Worm Breeder's Gazette 14(2): 41 (February 1, 1996)
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.
Department of MCDB, University of Colorado, Boulder
We have reported previously on the cloning, molecular
characterization, expression, and ectopic overexpression of a C.
elegans TGF-beta homologue (previously called ceg-1: WBG
13(4):65, 1994; 1995 Meeting Abstracts: 555), which we have now
named dbl-1 (dpp-BMP2,4-like). We have continued analysis of
dbl-1 expression, using a transgenic him-8 line isolated after
coinjection of two dbl-1::GFP constructs containing 4.4 kb of
upstream sequence and differing only in the presence or absence
of a nuclear localization signal. The expression of both
reporters allows us to identify cells both by nuclear position
and by cell body and axonal morphology. Fluorescing cells in
adults include the anal depressor muscle, neurons of the retro-
vesicular ganglion, and VNC motor neurons including those of the
pre-anal ganglion and the PDA motor neuron. Male-specific
expression is seen in the CP neurons of the VNC, in cells of the
dorsal-rectal ganglion, and in two spicule associated neurons of
adults, as well as transiently in several still unidentified tail
cells of L3-L4 larvae.
We have also continued to analyze effects of ectopic dbl-1
overexpression in transgenic lines carrying a dbl-1 cDNA
construct (hsp::dbl-1) made with the heat-shock promoter plasmid
pPD49.78, which induces well in neuronal and hypodermal cells (A.
Fire, personal communication). Since both the dbl-1 expression
pattern and existing evidence (Savage et al., 1995 Meeting
Abstracts: 56) suggest a role for signaling mediated by a TGF-
beta homologue during ray and spicule formation, we examined the
effects of dbl-1 overexpression on ray morphogenesis. Heat
shocks of hsp::dbl-1 transgenic him-8 L3 males result in a
variety of ray transformations, the most common of which (50%
penetrance) is transformation of ray 4 to ray 3 with 4-3 fusion,
a phenotype also associated with decreased mab-5 activity (Chow
and Emmons, Development 120:2579, 1994). Other phenotypes seen
at lower frequencies include extra V-rays and multiple-ray
fusions. These results suggest a role for dbl-1 in the
specification of ray identities.
We have refined the genetic map position of dbl-1 on LGV by
using PCR to assay for dbl-1 sequences in DNA from inviable
embryos homozygous for each of several mapped deficiencies in the
region. The results locate dbl-1 in a small region of <.5 map
unit (uncovered by nDf32 but not by nDf18) which includes unc-70
and four other genes defined by lethal mutations (Johnsen and
Baillie, Genetics 129:735, 1991). To determine whether any of
these genes could be dbl-1, we have done DNA coinjection rescue
experiments with a rol-6 marker plasmid and cosmid ZC421, which
contains the dbl-1 coding region and approximately 20kb of
upstream sequence. We have so far injected homozygous unc-70
hermaphrodites as well as heterozygous hermaphrodites from
balanced strains carrying lethal mutations representing three of
the four essential genes. None of these mutants was rescued. We
have also undertaken PCR screening of a high-copy-number Tc1
strain (Andachi and Kohara, 1995 Meeting Abstracts: 96) in
attempts to identify nearby Tc1 insertions that would allow
generation of a dbl-1 allele.