Fig. 8. Ectopic expression of the GRIP domain causes centrosome hypertrophy. To investigate the function of the Cbs GRIP domain we expressed a GFP::GRIP fusion protein during embryogenesis under control of nanos::Gal4 and immunostained embryos for Cnn (green, A,E,I) or microtubules (green, M), GFP (red, B,F,J,N), and Cbs (blue, O) or DNA stained with TOTO-3 (blue, C,G,K). (A-D) During early prophase the GRIP domain initially localizes to one centrosome, resulting in an asymmetric accumulation of Cnn at replicated centrosomes (arrowheads). A cytoplasmic pool of the GRIP domain is also present around nuclei during prophase. (E-H) When the GRIP domain remains concentrated at centrosomes during prophase, individual centrosomes (arrowhead) and pairs of centrosomes (arrow), undergo additional rounds of replication and frequently become dissociated from nuclei. (I-P) During metaphase the GRIP domain accumulates as cytoplasmic particles and localizes strongly at centrosomes and the microtubule spindle, but does not cause further accumulation of Cnn at centrosomes or centrosome hypertrophy. Low levels of the GRIP domain are present at DNA (J,N), but the transport efficiency and DNA-binding affinity of endogenous Cbs appears higher (O). However, the GRIP domain does cause excessive accumulation of cytoplasmic Cbs particles that are not seen in wild-type embryos during mitosis. Bars, 20 µM (A-D, I-P); 5 µM (E-H).