Date of Award
Ryan, Littlefield, Ph.D.
Striated muscles consist of contractile myofibrils that generate force. The repeating units (sarcomeres) of striated myofibrils contain thin & thick filaments composed of actin and myosin. In vertebrates, titin connects thin & thick filaments during myofibril assembly and produces passive tension. Invertebrate homologs of vertebrate titin are similar in structure molecularly. However, invertebrate titin homologs are smaller than vertebrate titin, and likely function differently within invertebrate muscles. Here, I used Ce-TTN-1, a titin homolog in the invertebrate roundworm C elegans. To visualize Ce-TTN-1 in C. elegans, I used Nested CRISPR gene editing to generate translational fusions of a red fluorescent protein at Nterm- TTN-1 or Mid- TTN-1. I then colocalized Nterm-wrmScarlet-TTN-1 and Mid-wrmScarlet-TTN-1 proteins with DEB-1-GFP. By confocal microscopy, Ce-TTN-1 appeared as narrow striations within BWM while DEB-1-GFP was punctate. Statistical analysis done with Google Colaboratory showed distances between and the locations of Nterm-wrmScarlet-TTN-1 and Mid-wrmScarlet-TTN-1 were indistinguishable from each other. Using software called JACOP, Nterm-wrmScarlet-TTN-1 and Mid-wrmScarlet-TTN-1 crossed with DEB-1-GFP showed some colocalization but showed no evidence of localization of wrmSclt-TTN-1 to the DBs, indicating that Ce-TTN-1 may be unstretched.
Prince, Gaberielle S., "Location and Conformation of Titin in Caenorhabditis elegans Muscle" (2022). Theses and Dissertations. 81.