One major focus in our lab is to elucidate the role of
unc-119 in nervous system development. Through the course of this study we became interested in the predicted gene
pdl-1 ( p hosphodiesterase 6 d elta l ike), which shows weak similarity to
unc-119 . A
pdl-1 promoter::GFP fusion shows pan-neuronal expression beginning early in embryogenesis and maintained throughout the life of the worm. This suggests that
pdl-1 , like
unc-119 , is involved in nervous system development or function so we have decided to characterize this gene further. Additionally,
pdl-1 is highly conserved exhibiting 85% similarity to the mammalian protein PDE6D (rod cGMP phosphodiesterase delta). PDE6D is widely expressed in mammalian tissue with enrichment in the retina and appears to play a regulatory role through its interactions with other proteins. Specifically, PDE6D binding affects the solubility and/or nucleotide binding properties of the proteins with which it interacts. These include rod cGMP phosphodiesterase (PDE6), Arl3, and Rab13. Interestingly, C. elegans PDL-1 is able to bind and solubilize mammalian PDE indistinguishably from PDE6D 1 , suggesting that the function of these proteins has been conserved. To determine the function of
pdl-1 we are attempting to determine a knockout phenotype as well as studying its expression and protein interactions. i. Knockout Phenotype Deletion screening is ongoing through the C. elegans Knockout Consortium. We have made several attempts at RNAi, but so far we have been unable to knock out expression of
pdl-1 in the nervous system. We have also looked for uncloned mutations in the region of
pdl-1 which seem like possible candidates. Neither
dyf-13 nor
egl-42 is rescued by
pdl-1 . ii. Expression Northern analysis has confirmed that
pdl-1 is expressed as a single transcript of predicted size. Transcriptional and translational GFP fusions show expression throughout the nervous system with additional expression in pharyngeal and vulval muscle. As well, we are in the process of purifying antibodies which will allow us to determine the subcellular localization of PDL-1. iii. Protein Interactions We have performed a yeast 2-hybrid screen using PDL-1 as bait. We have identified six putative positives, two of which show interaction with mammalian PDE6D. Further characterization of these interactions, including domain analysis and CoIp, is underway. 1. Li and Baehr (1998). FEBS Letters. 440: 454-457.