Identification of Unique Regulatory and Coding Regions in Mouse Myosin V Introns: A Comparison of Proteomics- and Genomics-Based Methods

Abstract

Myosins are motor proteins that contribute to many intracellular biological processes in eukaryotic cells, including a major role in transporting various molecules in nerve cells. Cargos delivered by myosin motors contribute to synapse development, a process important in human learning and memory. Caenorhabditis elegans uses alternative splicing to code for a motor-less version of myosin V (HUM-2C) that interacts with an RNA binding protein called VIG-1. The objective of this research was to find a homologous version of HUM-2C in mammals. Plasminogen Activator Inhibitor RNABinding Protein (PAI-RBP1), a mammalian homologue to VIG-1, was used as a yeast two-hybrid bait to identify an interacting mouse myosin V. A yeast strain carrying PAIRBP1 was mated to a yeast strain carrying the mouse brain transcriptome library. The mated cells were screened for activation of reporter genes and a protein interaction with PAI-RBP1. I sequenced 65 randomly selected colonies showing positive interaction. Although no prey gene corresponded to a myosin, I identified some common binding sequences using this method. Multiple sequence alignments were utilized to identify conserved regulatory and coding sequences located in myosin V genes of five mammals, including the common house mouse (Mus musculus). Conserved sequence and transcription factor binding sites were identified within one intron of MYO5A. The intronic region may act as a 5’ UTR in the expression of a truncated Myo5a protein in mammals. Identifying this version of myosin V in mammalian brain tissue may provide a new target in studying processes that affect neuronal plasticity and learning in humans.