Biological functions of galectin 15 (lgals15) in the ovine uterus
Galectins are proteins with 15 known members found in nearly all living organisms. They share a conserved CRD that binds beta-galactoside sugars, and functions to cross-link glycoproteins as well as glycolipid receptors on the surface of cells to initiate biological responses. Functional studies on the extracellular and intracellular roles of galectins implicate them in cell adhesion, chemoattraction and migration as well as growth, differentiation and apoptosis. Therefore, studies were conducted to identify functional roles of galectin 15 (LGALS15) during the periimplantation period of pregnancy in the sheep. The first study was designed to develop and characterize primary ovine trophectoderm cell lines for the study of the biological functions of LGALS15. Once characterized, these cell lines were used to investigate the role of LGALS15 in trophectoderm gene expression, development, growth, and survival. Two primary trophectoderm cell lines (oTr1 and oTrF) were developed, and they had characteristics similar to in vivo conceptus trophectoderm relative to gene expression, morphology, and migration and proved suitable as an in vitro model to investigate functional roles of LGALS15. The second study investigated LGALS15 function in trophectoderm cell adhesion. A dose-dependent increase in oTr cell attachment to LGALS15 was found that could be inhibited by cyclic GRGDS, but not GRADS, peptides. Mutation of the LDVRGD integrin binding sequence of LGALS15 to LADRAD decreased its ability to promote oTr cell attachment, whereas mutation of the CRD had little effect. LGALS15 induced formation of robust focal adhesions in oTr cells that were abolished by mutation of the LDVRGD sequence. The third study tested the hypothesis that LGALS15 is a secreted regulator of trophectoderm development and gene expression, as well as growth, migration, and apoptosis of trophoblast. LGALS15 moderately increased cellular proliferation, partially inhibited staurosporine elicited apoptosis, stimulated migration that was dependent on Jun N-terminal kinase (JNK), and initiated differential gene expression of oTr cells. Collectively, these results support the hypothesis that LGALS15 has a biological role in the peri-implantation stage of early pregnancy in the ovine uterus and stimulates trophectoderm cell gene expression, migration and attachment via integrin binding and activation which are critical to blastocyst elongation and implantation.