2000 West Coast Worm Meeting abstract 91
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| 1 | Department of Biology, Dartmouth College, Hanover, NH 03755 |
| 2 | Fox Chase Cancer Center, Philadelphia PA |
The heterochronic gene pathway controls the timing of larval developmental events. Central to the action of this pathway is the developmental down-regulation of lin-14 and lin-28, whereby high levels specify L1 events, and lower levels specify L2 and L3 events. The decrease in lin-14 and lin-28 results from translational repression by the small RNA product of lin-4. lin-4 RNA is complementary to sequences in the 3' UTR of its target mRNAs, and recent experiments suggest that lin-4 acts by gating access to the 3'UTR by other positive and negative inputs. These other inputs include a mutual positive feedback between lin-14 and lin-28 and a daf-12-dependent negative regulatory input.
The outputs of the heterochronic gene pathway are the expression of cell-type specific developmental programs. To seek downstream targets of lin-14 for L2 events, we employed a microarray of 12,000 ORFs prepared by the Kim lab at Stanford University. Probes were prepared from mRNAs of synchronized mid-L1 populations of N2 and lin-14(n179ts) animals. Among the genes that displayed lin-14-dependent expression by microarray analysis, we chose four genes for follow-up by Northern and GFP-fusions. By Northern analysis, three of these four genes show precocious mRNA expression in lin-14(lf) animals, consistent with their temporal regulation by lin-14 in the wild type. The expression of GFP promoter fusions suggests that at least one of the genes, ins-33, appears to be developmentally regulated by lin-14 at the transcriptional level.
For L3 events, the chief effectors downstream of lin-14 and lin-28 appear to be daf-12 and lin-46. lin-46 encodes a member of a conserved family of proteins involved in protein-protein interactions, suggesting a complex of regulators controling L3 targets. In the vulva precursor cells, L3 events controled by the heterochronic genes are progress through the G1/S phase of the cell cycle and the acqusition of competence to express vulval cell fates. VPC G1/S is regulated by the transcriptional activation of a cyclin kinase inhibitor gene, cki-1. We are identifying regions of the cki-1 promoter which mediate the VPC-specific control of cki-1 transcription by the heterochronic gene pathway.