PROJECT SUMMARY/ABSTRACT
This proposal describes a five-year mentored research career development project focused on the in vivo
efficacy of and mechanisms underlying a novel restriction factor, retroCHMP3. Host restriction factors evolve to
protect genomes from threats from viruses and transposable elements. The cellular Endosomal Sorting
Complexes Required for Transport (ESCRT) pathway is targeted by many viruses for budding and is implicated
in retrotransposition, yet is essential for host membrane remodeling, including during cytokinesis. Host evolution
to restrict viruses or retrotransposons must therefore preserve key cellular functions, with little room for error.
retroCHMP3 is a retrotransposed, truncated and mutated copy of the essential ESCRT gene chmp3 that
dominantly inhibits virus budding and LINE-1 retrotransposition in cells. Remarkably, retroCHMP3 has been
detoxified so that it does not block cytokinesis. retrochmp3 arose and evolved independently in mice and new
world monkeys, suggesting that it confers selective advantages. This proposal tests the hypothesis that
retroCHMP3 is a broad-acting restriction factor that evolved for mammalian defense. The experiments in Aim 1
will elucidate 1) the in vivo efficacy of retroCHMP3 against LINE-1 retrotransposition with a mouse model that
reports new LINE-1 insertions in the presence or absence of retroCHMP3, and 2) the mechanism by which LINE-
1 activity requires the ESCRT pathway with a combination of cell culture and biochemical assays. Aim 2
describes the generation of a knock-in mouse which will reversibly and inducibly express retroCHMP3 and will
be used to test 1) the extent of L1 retrotransposition restriction in vivo, 2) the in vivo efficacy of retroCHMP3
against viral infection, and 3) the degree of toxicity from in vivo expression of retroCHMP3. The two most
important impacts are: 1) understanding the functionality of a new viral and retrotransposition restriction factor
in vivo has implications for evolutionary biology, development of anti-viral interventions, and understanding the
balance between host cell toxicity and pathogen restriction, and 2) the role of the ESCRT pathway in LINE-1
biology may reveal new functions for the ESCRT pathway, since known ESCRT functions have been shown to
be non-essential for LINE-1 retrotransposition. The investigator is a postdoctoral researcher at the University of
Utah and has enlisted two strong mentors with expertise in the ESCRT pathway, virus budding, evolutionary
biology, and postdoc mentoring. The candidate is a veterinary scientist who proposes to use this mentored
training period to augment her expertise in infectious disease pathogenesis with virology, host-pathogen
coevolution, and mechanistic cell biology. The scientific proposal is complemented by a thorough training plan
which emphasizes proposal writing, idea generation, management and leadership, in addition to the
development of research skills in a new field. The short-term goals and specific training opportunities described
in the training plan contribute to the candidate’s long-term goal of leading an independent research program in
infectious disease.