Characterization of a mutant with abnormal flagellar morphogenesis in Chlamydomonas reinhardtii.

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I have characterized a novel flagellar mutant in Chlamydomonas reinhardtii. When the mutant was cultured under the light (-4,800 lux), most cells had no flagella. However, when the cultures were put in the dark, greater than 70% of the cells flagellated within 16 h after the transfer. The flagellar morphology varied from "rod-shape" (same as the wild-type flagella) to "disk-shape" . Electron microscopy showed that the disk-shaped flagella had the axonemes which were curved into a loop within the swollen membrane. On the other hand, flagellated cells (70%*) in the dark were put under the light, all cells had no flagella within 8 h after the transfer. Light inhibition of flagellation was restored in the presence of 10^<-6>-10^<-5> M DCMU. The spectral dependency of the photo-inhibition of flagellation showed maximal effectiveness at 400-420nmand 660-680nm. Flagellar regeneration of the mutant was also inhibited by light, and the inhibition pattern was similar to that of cycloheximide (20 mug/ml). Axonemes of the mutant maintained looped shape after being demembranated with 0.5% Nonidet P-40. SDS PAGE of the mutant flagella showed that two bands with molecular weights of 250 kDa and 7.4 kDa were missing and three other bands with molecular weights 240 kDa, 150 kDa and 130 kDa increased their densities. From these results, I suppose that product(s) of photosynthesis inhibit flagellation of the mutant, and formed flagella have defect(s) in the binding between the axoneme itself. Tetrad analysis showed that phenotypes of the mutant resulted from a single gene mutation.