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.
Shin
Jihye
J
Division of Marine Molecular Biotechnology, Faculty of Marine Bioscience and Technology, Kangnung National University, Gangneung 210-702, Republic of Korea.
Oh
Dahlkyun
D
Sohn
Young Chang
YC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D005412:Flatfishes / Q000235:genetics* / Q000378:metabolism; D020869:Gene Expression Profiling; D006023:Glycoproteins / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D010802:Phylogeny; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D014018:Tissue Distribution / Q000235:genetics
D006023:Glycoproteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; C051795:teleocalcin