Assistant Professor Midwestern University Glendale, Arizona, United States
David Carroll (Midwestern University)| Lauren Bates (Florida Institute of Technology)| Emily Wiseman (Florida Institute of Technology)
Multiple events occur between the sperm and egg membranes at fertilization, including sperm-egg binding, species-specific discrimination, membrane fusion, and egg activation. To date, only a handful of egg membrane proteins have been identified, and none of these are known to be involved in the egg activation pathway at fertilization. An excellent model organism to address how these events take place is the Pacific bat star, Patiria miniata, due to the quantity of synchronized oocytes that are large in size, nearly transparent, and amenable to many different techniques. To address this lack of knowledge regarding the egg cell surface, three databases were developed to explore the membrane proteome of the mature sea star egg. These databases utilized the first sea star mature egg transcriptome for the sea star, which has 96,723 unigenes and is available on NCBI (BioProject ID: 389668). The first database is a prediction of all mature egg transcripts that have putative transmembrane alpha-helical sequences using two different software programs: Phobius and TMHMM, which use different methods for prediction. After combining results from both programs, 15,501 transcripts were predicted to encode a transmembrane domain. The majority possessed a single transmembrane domain (9744); however, many were predicted to have multiple transmembrane domains. The second database categorized proteins into five major signaling families, each of which have been hypothesized to play a role in gamete interaction in different species. This database identified unique representatives of 26 Tetraspannins, 28 Receptor tyrosine kinases, 8 A Disintegrin and Metalloproteases (ADAMS), 140 G-protein coupled receptors, and 9 Integrins. The final database represents the membrane proteins currently present in the mature egg as identified by mass spectrometry. In the mature egg membrane preparation, 951 proteins were identified by mass spectrometry. Of these, 167 matched with the transcripts identified using the topology prediction software (Database 1) and 272 were matched with transcripts belonging to the five families of interest (Database 2). Interestingly, overlap between all three databases identified five proteins that are present in the unfertilized egg and belong to one of the signaling families predicted to be involved at the surface. These proteins were an integrin alpha and beta-like isoform, and three different tetraspannins. A major goal of this work is to provide targets for future functional studies of receptors involved in activation of fertilization at the egg surface.