EOR-2 is an obligate binding partner of the BTB-zinc finger protein and putative ERK target EOR-1.

摘要

BTB-zinc finger transcription factors play many important roles in metazoan development. The BTB domain of these proteins is critical for self-association and for recruiting cofactors to promoters of target genes. Identification of these binding partners is important for understanding how BTB-zinc finger proteins influence transcription. Caenorhabditis elegans serves as a good model to study BTB-zinc finger proteins as only two of these proteins are encoded in the worm. The BTB-zinc finger protein EOR-1 was previously identified in our lab for its positive role in mediating the Ras/ERK signaling pathway. Based on genetic arguments, the novel but conserved protein EOR-2 is a predicted partner of EOR-1. For my thesis, I sought to characterize the relationship between EOR-1 and EOR-2 and to better understand how both proteins function to mediate Ras/ERIC signaling.;Here, we show that EOR-2 is an obligate binding partner of EOR-1. A point mutation (L81F) in the EOR-1 BTB domain reduces EOR-2 binding and eliminates all detectable biological function. This point mutation lies near the predicted charged pocket region of the EOR-1 BTB dimer, a region that, in other BTB-zinc finger proteins, has been proposed to interact with corepressors and coactivators. In addition, we demonstrate that the conserved CHHC zinc finger motif of EOR-2 is required for binding to EOR-1 and for EOR-2 function in vivo. Furthermore, we show that EOR-2 can bind to the human BTB-zinc finger protein PLZF. We propose that EOR-2 defines a new family of binding partners for BIB-zinc finger transcription factors that may have conserved roles in other organisms.;We also show that EOR-1 may be a direct target of MPK-1 ERK in vivo. EOR-1 functions in the same cells in which Ras is active, and EOR-1 is a robust substrate of ERK in vitro. The EOR-1 BIB point mutation (L81F) that reduces EOR-2 binding also disrupts an ERIC docking site in EOR-1, resulting in a reduction of ERK phosphorylation. However, phosphorylation of EOR-1 is not absolutely required for its interaction with EOR-2. The majority of phosphorylation maps to the N-terminus of EOR-1 and mutation of these sites reduces EOR-1 function in vivo. Our future studies will address how EOR-1 regulates transcription of target genes and how EOR-2 binding and MPK-1 ERK phosphorylation affects this regulation.