Anti-BSND (aa 1-320) polyclonal antibody

BSND protein, which is involved in chloride transport, is expressed in normal kidney and the inner ear and is known as an immunohistochemical marker for chromophobe renal cell carcinoma (RCC) and renal oncocytoma; however, other organs and tumor types exhibiting BSND expression have not yet been reported. In this study, we investigated the expression of BSND using data from the Cancer Genome Atlas (TCGA) database and by performing immunohistochemical analyses. As a result, we found that BSND was also expressed in the striated duct cells of normal salivary glands. Next, BSND expression was examined immunohistochemically in 7 types of salivary gland tumors, and BSND positivity was found in Warthin's tumor (25 out of 25 cases; 100%) and oncocytoma (4/4; 100%), both of which are usually classified as oncocytic tumors, whereas BSND negativity was observed for pleomorphic adenoma (0/11), adenoid cystic carcinoma (0/7), acinic cell carcinoma (0/6), mucoepidermoid carcinoma (0/6), and salivary duct carcinoma (0/5). Finally, the expression of BSND mRNA in 30 types of tumors other than chromophobe RCC and salivary gland tumors was examined using data from the TCGA database, but none of these tumors exhibited BSND expression. These results suggest that BSND is expressed only in normal salivary glands and oncocytic salivary gland tumors such as Warthin's tumor and oncocytoma in addition to the two known organs and the two known renal tumor types mentioned above. The selective expression pattern of BSND suggests that BSND is an excellent novel immunohistochemical marker for oncocytic salivary gland tumors.

Barttin, a gene product of BSND, was identified as a fourth gene responsible for Bartter syndrome. The co-expression of barttin with CLC-K chloride channels has been demonstrated to dramatically induce the expression of CLC-K current. However, it remains unknown how barttin interacts with CLC-K channels in mammalian cells and how the mutations of barttin lead to Bartter syndrome. In an attempt to clarify the effect of barttin expression on CLC-K2 cellular localization, we examined the expression of the CLC-K2 chloride channel and barttin, solely and in combination, in transient and stable expression systems in mammalian cells. In addition, we generated several stable cell lines expressing mutant barttins to clarify the consequence of the previously reported barttin mutations in Bartter syndrome. In immunocytochemistry, CLC-K2 was retained in the Golgi in the absence of barttin expression, but delivered to the plasma membrane when barttin was present. Barttin was coprecipitated with CLC-K2, suggesting a protein-protein interaction. Disease-causing mutant barttins, especially R8L, were retained intracellularly, but their binding ability to CLC-K2 was preserved. This led to a retention of CLC-K2 in intracellular organelles with barttin, and a loss of plasma membrane localization. The stability of the CLC-K2 protein was also markedly increased by coexpression with barttin. These results clarified that barttin determined cellular localization of CLC-K2 by protein-protein interaction. Thus, the mislocalization of CLC-K2 was identified as the molecular pathogenesis of Bartter syndrome by mutant barttins.