Localization of Selenoproteins in the Perinuclear Structure of the Rat Kidney

Abstract

Investigations were performed on rats to obtain information on the distribution of selenoproteins in the microsomal fraction, especially in the perinuclear structure [endoplasmatic reticulum (ER) and Golgi apparatus] of the kidney. For the determination of the selenium levels instrumental neutron activation analysis (INAA) via ⁷⁵Se was used. The membrane‐bound selenium‐containing proteins were detected by labeling of rats with ⁷⁵Se‐selenite, subcellular fractionation of the tissue homogenates, separation of the proteins by SDS‐PAGE, or two‐dimensional electrophoresis and autoradiography of ⁷⁵Se. In this way, at least 17 membrane‐bound selenium‐containing proteins could be distinguished in the microsomal fraction of the kidney. Of those, six selenium‐containing proteins with molecular masses of about 15, 16, 20, 23–25, 42–44, and 58–60 kDa were found in the ER fraction, which was isolated from the microsomes by differential centrifugation and further purification steps with detergents. The ⁷⁵Se‐labeled bands with molecular masses of 15 and 20 kDa and those in the range of 23–25, 42–44, and 58–60 kDa, could be attributed to known selenoproteins. A further ⁷⁵Se‐labeled 16 kDa protein, which had been detected in an earlier study, was characterized as a selenocysteine (Sec)‐containing selenoprotein with an isoelectric point of 5.4. The localization of the different selenoproteins in the ER is of great interest with regard to the functions of these cellular organelles.

Keywords:

• Selenoproteins
• Electrophoresis
• Autoradiography
• HPLC
• MALDI‐MS
• INAA
• Rat
• Kidney
• Endoplasmatic reticulum

Acknowledgments

This study was partly supported by the Deutsche Forschungsgemeinschaft (DFG). We would like to thank Prof. Dr. N.C Juhr, Institut für Versuchstierkunde und Versuchstierkrankheiten, Freie Universität Berlin, and Mr. Franke, Klinikum Benjamin Franklin, Freien Universität Berlin, for their help with the animal experiments.