral prostate was investigated. The expression of mRNA for TGF-␣ and EGFR was analyzed by a quantitative RT-PCR (QRT-PCR) procedure developed. Observations from this assay demonstrated that both epithelial and stromal cells expressed the mRNA for TGF-␣ and EGFR. TGF-␣ mRNA expression was constant during postnatal, pubertal, and adult development of the prostate. EGFR mRNA expression was elevated at the midpubertal period and decreased with age. After castration of 60-day-old adult rats, both TGF-␣ and EGFR mRNA were significantly enhanced. TGF-␣ mRNA expression was stimulated by EGF in stromal cells (4.5-fold increase) but was not changed by any treatment in epithelial cells. EGFR mRNA levels were stimulated by EGF and keratinocyte growth factor treatment and inhibited by testosterone treatment in epithelial cells. Stromal cell EGFR mRNA levels were not affected by any treatment. Both testosterone and EGF stimulated incorporation of 3 H-thymidine into prostatic stromal and epithelial cells. Anti-TGF-␣ antibody significantly inhibited testosterone-stimulated Results indicate that testosterone does not directly regulate TGF-␣ mRNA levels but does inhibit EGFR mRNA levels. Interestingly, anti TGF-␣ antibody suppressed the effect of testosterone on 3 H-thymidine incorporation into prostatic stromal and epithelial cells. This finding suggests that testosterone may act indirectly on prostatic cells to influence TGF-␣ actions. TGF-␣ mRNA levels were influenced by EGF in stromal cells only, and EGFR mRNA levels were influenced by testosterone, EGF, and keratinocyte growth factor in epithelial cells. These observations suggest that regulation of TGF-␣ and EGFR is distinct between the cell types. In conclusion, a network of hormonally controlled growth factor-mediated stromal-epithelial interactions is needed to maintain prostate development and function. (Endocrinology 139: 1369 -1377, 1998)

What is it? γ γ-Tubulin is a member of the tubulin superfamily and is required for nucleating the polymerization of microtubules in vivo. Microtubules are dynamic cytoskeletal polymers that assemble from two other members of the tubulin superfamily, α α-tubulin and β βtubulin. Microtubules make up the mitotic and meiotic spindles, and are important for establishing cell polarity and vesicle trafficking. The tubulin superfamily is still growing, and now includes tubulins δ δ through η η (delta through eta, in case you've forgotten your Greek). α α-, β βand γ γ-tubulin are conserved in all eukaryotic groups, whereas the other superfamily members are not.The editors of Current Biology welcome correspondence on any article in the journal, but reserve the right to reduce the length of any letter to be published. All Correspondence containing data or scientific argument will be refereed. Items for publication should either be submitted typed, double-spaced to: The Editor, Current Biology,