MT Tech

Tripartite motif (TRIM) proteins are E3 ubiquitin ligases that regulate cellular processes, such as interferon responses and autophagy.  These activities influence antiviral immunity and are potential therapeutic targets.  Our hypothesis is that TRIM proteins interact with and post-translationally modify proteins that influence antiviral processes.  To test this hypothesis, we will utilize teams of undergraduate researchers to identify TRIM-binding proteins, characterize any post-translational modifications to the targeted proteins, and weave these observations together with our aims looking at TRIM protein influence on interferon responses, autophagy, and viral replication.  While the 82 TRIM protein family members are named after their conserved “RING/B-box/coiled-coil domain” tripartite motif near their N terminus, the C terminus is less conserved and is often important for interactions with TRIM-binding proteins.  Through alternative splicing of mRNA, most TRIM proteins can have several different isoforms, and these can lack different domains within the tripartite motif or the variable region. 

Our knowledge that TRIM proteins influence interferon signaling and autophagy pathways comes from screens of TRIM protein panels using the longest isoforms possible for each TRIM protein.  There is precedent for shorter TRIM protein isoforms to interfere with the long TRIM protein isoforms.  Thus, there is a knowledge gap in our understanding of TRIM proteins on two levels: the TRIM protein panel screens produced many “hits” that have not been further characterized, and the impact of shorter TRIM protein isoforms are largely unknown.  The experimental design in this research proposal is straight-forward and intended to be formative for undergraduates seeking medical- and research-based careers.

Specific Aims

  • Aim 1: Identify protein targets that are post-translationally modified by TRIM proteins.  Our working hypothesis is that proteins bound by TRIM proteins will be post-translationally modified.  The RING domain within the tripartite motif will be responsible for the enzymatic activity and the type of modification (mono- or polyubiquitination, SUMOylation, etc.) made to the target protein. Protein-protein interaction assays will be the foundation for this aim.
  • Aim 2: Pinpoint the step or steps of antiviral processes acted upon by each TRIM protein.  Large-scale screens have indicated that most TRIM proteins influence interferon signaling and autophagy pathways.  Our working hypothesis for this aim is that these pathways could be dissected to better characterize the point at which TRIM proteins play a role.  Diverse assays for analyzing gene expression, protein modification and localization, as well as fluorescent-based autophagy analysis will be used by students to complete this aim.
  • Aim 3: Assess TRIM protein function in the context of specific viral infections.  In addition to influencing cellular proteins and processes, there are numerous examples of specific TRIM proteins influencing specific viruses.  The working hypothesis for this aim is that screening the effect of TRIM protein expression levels on a panel of viruses is the most direct method for detecting these virus-specific TRIM roles.

Primary Contact

Joel Graff jgraff@mtech.edu