CAREER:Evolution of a vertebrate-specific plug-and-jack interaction

Montana State University, Bozeman MT

Investigators

Abstract

The C-terminal core of the TATA-binding protein, TBP, is one of nature's most conserved protein domains; however the TBP N-terminal region varies greatly among phyla. A novel domain arose at the pre-vertebrate/vertebrate transition and has been conserved along all subsequent lineages. The investigator hypothesizes that the N terminus functions as a signaling port by which the basal transcription machinery receives specific regulatory signals via protein:protein interactions, reminiscent of the diverse task-specific 'plug-and-jack' communication connections on the back of computers. Variation between N termini likely represents commitment to phylum-specific gene-regulation. This project investigates co-evolution of the vertebrate TBP N terminus with its interaction partners. The work will begin with two-hybrid protein:protein interaction screens in representatives of advanced (mouse) and primitive (hagfish) vertebrates. Putative interactors are being confirmed first biochemically and then in live cells via co-transfection assays. Various cell lines from different phyla, as well as mammalian cells lacking the vertebrate N terminus, are being used to represent major variations in metazoan TBP N termini. Lastly, functional evolution of the vertebrate N terminus is being investigated by precisely replacing the N terminus protein-coding region of the mouse tbp gene with those from hagfish (the lowest extant vertebrate) or amphioxus (an advanced pre-vertebrate chordate). This project is expected to provide a paradigm for co-evolution of protein:protein interactions. The mammalian version of the vertebrate N terminus plays an important, yet only partially understood, role in maintaining pregnancy. These studies on the pre-vertebrate and early vertebrate functions of this domain, and of the molecular evolutionary history of this domain along vertebrate lineages, will provide the basic understanding to help the scientific community to understand such phenomena. This CAREER project integrates advanced research on evolution of the transcription machinery with hands-on research experience for students at many levels. The underlying hypothesis is that the vertebrate N terminus co-evolved with interactor proteins as 'plug-and-jack' interactions, and that these diverged to assimilate novel functions during vertebrate evolution. This provides an interesting and novel topic that is being integrated into an advanced undergraduate course taught by the investigator. Moreover, most of the experiments and approaches used in this study are compatible with providing research training to relatively inexperienced students. Therefore, the students in the class also participate on a small aspect of this CAREER project each year. This serves both to demonstrate the type of empirical investigation that underlies molecular evolutionary research, and helps make the lecture materials more tangible. Despite the relative technical simplicity of most aspects of this project, the hypotheses are novel and important, the training value is high, and the scientific importance of the evolutionary processes that will be investigated is great.