Worm Breeder's Gazette 16(5): 39 (February 1, 2001)
Samuel Lunenfeld Research Institute of Mt. Sinai Hospital, Toronto, Ontario, Canada M5G 1X5
In C. elegans hermaphrodites, the gonad acquires a U-shape by the directed migration of distal tip cells (DTCs). They start migration at late L2 stage from the ventral mid body of the animal, and by late L4 stage they finish migration. The first phase of DTC migration is longitudinal migration along the ventral muscle bands. The second phase is a circumferential migration, where DTCs migrate from ventral to dorsal along the epidermis, then finally they migrate retrogradely along the dorsal muscle band back to mid-body. Various mutants showing defects in different phases have been described. In gon-1, the first longitudinal phase is defective and the DTC stays there forming a very short gonad, which is sterile (1). unc-5, unc-6, unc-40, and unc-130 have defects in the second phase. DTCs fail to migrate dorsally, resulting in ventral reflection of the gonad arms (2,3). In mig-17, both longitudinal and circumferential guidance are affected resulting in supernumerary turns of the DTC (4).
We have characterized two phenotypic classes of mig-6 mutants. In conventional class 1 mutant, as described previously (5), the first longitudinal migration is defective, but ventral to dorsal migration is normal, which is partly similar to that of gon-1. By a large scale F1 screen for DTC migration mutants, we have obtained a new class of mig-6 mutant (class 2). Heterozygotes of class 2 mutants display ventral reflection of DTCs that is similar to the defect in unc-5, 6, 40, and 130. Interestingly, homozygotes of this class 2 are embryonic/early larval-lethal but escapers have DTCs that wander along both body axes, similar to mig-17. Thus, mig-6 affects all of the three phases of DTC migration but separately.
Both classes of mig-6 are rescued by a genomic fragment containing a single locus. There are two possible ORFs predicted by genome sequencing consortium and Yuji Kohara's EST clones. The two alternative transcripts, designated mig-6a and mig-6b, have the same exon structure down to exon 10 and have different 3' extensions. Northern blot analysis revealed two mRNA species, corresponding to the two predicted sizes. By designing dsRNA based on transcript specific 3' sequence, each transcript was disrupted independently or together. Quite surprisingly, it was shown that RNAi of mig-6b phenocopied the class 1 mutant, and RNAi of mig-6a alone or together with mig-6b phenocopied class 2 mutant recessive phenotype. All four class 1 alleles are nonsense mutations that affect just the mig-6b transcript. At least one of the class 2 mutations is a missense codon affecting both transcripts. Preliminary results suggest that expression of the two transcripts is under distinct regulation. Hence the expression patterns of the two transcripts, in addition to structural difference may define the class 1 and class 2 phenotypes.
(1) Blelloch and Kimble (1999) Nature 399, 586-590
(2) Hedgecock et al. (1990) Neuron 2, 61-85
(3) Nash et al. (2000) Genes & Development 14, 2486-2500
(4) Nishiwaki et al. (2000) Science 288, 2205-2208
(5) Hedgecock et al. (1987) Development 100, 365-382