A newly identified Galectin-8 from sea urchin embryos

Galectins are carbohydrate binding proteins that specifically bind beta-galactoside derivatives (Jun Hirabayashi et al, 2002; Karin Pfeifer et al, 1993; Liu et al, 2011). The members of the galectin super-family interacting with cell-surface glycoconjugates regulate diverse cellular events, including signaling pathways, apoptosis, innate immune and inflammatory responses (Liu and Rabinovich 2005). Some galectins involved in biomineralization are found in mammalian osteoblasts and osteocytes (Tanikawa et al. 2010). In sponges, the matrix guided formation of silicatein-mediated silica spicules is strongly increased when associated with a galectin (Schröder et al, 2006; Müller et al 2009). This study was undertaken to isolate and characterize galectin cDNAs from the Paracentrotus lividus sea urchin embryo. By RT-PCR and 3' RACE we amplified a putative galectin family member and cloned it in pGEM-T-Easy vector. The 1309nt clone includes a 933nt coding sequence, encoding a 34.7kDa protein, containing two tandem carbohydrate-recognition domains. The sequence homology, obtained by Blast analysis, suggested Pl-galectin as a novel member of the Galectin-8 family. We characterized the deduced aminoacid sequence by in silico analysis and modeling based on the high structural similarity of the N-Terminal domain of Human Galectin-8 crystal structure. The expression levels of the Pl-galectin-8 mRNAs were monitored during the development of the P. lividus embryo by whole mount in situ hybridization and comparative Q-PCR. In order to perform functional assays in view of future biomedical applications we cloned the CDS in the pCOLD-TF expression vector and isolated the recombinant Pl-Galectin protein expressed in E. coli (BL21.AI). As potential calcium binding sites were identified by similarity on the tridimensional model, the calcium binding activity was assayed using the isolated recombinant protein. This work was supported by the BIOMINTEC Project (European Commission PITN-GA -2008-215507 grant).