Isolation and sequencing of swine carbonic anhydrase VI, an enzyme expressed in the swine kidney

Toshiho Nishita,Shino Kamoshida,Hideharu Ochiai,Kensuke Orito,Masaru Murakami,Kazuyoshi Arishima,Nobutune Ichihara,Juro Yatsu

doi:10.1186/1756-0500-7-116

Toshiho Nishita, Shino Kamoshida + Show 6 more

Open Access

https://doi.org/10.1186/1756-0500-7-116

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Abstract

BackgroundCarbonic anhydrase VI (CA-VI) is produced by the salivary gland and is secreted into the saliva. Although CA-VI is found in the epithelial cells of distal straight tubule of swine kidneys, the exact function of CA-VI in the kidneys remains unclear.ResultsCA-VI was located in the epithelial cells of distal straight tubule of swine kidneys.A full-length cDNA clone of CA-VI was generated from the swine parotid gland by reverse transcription polymerase chain reaction, using degenerate primers designed based on conserved regions of the same locus in human and bovine tissues.The cDNA sequence was 1348 base pairs long and was predicted to encode a 317 amino acid polypeptide with a putative signal peptide of 17 amino acids. The deduced amino acid sequence of mature CA-VI was most similar (77.4%) to that of human CA-VI. CA-VI expression was confirmed in both normal and nephritic kidneys, as well as parotid. As the primers used in this study spanned two exons, the influence of genomic DNA was not detected. The expression of CA-VI was demonstrated in both normal and nephritic kidneys, and mRNA of CA-VI in the normal kidneys which was the normalised to an endogenous β–actin was 0.098 ± 0.047, while it was significantly lower in the diseased kidneys (0.012 ± 0.007). The level of CA-VI mRNA in normal kidneys was 19-fold lower than that of the parotid gland (1.887).ConclusionsThe localisation of CA-VI indicates that it may play a specialised role in the kidney.

Highlights

Carbonic anhydrase VI (CA-VI) is produced by the salivary gland and is secreted into the saliva
CA-VI was located in the epithelial cells of distal straight tubule of swine kidneys
Expression levels of CA-VI mRNA were measured by qRT-PCR in FFPE samples of normal and diseased kidneys (Figure 4)

Summary

Introduction

Carbonic anhydrase VI (CA-VI) is produced by the salivary gland and is secreted into the saliva. Carbonic anhydrase (CA; EC 4.2.1.1) is a well-characterised enzyme that catalyses the reversible hydration of CO2 to form HCO3− and protons according to the following reaction: CO2 + H2O ↔ H2CO3 ↔ HCO3− + H+. The physiological function of carbonic anhydrase is to maintain the acid–base balance in various tissues and biological fluids [4]. The enzyme is localised in the serous acinar and demilune cells of the parotid and submandibular glands [7,10], from which it is secreted into saliva. Hooper et al [12] suggested that the unique oligosaccharide structures on bovine CA-VI might have an antibacterial function. Karhumaa et al [13] suggested that the glycoproteins on CA-VI confer multifunctionality on the enzyme

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