Worm Breeder's Gazette 14(4): 33 (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.
Dept. Neuroscience, Albert Einstein College of Medicine Bronx, NY 10461
Immunofluorescence studies using MH27 antibody have shown a wide
variety of tissues to be labelled in patterns which define apical
cell borders in C. elegans. Disappearance of MH27 binding during
development often marks cell fusions in hypodermis and in the male
tail (Podbilewicz and White, 1994; Fitch and Emmons, 1995). We
have used an immunoEM method to learn the nature of membrane
structures marked by MH27. A post-embedding technique was used to
apply MH27 Ab to thin sectioned worms, followed by a gold-linked
secondary Ab (for methods, see Selkirk et al., 1991; Hall, 1995).
Electron microscopy reveals that MH27 binds only to a few types of
cell junctions.
By immunoEM, MH27 can be seen to bind to zonula adherens
junctions in intestine, at hypodermal/seam cell borders, and at
pharyngeal muscle/support cell borders. In adult pharyngeal
muscle, small longitudinal stripes of adherens "junction" are also
retained on the apical surface where pairs of muscle cells had
fused to become syncytial. Thus MH27 binding can sometimes mark
the vestiges of an old cell border, long after cell fusion.
Spermathecal cells are held closely together by two types of
"septate" junction, which together cover almost all of their
lateral borders. Extensive, dark-staining apical junctions, which
look vaguely adherens-like, are not labelled by MH27. In more
basal regions, MH27 binds heavily to another class of extensive
sinuous junctions, which lack any dramatic staining
characteristics. Neither class of junction has osmiophilic septa
visible in ordinary sections. The apical junctions are rather
closely apposed and show periodic striping of the thick cytoplasmic
density bordering the junctions. The basal, sinuous junctions show
an even, widened extracellular space between adjacent plasma
membranes, with faint periodic striations crossing the
extracellular space between the cells. There is no cytoplasmic
density associated with the sinuous junctions. Lanthanum
infiltration has been used to negatively stain extracellular septa
in the spermathecal junctions. The apical junctions have long,
wavy septa; they may comprise a novel class of septate junction,
characterized by the thick densely-staining coat on the cytoplasmic
face of each cell. The sinuous junctions have not been well
infiltrated with lanthanum yet. In one instance we did observe
short septa at regular intervals, spanning the extracellular space,
similar to those expected in "smooth septate junctions" as
described in other invertebrate species. These two classes of
septate junction may resist the stresses which would otherwise tear
apart the spermatheca during the passage of an oocyte.
Our current technique does not permit resolution of the exact
locus of MH27 antigen within the adherens junction or the smooth
septate junction. It could be cytoplasmic, extracellular, or
within the plasma membrane. In glancing sections of both types of
junction, it is clear the antigen is found evenly along the entire
face of the cell-cell appositions, possibly at a fixed distance in
relation to the plasma membrane. High resolution studies would
require a more direct labelling method, such as attaching a small
gold tag directly to a Fab fragment of the MH27 antibody. This
would bring the gold tag into closer proximity to the true
antigenic site within a thin section.
We thank Jim Waddle and Ross Francis (Washington U.) for
generously supplying MH27 antibody.
Fitch and Emmons (1995) Dev. Biol. 170: 564-582.
Hall (1995) In C. elegans: Modern Biological Analysis of an
Organism. H.F. Epstein and D.C. Shakes (eds.). Academic Press, New
York, pp. 395-436.
Podbilewicz and White (1994) Dev. Biol. 161: 408-424.
Selkirk et al. (1991) J. Biol. Chem. 266: 11002-11008.