Cloning, Characterization and Expression Analysis of a cDNA Encoding Granulin Gene in Nile Tilapia (Oreochromis niloticus)

Granulins are a group of highly conserved growth factors that have been described from variety of some organisms spanning the metazoan. It has been shown to have roles in multiple processes involved in cell growth, development inflammation, wound healing and tissue remodeling. In the present study, we isolated the full-length cDNA encoding Nile tilapia granulin gene by in silico cloning and 5iRapid Amplification Complementary DNA Ends (RACE) techniques. The complete cDNA sequence of this gene consisted of 1,270 bp containing an open reading frame (ORF) of 618 bp (206 amino acid residues), including 44 and 608 bp of 5i and 3i untranslated regions, respectively. Two conserved-cystein granulin motifs CX5CX5CCX8CCX6CCX5CCX5CX6C similar to those of vertebrates were clearly characterized. Sequences analysis revealed that Nile tilapia granulin was closely similar to Mozambique granulin I and II. Phylogenetic analysis strongly showed that Nile tilapia granulin was clustered in the tiny sister group of vertebrate granulin genes closed to Mozambique granulin. Expression analysis using RT-PCR revealed that low levels of granulin mRNAs were present in cells obtained from heart, head kidney, spleen and lower levels in liver, respectively. Quantitative real-time PCR revealed that injection fish with Streptococcus agalactiae could clearly upregulate the levels of granulin mRNAs in head kidney.