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Senior Theses 2007
Elise Coffey (2007). Consequences of RNA Interference of the muscle gene Titin in the development and muscle function of Caenorhabditis elegans
Faculty Advisor: Denise Flaherty
Caenorhabditis elegans is often used as a model organism for studies of muscle function. Titin, the enormous striated muscle protein in C. elegans was examined in this study through the process of RNA interference (RNAi). Both feeding and microinjection strategies were employed for this study. Double-stranded RNA targeting a unique coiled-coil motif region of titin (ttn-cc) was used to determine if this region of titin was functionally or structurally important to the developing, or adult, muscle sarcomeres. It was hypothesized that this coiled-coil region may be important in titin-proteinbinding interactions, or that it may be another functional spring portion of the titin molecule. To ascertain the consequences of titin RNA interference, affected and unaffected progeny were evaluated with motility assays and polarized light microscopy.
N2 (wild type) and the two titin mutant strains ttn-l (gk135) and ttn-l(okl018) were examined for effects. The most substantial effect on the muscle myofilament lattice was seen in ttn-l(okl018) with a "patchy" polarized light phenotype showing a broadening of the myosin muscle A-band thick filaments and disorganization of the actin filament containing thin I-band filaments. These data suggest that when several titin isoforms are compromised the structure of the muscle sarcomere is adversely affected. Beyond a polarized light phenotype, all RNAi affected animals (wild type or titin mutant) had slower motility by the liquid beat assay that was statistically significant (student two- tailed t-test). As new data in the field of titin molecular biology examine titin's many structural protein motifs, this study is the first to demonstrate a functional consequence of RNA interference of the titin gene.