Integrin Distribution in Collagen Iv Mutants and Use of Rna Interference to Identify a Collagen Iv Receptor in C. Elegans

Umme Rumana

Date of Graduation

Spring 2004

Degree

Master of Science in Cell and Molecular Biology

Department

Biomedical Sciences

Committee Chair

Colette Witkowski

Abstract

Perlecan, a basement membrane protein, was found to be significantly decreased in collagen IV mutants. To better understand the role of collagen IV in perlecan-integrin dependent sarcomere organization, an extrachromosomal array containing a functional ßpat-3::GFP fusion gene was mated using genetic crosses into a collagen IV (emb-9) mutant background. Generation of a stable transgenic strain was not successful, but GFP distribution in M-lines and dense bodies was analyzed on a potential crossed animal to determine integrin distribution directly and ligand perlecan organization indirectly in live, mutant embryos. The distribution of integrin as assessed by the GFP tag in the mutant embryos demonstrated several defects including disorganized integrins, gaps in M-lines and dense bodies, and potential cellular accumulation of integrins. These data suggest that collagen IV is possibly critical for perlecan distribution and organization and in maintaining the perlecan-integrin dependent sarcomere organization. The second part of this research attempted to identify a potential collagen IV receptor, a putative protein, M01E10.2, based on its homology to human integrin a1, a2 and aM subunits and signature consensus sequences, by using the RNA interference (RNAi) technique. Microinjection of dsRNA targeting the putative receptor gene was hypothesized to phenocpy the collagen IV null phenotype. Observation of dumpy animals suggests that the putative protein is possibly involved in body shape and size regulation. Absence of a significant percent of embryonic lethality as hypothesized from the M01E10.2 gene silencing suggests that this hypothetical protein is not a collagen IV receptor.

Keywords

extracellular matrix, collagen IV, collagen IV receptor, integrin, RNAi

Subject Categories

Medical Molecular Biology

Copyright

© Umme Rumana

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Dissertation/Thesis

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