Research Mentor and Department
Douglas Chalker, PhD, Department of Biology
Ciliary function is known to play an important role in many human conditions, including chronic sinus and pulmonary diseases and problems with infertility. Cilia are cytoskeletal structures that protrude from the cell body to facilitate movement. Ciliary structure is conserved throughout eukaryotes, from unicellular to multicellular organisms, including humans. A clear understanding the proteins that compose cilia and how they interact with one another will increase our knowledge about important cytoskeletal elements. Because cilia are difficult to study in multicellular organisms, the unicellular eukaryote Tetrahymena thermophila serves as a useful model for the study of cytoskeletal genes, due to their robust and longitudinally organized cilia. Here, we examine two novel kinetodesmal fiber proteins in T. thermophila, BBC29 and BBC39. Kinetodesmal fibers stabilize and organize cytoskeletal elements to ensure that all the cilia beat together and in the same orientation to maximize the effects of their beating. Genes BBC29 and BBC39 were isolated using PCR amplification, cloned into pICY-gtw vectors, and transformed into T. thermophila. The pICY vector included an inducible 3’ MTT cadmium promoter and a 5’ yellow fluorescent protein tag to allow visualization of the proteins’ location in live cells under UV light. Both genes localized to the kinetodesmal fiber structures, indicating their role to help stabilize and coordinate ciliary movement. We examined the interaction between the two proteins using co-immunoprecipitation to determine if the two proteins are dependent on each other for proper function. Our results demonstrate the complex interactions that occur between cytoskeletal proteins, such as those in the kinetodesmal fibers of cilia, and illuminate the need to further understand the relationship among the many proteins that allow for proper ciliary function. Because cytoskeletal proteins are often highly conserved between species, results gleaned through study with T. thermophila may be extended to elucidate the biochemical basis of human ciliary diseases.