RNAi characterization of a putatuve collagen IV cell surface receptor.

Collagen IV is a major component of the Basement membrane, a thin sheet of extra cellular matrix. In vertebrates collagen IV is recognized by cell surface receptors and interacts with other basement membrane components. The cell surface receptor that recognizes collagen IV in C. elegans is unknown. The goal of this research is to identify a potential cell surface receptor that recognizes collagen IV by the combination of bioinformatics with RNAi techniques in C. elegans.

The C. elegans sequenced genome has allowed for advanced genetic analysis. The previously established RNAi technique has proved useful in genetic studies, RNAi acts at the posttranscriptional level by targeting the mRNA for degradation resulting in reduced gene expression. In this way RNAi has been a powerful tool for determining the function of specific genes. Microinjection was utilized to introduce dsRNA into the C. elegans syncytial gonad to expose early embryos to dsRNA. Embryos are visually analyzed over several weeks to determine an RNAi effect. To date preliminary dsRNA has been generated from cDNA clones. The dsRNA control gene fragments have been used in RNAi studies and a twitcher phenocopy has been achieved, dsRNA for putative collagen IV cell surface receptors (ESTs) are in the process of being generated and will be utilized in RNAi studies. An RNAi phenocopy for the collagen IV cell surface receptor is expected to be embryonic lethal. Digital microscopy will be used to analyze and record results.

Other putative collagen IV cell surface receptors will be identified using additional C. elegans database searches for other potential genes that may encode for a receptor that recognizes collagen IV. These searches will expand our library of potential genes.

* Burcks, S. and C.M. Witkowski. Department of Biomedical Sciences, Southwest Missouri State University.