Color Code for “S1”, a sma-2;unc-30 double mutant

Nov. 14, 2012

DHH interpretation

This mutant hermaphrodite adult was prepared by Nichol Thomson for Sydney Brenner to assess the state of the synaptic wiring in the retrovesicular ganglion (RVG) and anterior ventral nerve cord. Relatively few neurons were traced on the archival prints, and we do not have a color code from the MRC files. It was originally called allele “S1”, consisting of a genetic cross between sma-2 (e502) and unc-30 (e318).

Why this double mutant was created and examined by TEM in serial sections is not entirely clear. Given what we know today about these two genes (see below), one might expect that in the RVG and ventral cord, the unc-30 phenotype might be more easily scored than any effect of sma-2. At the time of this reconstruction, probably in the early 1970s, nothing was known about the identity of these two genes beyond their general effect on body shape and/or motor coordination. Perhaps it was considered advantageous to cross these two genes to produce a shorter animal that would require fewer thin sections to explore the RVG?

The animal is sectioned transversely. A system of blue and blue green markers were used to trace some sets of axons, mostly using an X or O to indicate the locales of several motor axons where they approach the muscle plate. In addition, many axon shapes were carefully followed on tracing paper overlays to follow the exact contours of individual neurons within the main fascicle, using another schema to indicate separate axon identities (not shown). We believe that the annotations and tracings were done by Brenner, as they do not match the style of other MRC staff.

In our collection of mutant animals from the MRC, there are several more annotated print sets from unc-30 single mutants. A typical phenotype for unc-30 nerve cord cells was as follows:

A few motoneuron cell bodies seem to be slightly mispositioned along the A/P axis (DD2 is about 3 cell body positions too posterior, as was also noted in one of the adults from 1984).  The most striking defects are the incorrect NMJs formed by VDs and DDs in this animal.  VDs retain NMJs to dorsal muscles (as in normal L1s) and have few or none to ventral muscles (where there ought to many).  DDs have too few NMJs to dorsal muscles (should have many more), and apparently none to ventral muscles (which is normal).

From WormBase:

sma-2 encodes one of three C. elegans Smad proteins; during development, SMA-2 functions in a TGF-beta signaling pathway to regulate body size and specification of sensory structures in the male tail. sma-2, through this pathway, also regulates reproductive aging. sma-2 is expressed in pharynx, intestine and hypodermis.

unc-30 encodes a homeodomain-containing protein that is orthologous to the Pitx family of homeodomain transcription factors. During development, UNC-30 controls the terminal differentiation of all 19 type D GABA-ergic motor neurons by directly regulating the expression of UNC-25/GAD and UNC-47/VGAT, which regulate GABA formation and secretion, respectively. UNC-30 is expressed most strongly in type D motor neurons during early L1 and L2 larval stages.