Worm Breeder's Gazette 11(4): 92

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.

A Zygotic Embryonic Lethal Mutant Defective in Specifying Multiple Embryonic Cell Fates, Gastrulation, and Cell Adhesion

Joel H. Rothman

Among the collection of zygotic embryonic lethal mutants described 
in the accompanying article are five mutants (e2482, e2500, e2504, 
ZE21, and ZE334) that arrest with ~500 cells but in which the 
hypodermis has not enclosed the embryo.  The hypodermal cells cluster 
on one side of the embryo, exposing a field of neurons on the opposite 
side; in e2482 and e2500, and probably the other mutants, this 
hypodermal cluster resides on the dorsal side.  In the hope of 
learning more about embryonic hypodermal development, I have begun an 
in-depth analysis of this class of mutants.
e2482 V, located between unc-76 and dpy-21, exhibits a number of 
cell fate and lineage alterations.  e2482 is probably a null, since 
yDf7, which covers the region in which e2482 maps, yields an identical 
phenotype when homozygous; other nearby deficiencies do not.  e2482 
mutants differentiate hypodermal antigens (hypodermal desmosomes and 
an anti-engrailed-reactive antigen; see WBG, v.10, no.3, p.158) but 
appear to make too few hypodermal cells.  An antigen specific to seam 
hypodermal cells is absent, indicating that seam cell differentiation 
is blocked.  Only about 36 body wall muscle cells are made (the exact 
number is variable) instead of the usual 81, and most of these muscles 
are located at the posterior of the embryo.  Differentiation of gut 
cells occurs, but most embryos lack pharynx muscles, as determined by 
staining for a late antigen (myosin C) and the earlier pharynx muscle 
antigen recognized by MAb 3NB12 (Priess and Thomson, Cell 48, 241-250 (
1987)).  Some embryos do produce a few (~1-3) pharynx muscle cells.  
The failure to express the 3NB12 antigen is unusual - of hundreds of 
other mutants (zygotic and maternal) isolated by the Schnabel lab (p.c.
) and by me, none fails to express the 3NB12 antigen, as long as a 
complete cell number is made.  The absence of 3NB12-reactive cells may 
indicate that cells normally destined to become pharynx muscle 
actually express non-muscle fates, as opposed to simply failing to 
differentiate, since mutants such as pha-1, which don't complete 
pharynx differentiation, nonetheless produce 3NB12-reactive cells.
All of these defects indicate that the fates of certain cells in 
e2482 embryos are altered.  To track down specific cell fate defects, 
I have followed a number of lineages in developing e2482 embryos using 
a time-lapse video system, put together by John White, which records 
in multiple focal planes.  The lineages of hyp6 and hyp7 cells from 
ABarpaa, ABplap, and Ca, seam hypodermal cells H0L, H1L, V14L, and V6L,
and the 8 body muscles arising from Capa were wild-type.  However, 
the lineages normally engendering all of the ventral hypodermal Pn/n+1 
cells on the left side of one embryo produced cells whose morphology 
appeared non-hypodermal in that they were small round cells containing 
grainy looking nuclei with no nucleolus, and ended up on a region 
removed from the dorsal patch of hypodermis.  In some cases these 
apparently non-hypodermal cells underwent an extra division (in two 
different embryos in the case of P7/8L).  It is striking that the 
failure to produce a Pn/n+1 cell of hypodermal appearance, but not a 
seam cell, can occur at the very last normal division: although V3L 
and P7/8L are sisters, only the latter failed to become hypodermal and 
underwent a supernumerary division.  More lineages must be performed 
before it is possible to determine whether the Pn/n+1 cells are the 
only hypodermal cells that fail to be properly specified, and to 
assess the fates of the missing pharynx and body wall muscle cells.
The following additional defects of developing e2482 embryos were 
observed.  The earliest was a weak gastrulation defect (at ~28 cells) 
in most of the embryos observed: the two E cells, and their daughters, 
generally failed to enter the interior of the embryo to the same 
extent as control wild-type embryos.  This is one of the earliest 
demonstrated requirements for zygotic gene expression, and this defect 
has proven useful in identifying e2482 homozygotes at an early stage 
in development.  Later, an apparent defect in cell adhesion appears (
conspicuous after about 200 cells): the cells at the surface fail to 
become tightly packed, and the embryos take on the appearance of 
clusters of grapes.  It is not at present clear whether these defects 
are the cause of Lhc altered cell fates observed in e2482, or the 
manifestation of altered precursor cell fates.
What are the ultimate fates of the 'lost' hypodermal, muscle, and 
pharynx cells? The nuclei of the aberrant cells arising from the 
Pn/n+1 lineages look neuronal, and in the few cases in which these 
cells appeared at the surface of the embryos, tiny processes were 
sometimes seen to protrude from them.  It is therefore possible that 
e2482 is a neurogenic mutant, resulting in transformation of certain 
cells into neurons.  Electron microscopy of one fixed embryo, whose 
development was recorded by time-lapse, is forthcoming.
The other pre-elongation mutants may also have hypodermal lineage 
defects.  e2504 V embryos contain about 70 anti-desmosome-reactive 
hypodermal cells (instead of the 85 hypodermal cells in wild-type) and 
generally fail to differentiate the seam cell-specific antigen, 
although expression of pharynx myosin and body wall muscle appears 
normal.  In contrast, e2500 expresses all differentiation markers 
tested, and the few cursory hypodermal lineages examined were normal --
this mutant may be specifically defective in migration of hypodermal 
cells.  It will be illuminating to follow the lineages and migration 
of hypodermal cells in e2504 and other pre-elongation mutants that 
arise in the ZEL screen.