Figure 4. Depletion of CeGrip-1 blocks recruitment of centrosomal gamma-tubulin. (A) CeGrip-1 contains the two grip motifs found in proteins that form complexes with gamma-tubulin (Gunawardane et al., 2000). By performing further BLAST searches using the grip motifs as a query sequence, we recently identified a second predicted C. elegans protein, CeGrip-2 (C45G3.3), that contains the grip motifs. Here, grip motifs 1 and 2 from CeGrip-1 and CeGrip-2 are aligned with the grip motifs from other proteins homologous to Dgrip91/Spc98p. Amino acids conserved in 80-100% of the sequences are highlighted in yellow. The conserved amino acids that define grip motifs 1 and 2 (Gunawardane et al., 2000) are shown beneath the alignments in red and green, respectively. (B) Panels summarizing a time-lapse sequence of a wild-type (left) and a
gip-1(RNAi) embryo (right) expressing both GFP-histone and GFP-gamma-tubulin. In
gip-1(RNAi) embryos, the gamma-tubulin signal was greatly reduced or, as in this case, absent (see Videos 1 and 2 located at
http://www.jcb.org/cgi/ content/full/jcb.200202047/DC1). (C) Mitotic wild-type and
gip-1(RNAi) embryos stained for MTs and DNA (green and red, top), gamma-tubulin (middle), and CeGrip (bottom) are shown. CeGrip-1 and gamma-tubulin colocalize at centrosomes. Depletion of CeGrip-1 prevents gamma-tubulin recruitment to centrosomes and results in an MT phenotype essentially identical to that observed in
tbg-1(RNAi) embryos. Depletion of CeGrip-2 also results in an MT phenotype essentially identical to that in
tbg-1(RNAi) embryos (unpublished data). Bars, 10 um.