Molecular characterization and expression analysis of regucalcin in disk abalone (Haliotis discus discus): intramuscular calcium administration stimulates the regucalcin mRNA expression.
Regucalcin is a novel calcium (Ca(2+)) binding protein and it has been demonstrated to play a multifunctional role in many organisms. Here, we report the molecular cloning of invertebrate regucalcin cDNA from disk abalone Haliotis discus discus. The full length cDNA showed 1321 bp of nucleotides with a polyadenylated sequence (AATAAA). Abalone regucalcin (HdReg) open reading frame (ORF) consists of 918 nucleotides encoding 305 amino acids (aa). Estimated molecular mass was 33 kDa and predicted isoelectric point (pI) was 4.9. The HdReg aa sequence did not contain the EF-hand motif as a Ca(2+) binding domain, suggesting a novel class of Ca(2+) binding protein. Moreover, it showed 45% identity to chicken and zebrafish, and 44% to rat and mouse regucalcin in deduced aa level. The tissue expression analysis of HdReg mRNA was investigated by RT-PCR and it was expressed in all the tissues tested such as gill, mantle, digestive tract, and abductor muscle. Semi-quantitative RT-PCR results showed that an intramuscular administration of calcium chloride (CaCl(2)) (0.5 mg CaCl(2)/g of abalone) could significantly induce regucalcin mRNA in abductor muscle after 30 min of administration and reached maximum after 1 h. Subsequently, the expression level was decreased after 2 h. This indicates that the expression of regucalcin mRNA is constitutive, and specifically up regulated in abalone abductor muscle by Ca(2+) administration.
Nikapitiya
Chamilani
C
Department of Marine Biotechnology, Cheju National University, 66 Jejudaehakno, Jeju 690-756, Republic of Korea.
De Zoysa
Mahanama
M
Kang
Hyun-Sil
HS
Oh
Chulhong
C
Whang
Ilson
I
Lee
Jehee
J
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000008:administration & dosage* / Q000494:pharmacology*; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D049851:Gastropoda / Q000187:drug effects* / Q000378:metabolism*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects*; D007273:Injections, Intramuscular; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000187:drug effects; D010802:Phylogeny; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein
D002135:Calcium-Binding Proteins; D012333:RNA, Messenger; D002118:Calcium