Worm Breeder's Gazette 14(4): 70 (October 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.
| 1 | Dept. of Molecular Biology and Biochemistry, Rutgers University, New Brunswick, NJ 08855 |
| 2 | Dept. of Biochemistry, New York University, Medical School, New York, NY 10016 |
unc-8 encodes a member of the C. elegans degenerin family (as
was predicted by Shreffler and co-workers), that is extensively
expressed in the nervous system including motor neurons, interneurons
and sensory neurons. Degenerins are thought to assemble into
mechanically-gated ion channels. Gain-of-function mutations in unc-8
result in transient swelling of neurons, reminiscent of the vacuolation
phenotype induced by specific mutations in other family members. As a
consequence, movement is severely uncoordinated and animals are unable
to back up. These phenotypes are suppressed by lesions in the mec-6 gene
that also suppress the effects of analogous mutations in other
degenerins.
We have sequenced the semi-dominant e15, e49 and n491 unc-8
alleles. The two strongest mutations, e15 and n491, alter the same
residue in the extracellular region before the cysteine-rich domain II
(G387E). Interestingly, these mutations map in the center of a domain
previously implicated by Garcia-Anoveros and co-workers in regulation of
channel closing; mutations affecting this domain in two other
degenerins, deg-1 and mec-4, result in vacuolation and degeneration of
cells expressing the mutant alleles. The mutation in e49 affects a
conserved residue within cysteine-rich domain III (A586T).
Shreffler and Wolinsky had generated additional second-site
mutations in unc-8 that either suppressed (allele e15lb129) or enhanced
(allele n491lb82) the phenotype of the e15 and n491 unc-8 alleles when
in trans to the wild type gene. lb129 results in a substitution of
tyrosine for an absolutely conserved histidine at position 114, in the
intracellular part of the protein before the membrane-spanning domain I.
lb82 encodes a threonine to isoleucine change at position 664 in the
extracellular region between cysteine-rich domain III and
membrane-spanning domain II. The unusual genetic properties of these two
alleles suggest that at least two UNC-8 molecules participate in the
formation of a channel and that these molecules interact both through
intracellular and extracellular domains. These interactions might serve
to ensure a rigid channel structure that could respond to rather than
absorb mechanical forces.
C. elegans strains carrying unc-8 loss-of-function mutations do
not exhibit readily noticeable defects in locomotion. However, close
examination of the sinusoidal path inscribed by unc-8(lf) worms on
bacterial lawns revealed that the amplitude as well as the wave-length
of the track is substantially reduced. This striking phenotype suggests
that UNC-8 functions to regulate sinusoidal locomotion. unc-8 together
with an additional member of the degenerin family, del-1 (for
degenerin-like) that resides on cosmid clone E02H4 derived from the X
chromosome, is expressed at high levels in command motor neurons of the
ventral nerve cord (unc-8 in the D and V class and del-1 in the V class
of motor neurons).
In early electron-microscopy studies of the C. elegans nervous
system White and co-workers had noticed an unusual feature of these
neurons. The endings of their long longitudinally running processes were
undifferentiated and devoid of any synaptic input. This morphology
(which resembles the morphology of another group of sensory neurons-the
six touch receptors) prompted investigators to hypothesize that motor
neuron endings functioned as stretch receptor areas that could modify
the overall activity of the neurons, thus modulating locomotion. This
attractive scenario called for the existence of mechanically-gated
channels in motor neurons. Given the expression of two degenerins, unc-8
and del-1, in these neurons and the striking phenotype of unc-8(lf)
mutations, we propose that UNC-8 and DEL-1 assemble into
stretch-activated channels in motor neurons. By receiving input from
parts of the body stretched by muscle contraction, such channels would
provide the feedback needed to accordingly modulate muscle activity and
sustain a normal sinusoidal pattern of movement.