Program of the 2001 International Worm Meeting - Sternberg Lab ...

258
ClC-2 and CLH-3 share 40% amino acid
identity. The strong conservation of biophysical
and structural properties suggests that CLH-3
and ClC-2 may have similar physiological roles
and regulatory mechanisms. Because ClC-2 is
activated by cell swelling, it has been proposed
to function in cell volume homeostasis, a
housekeeping process conserved in widely
divergent species. However, knockdown of
CLH-3 activity by dsRNA has no effect on the
rate or extent of oocyte volume regulation.
Basic mechanisms of cell cycle control are
conserved in all eukaryotic organisms. We
observed that CLH-3 is constitutively active in
oocytes undergoing meiotic maturation. These
findings suggest that the channel may regulate
cell cycle-associated physiological processes
important for reproduction. To test this
hypothesis, we quantified several reproductive
events. Brood size and the timing of meiotic
maturation, ovulation and fertilization, and
embryonic development were unaffected by
dsRNA-induced knockdown of CLH-3.
Meiotic maturation induces ovulatory
contractions of gap junction-coupled gonadal
sheath cells. Sheath contractions were initiated
~2 min earlier in dsRNA-injected versus
buffer-injected control animals. These results
indicate that CLH-3 functions in part to
modulate ovulatory contractions of gap
junction-coupled gonadal sheath cells. We
suggest that this modulation occurs by
CLH-3-induced membrane depolarization,
which in turn may modulate sheath cell Ca 2+
signaling pathways.
259. DYSTROPHIN AND
ASSOCIATED PROTEINS IN C.
elegans.
Gieseler, K, Grisoni, K, Mariol, M-C,
Segalat, L
University of Lyon, CGMC, 43 bd du 11
Novembre, 69100 Villeurbanne, France.
259
Dystrophin is the product of the gene mutated in
Duchenne muscular dystrophy, a neuromuscular
disease leading to muscle necrosis. The function
of the dystrophin protein is not known. In
mammals, dystrophin is located under the
muscle plasma membrane, and is associated
with a protein complex (DGC) spanning the
membrane. We reported earlier that the C.
elegans genome contains a dystrophin homolog
named dys-1. Our goal is to understand
dystrophin function in C. elegans.
Loss-of-function mutations in the dys-1 gene do
not alter the muscle structure, but make animals
hyperactive and slightly hypercontracted. In a
forward genetics approach, we isolated
additional mutations with dys-1-like
phenotypes. Mapping and complementation
analysis revealed that these mutations account
for 4 additional genes. We cloned two of them :
dyb-1 and dyc-1. dyb-1 is the C. elegans
homologue of dystrobrevins, a family of
proteins belonging to the dystrophin complex
and known to bind dystrophin directly through
coiled-coil domains. dyc-1 has no known
homologue. Like DYS-1 and DYB-1, it contains
a putative coiled-coil domain. Preliminary data
indicate that DYC-1 may bind to DYB-1 in
vitro, possibly through the coiled-coil domain.
Interestingly dyc-1 has muscular and neuronal
isoforms. A gfp protein fusion with the neuronal
isoform stains approximately 20 neurons in the
animal. Staining is these neurons appears as dots
along the neuronal processes. We are currently
investigating the nature of these dots. We also
created a dystrophin-dependent myopathy in C.
elegans by associtaing dys-1 with a mild MyoD
mutation that potentializes dys-1. This provides
us with a tool to undertake suppresor screens
which are under way.