Kari Kaunisto

Expression of the Membrane-associated Carbonic Anhydrase Isozyme XII in the Human Kidney and Renal Tumors

Carbonic anhydrase isozyme XII (CA XII) is a novel membrane-associated protein with a potential role in von Hippel–Lindau carcinogenesis. Although Northern blotting has revealed positive signal for CA XII in normal human kidney, this is the first study to demonstrate its cellular and subcellular localization along the human nephron and collecting duct. Immunohistochemistry with a polyclonal antibody (PAb) raised against truncated CA XII revealed distinct staining in the basolateral plasma membrane of the epithelial cells in the thick ascending limb of Henle and distal convoluted tubules, and in the principal cells of the collecting ducts. A weak basolateral signal was also detected in the epithelium of the proximal convoluted tubules. In addition to the normal kidney specimens, this immunohistochemical study included 31 renal tumors. CA XII showed moderate or strong plasma membrane-associated expression in most oncocytomas and clear-cell carcinomas. The segmental, cellular, and subcellular distribution of CA XII along the human nephron and collecting duct suggests that it may be one of the key enzymes involved in normal renal physiology, particularly in the regulation of water homeostasis. High expression of CA XII in some renal carcinomas may contribute to its role in von Hippel–Lindau carcinogenesis.

The identification of secreted carbonic anhydrase VI as a constitutive glycoprotein of human and rat milk

In addition to essential nutrients, human milk contains several classes of bioactive factors such as enzymes, hormones, and growth factors, many of which are implicated in infantile growth and development. Secretory carbonic anhydrase isoenzyme VI (CA VI) has been identified earlier as an essential component of mammalian saliva, and we demonstrate here by using biochemical and immunohistochemical techniques that it is also an elementary component of milk. The 42-kDa glycopolypeptide purified from human milk in CA inhibitor affinity chromatography shared 100% homology with salivary CA VI in the protein sequence analysis (40% coverage), and its digestion with PNGase F resulted in a polypeptide backbone similar in size to salivary CA VI. Quantification of CA VI in milk by using a time-resolved immunofluorometric assay revealed an approximately eight-times-higher concentration in human colostrum than in mature milk, the latter corresponding to the levels previously detected in human saliva. The high concentration in the colostrum, in particular its functional and structural stability in an acidic milieu, and its growth-supporting role in the taste buds suggest that milk CA VI is an essential factor in normal growth and development of the infant alimentary tract.

Secretion of Carbonic Anhydrase Isoenzyme VI (CA VI) from Human and Rat Lingual Serous von Ebner's Glands

Salivary carbonic anhydrase VI (CA VI) appears to contribute to taste function by protecting taste receptor cells (TRCs) from apoptosis. The serous von Ebners glands locating in the posterior tongue deliver their saliva into the bottom of the trenches surrounding the TRC-rich circumvallate and foliate papillae. Because these glands deliver their saliva directly into the immediate vicinity of TRCs, we investigated whether CA VI is secreted by the von Ebners glands, using immunochemical techniques. The immunohistochemical results showed that CA VI is present in the serous acinar cells, ductal cells, and ductal content of von Ebners glands and in the demilune and ductal cells plus ductal content of rat lingual mucous glands. More importantly, CA VI was also detected in taste buds and in the taste pores. Western blotting of saliva collected from the orifices of human von Ebners glands and CAs purified from rat von Ebners glands confirmed that CA VI is expressed in these glands and secreted to the bottom of the trenches surrounding the circumvallate and foliate papillae. These findings are consistent with the hypothesis that locally secreted CA VI is implicated in the paracrine modulation of taste function and TRC apoptosis. (J Histochem Cytochem 49:657–662, 2001)

Nuclear NonO/p54nrb Protein Is a Nonclassical Carbonic Anhydrase

The growing carbonic anhydrase (CA) gene family includes 11 enzymatically active isozymes in mammals. Each of them has a characteristic cellular and subcellular distribution pattern. In this report, we demonstrate for the first time a nuclear protein with CA activity. A polypeptide recognized by CA II antibodies was purified from several rat tissues using CA inhibitor affinity chromatography. This polypeptide of apparent 66 kDa mass was characterized using amino acid sequencing and CA activity measurements. It appeared to be identical to nonO/p54nrb, a previously cloned and characterized RNA and DNA binding nuclear factor. Recombinant nonO generated in baculovirus bound to the CA inhibitor affinity chromatography matrix and revealed detectable CA activity (25 units/mg). Hanssons histochemical staining of rat lymph nodes followed by light and electron microscopy showed nuclear CA activity in lymphocytes, suggesting that the nuclear nonO protein is catalytically active in vivo. These results demonstrate that a previously known transcription factor is a novel, nonclassical CA. Through its CA activity, the nonO may function in the maintenance of pH homeostasis in the nucleus.