Worm Breeder's Gazette 17(2): 36 (April 1, 2002)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
1Department of Molecular Genetics and 2Program in MCDB, Ohio State University, Columbus, OH 43210.
Pax transcription factors play an important role in organ development in animals. Pax factors are subdivided into four subclasses based on sequence similarity and the presence of other sequence motifs. In C . elegans there are five Pax genes with one member for three of the subclasses but two Pax 2/5/8 members: egl-38 and K06B9.5 (1).Although these two genes are highly similar in their coding region and bind similar DNA targets (2) they are not functionally redundant. egl-38 is essential for viability, and mutations can disrupt development of distinct organs, including the hindgut and the egg-laying system (3). We hypothesize that the unique functions of egl-38 and K06B9.5 result from differences in expression pattern between the two genes.
To test this idea, we have investigated the expression pattern of K06B9.5. We generated reporter transgenes by tagging the 3'end of K06B9.5 with gfp. We found that these transgenes express in two cell types: The PVC neurons in the tail and the D cells of the vulva (vulD). To confirm that the vulD expression is specific to cell type rather than position, we have crossed the K06B9.5::gfp transgenes into vulval lineage mutants (Table 1). In wild type two vulval cells express K06B9.5::gfp, with one vulD produced from each of the 2° lineages of P5.p and P7.p. In lin-15 mutants we frequently observe additional ectopic expression coincident with cells that produce ectopic 2° lineages. In addition, the expression in P7.p-derived cell is missing or misplaced in some lin-17 mutants consistent with the expected lineage defects. Using a deletion analysis, we have localized sequences important for vulD expression to 150bp of the K06B9.5 promoter. Prelimenary RNAi experiments indicate that both egl-38 and K06B9.5 are affected by double-stranded RNA from either gene. Currently we are trying to study the function of K06B9.5 alone by expressing sense and antisense RNA under the control of K06B9.5 promoter and tagging K06B9.5 with the activator VP16, and with the engrailed repressor domain.
To better understand the relationship between egl-38 and K06B9.5, we have isolated the Pax-2/5/8 family gene from C . briggsae and found that there is only one copy of this gene in this species. This result is confirmed by the recent completion of the C . briggsae genome sequence. A sequence comparison between the genes between C . elegans and
C . briggsae indicate that C . elegans egl-38 and K06B9.5 are more similar to each other than to the C . briggsae Pax-2/5/8 gene, suggesting egl-38 and K06B9.5 result from a gene duplication subsequent to the divergence of the C . elegans and C . briggsae lineages. We have generated a GFP reporter transgene for the C . briggsae Pax-2/5/8 gene. In the larva, this transgene is expressed in two tail neurons (PVC), two neurons in the head, uterine and D cells in the vulva. Characterization of embryonic expression is ongoing. RNAi of this gene in C.briggsae showed defects in egg-laying, hindgut, tail morphology and a low level of larval lethality.
Table 1
Genetic background | No of vulval cells expressing K06B9.5::GFP transgene | ||||
0 | 1 | 2 | 3 | 4 | |
N2 (Wild Type) | 30 | ||||
lin-15(n309) | 4 | 19 | 3 | ||
lin-17(n671) | 3 | 20 + 5 a | |||
a Two cells express but P7.p derived cell is misplaced.
References :
124 : 3919-3928