Our goal is to use the C. elegans pharynx as a model to understand the complexities of organogenesis. We have previously identified a sub-set of pharyngeal gland expressed Mucin-related phat genes, which are secreted by the glands into the pharyngeal lumen. The loss of PHAT secretion results in early larval arrest due to a severe pharyngeal feeding blockage. This is the case in mutants for
hlh-6, the bHLH transcription factor that directly activates the phat genes, and when we ablate the gland cells (Smit et al. 2008), alluding to a role for the glands in feeding and transport of bacteria. Each of the three gland sub-types secretes these proteins at a unique position along the pharyngeal lumen, suggesting that one of the functions of the gland sub types is to create a local concentration of PHAT proteins, completely coating the pharyngeal lumen.
From a genetic screen we have isolated a mutant,
glad-25, where the
g1a gland sub-types are missing, or displaced from the pharynx. The pharynx appears morphologically normal, and linearly related pharyngeal cells are correctly specified, suggesting this seems to be specific loss of the
g1a gland cells.
Coincidental with the loss or displacement of the
g1a gland cells and loss of PHAT-1 expression, is the presence of a feeding blockage initiating during the later larval stages and through adulthood. This blockage is more progressive and less severe when compared to the early larval arrest due to blockage in
hlh-6 mutants. Feeding defective
glad-25 animals grow slower, and are smaller. Previously we've had success rescuing feeding defects in
hlh-6 mutants using the viscous E. coli strain HB101. Feeding
glad-25 mutants HB101 was able to rescue the feeding defects, the small body size, and slow growth. This would suggest that loss of a gland sub-type results in a spatial disruption of secreted PHAT proteins, and subsequently a unique feeding defect.
We used array CGH to map the mutation between 7.5-9.5 Mb on chromosome IV, and performed whole genome sequencing identifying ~20 variants in the region that cause amino acid changes, with several of theses variants possibly causing a loss-of-function. We are currently testing theses candidates by cosmid rescue.