Worm Breeder's Gazette 15(2): 52 (February 1, 1998)
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.
Lab. of Molecular Biology, Toyohashi University of Technology, Tempaku-cho, Toyohashi 441, Japan.
Previously it has been shown that when the wild type dpy-20 gene is overexpressed ectopically in transgenic animals, it suppresses transcrption of the alpha-1 and alpha-2 tubulin genes in the ventral cord motor neurons very significantly, with little or no effect in the head and tail of the animals (Fukushige and Siddiqui,Transgenic Res.4; 332-340,1995).The dpy-20 IV encodes a non collagen novel protein of 359 amino acid with no significant similarity to any protein in various data bases (Clark et al.1995). We hypothesized that the dpy-20 gene may affect the transcription in ventral cord motor neurons not only of the alpha tubulin genes but perhaps also of other genes that are specific to motor neurons. We have previously shown that the dpy-20 gene interacts with unc-104, unc-116 and osm-3 genes that encode different kinesin motor proteins in C. elegans (C. elegans International Meeting, 1997). To test the notion whether the dpy-20 is a specific regulator of transcription in the ventral cord motor neurons, we have constructed a variety of double mutants using the dpy-20(e2017) amber allele (Hodgkin, 1985), with different uncoordinated mutants (Brenner, 1974), known to be affected in the ventral cord motor neuron development, such as the unc-4(e120) (White et al 1992; Miller et al., 1992), unc-5(e53) (Siddiqui, 1990; Hedgecock et al,1990; Leung-Hagesteijn et al,1992; Hamelin et al.1993), unc-13(e51) (Brenner 1974; Ahmed et al 1992; Nguyen et al. 1995), unc-104(rh1016,rh1017) (Otsuka et al.,1991) , unc-25(e156) (Thomas 1990;McIntire et al. 1993; Reiner and Thomas 1995), unc-51(e369) (Hedgecock et al. 1985; McIntire et al. 1992; Ogura et al.1994), unc-55(e402) (Walthall and Plunkett 1995), unc-70(e524)(Park and Horvitz 1986a; Johnsen and Baillie 1991), unc-86(e1416) (Chalfie et al. 1981; Finney et al.1988; Baumeister et al. 1996), and unc-32(e189)( Brenner 1974; Nguyen et al. 1995) etc. Interestingly, all of these double mutants are severely affected in growth and show great reduction in brood size. These double mutants are clearly slow to grow, mostly sterile, severely uncoordinated, and often die as bag of worms with only one or two progeny. In controls, the double mutants between the dpy-20(e2017) and muscle specific mutants, such as the unc-68(e540) Brenner 1974; Lewis et al. 1980), unc-54(e190) (Epstein et al; McLachlan and Karn 1982;Dibb et al. 1989; Bejsovec and Anderson 1990) unc-15(e73) (Kagawa 1989; Gengyo-Ando and Kagawa 1991; Epstein et al. 1993) were constructed and tested for their viability and brood size. No significant reduction in the brood size was observed. These observations suggest a role of dpy-20 gene in regulating the expression of genes that are expressed in the ventral cord motor neurons. The hypothesis that the dpy-20 gene regulates gene expression in ventral cord motor neurons got further support when we showed that a reporter gene dpy-20::lacZ, expresses in the set of ventral cord motor neurons and hypodermal cells (M. Y. Ali, Z. K. Siddiqui, Diana Janke, David Baillie, and Shahid Siddiqui, unpublished, see the accompanying abstract in this issue of WBG). Genetic data obtained from the double mutants is consistent with the observation that the dpy-20 gene is expressed in the ventral cord motor neurons. Based on these observations we have re-examined the protein sequence of DPY-20 and looked for its identity both from computer homology search and visual inspection. To our surprise (and delight) we found that indeed the DPY-20 shares significant homology in 18 small regions with a variety of transcription factors and regulatory proteins conserved across different species from the yeast to humans. For example, it shows homology with Drosophila ecdysone inducible protein(E75A,E75B) (Segraves WA and Hogness DS,1990), liver specific transcription factor, human(HNFA) (Bach I,1990), homeobox protein DLX-2 (HMD2, human) (Selski D.J, 1993), zinc finger protein, (HRX,mouse)(MA Q,et al.,1993), insulin receptor substrate(IRS1,human) (Nishiyama M, et al.,1992), zinc finger protein (MGF3,mouse) (Passananti C, et al.,1989), renal transcription factor KID-1(rat) (Witzgall R, et al.1993), human basic transcription factor BTF2 (Fischer L,et al.,1992), transcription factor GAGA, Drosophila (Soeller W.C. et al.,1993) and C. elegans homeobox protein lin-39 (Clark SG et al.,1993) etc. Dpy-20 protein shares significant homology(62%) with a small region(19 aa) of Lin -39. The overall structural similarity between the Dpy-20 and Lin-39 is 41%. We propose that the Dpy-20 may be a Lin-39 like transcription factor. It is also possible that dpy-20 is different from all other known transcription factors and defines a novel transcription factor. Interestingly, the dpy-20 genomic sequence contains different DNA binding sequences that are known to bind various transcription factors such as the NK-2 , POU factor and IRF factors etc. Currently, we are doing gel mobility shift assays to test the DNA/protein interaction with respect to examine the interaction of the Dpy-20 protein and different genomic sequences, and also the binding of various factors to the dpy-20 genomic sequences. Whether the Dpy-20 is regulating expression of various genes in the ventral cord motor neurons directly or perhaps controlling the transcription of other regulatory gene(s) remains to be seen. We thank David Baillie, Ken Nishikawa, Johji Miwa, Tony Otsuka, Y. Kohara, T. Motohashi, A. Fire, Ian Hope, R. Hosono, K. Harada, for their help and cooperation and T.Stiernagle for mutant strains.