Molecular characterization and expression analysis of stanniocalcin-1 in turbot (Scophthalmus maximus).

Stanniocalcin-1 (STC-1) is an anti-hypercalcemic hormone that is produced by the corpuscles of Stannius (CS) in the teleost. The hormone is a homodimeric glycoprotein involved in the calcium and phosphate regulation in both teleost fish and mammals. In the present study, we have cloned and characterized a full-length cDNA of STC-1 from the turbot (Scophthalmus maximus) CS and examined its expression pattern in various tissues. The turbot STC cDNA (1,246 nucleotides) encodes a putative preprohormone of 248 amino acids (aa) with a signal peptide of 18 aa and a pro-sequence peptide of 14 aa followed by a mature protein of 216 aa. The deduced aa sequence of turbot STC-1 shows the highest sequence identity (86.3%) with that of European flounder STC-1 among the fish and mammalian STC-1. The aa sequence of turbot STC-1 showed moderate sequence identity with those of salmoniforms (chum salmon and rainbow trout), cypriniforms (zebrafish), ancient fish (gar and bowfin), and humans (43.7-66.9%), respectively, whereas it showed less identity with those of zebrafish and human STC-2 (23.8-25.4%). RT-PCR analysis revealed that the turbot STC-1 gene is expressed in the CS, pituitary, brain, kidney, liver, heart, muscle, and gonad. In a transient transfection of turbot STC-1 cDNA construct into human embryonic kidney 293T cells followed by Western blot analysis, a recombinant STC-1 secreted into a medium was detected as a single band of 28 kDa under reducing conditions and a 50 kDa protein under non-reducing conditions. Deglycosylation of the recombinant STC-1 led to a decrease in the molecular mass. Our data indicate that the turbot STC-1 gene is widely expressed and the secreted protein exists as a homodimeric glycoprotein.

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