IOVS Applied and Environmental Microbiology
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(Investigative Ophthalmology and Visual Science. 2006;47:1397-1403.)
© 2006 by The Association for Research in Vision and Ophthalmology, Inc.
DOI:  10.1167/iovs.05-0602
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Distribution of CESP-1 Protein in the Corneal Endothelium and Other Tissues

Rieko Kinouchi,1 Tadatoshi Kinouchi,2,3 Toshirou Hamamoto,2 Takakazu Saito,4 Adriano Tavares,4 Tadahiko Tsuru,1 and Satoru Yamagami5

1From the Departments of Ophthalmology, and 2Biochemistry, Jichi Medical School, Tochigi, Japan; the 3Department of Radiation Life Science and Radiation Medical Science, Research Reactor Institute, Kyoto University, Osaka, Japan; the 4Department of Obstetrics and Gynecology, University of Utah Health Sciences Center, Salt Lake City, Utah; and the 5Department of Corneal Tissue Regeneration, University of Tokyo Graduate School of Medicine, Tokyo, Japan.

PURPOSE. The gene expression profile of human corneal endothelium (CE) was established with the gene signature system. A novel gene, GS3582, was abundantly transcribed in the CE compared with other tissues according to a human gene expression database. This protein was designated corneal endothelium–specific protein (CESP)-1. The tissue distribution and subcellular localization of CESP-1 was assessed in humans and mice, to investigate its physiological function.

METHODS. Rabbit and mouse CESP-1 cDNAs were cloned, and a polyclonal anti-human CESP-1 antibody (Ab) and anti-mouse N- or C-terminal ovary-specific acidic protein (OSAP)-1 Ab were produced. CESP-1 expression was investigated in human and mouse corneas by Western blot and/or immunohistochemical analysis. The distribution of CESP-1 in human tissues was also examined by Western blot analysis. To identify the subcellular localization of CESP-1, cultured human CE was colabeled with anti-human CESP-1 Ab and anti-cytochrome c monoclonal Ab or anti-GRP78 monoclonal Ab for confocal microscopy.

RESULTS. The rabbit and mouse CESP-1 cDNA sequences contained an open reading frame coding 242 and 283 amino acids, respectively. Mouse CESP-1 was entirely consistent with mouse OSAP. Western blot analysis showed that CESP-1 was expressed in the human corneal epithelium, CE, cultured CE, brain, testis, and ovary. Mouse CESP-1 was also expressed in mouse corneal epithelium and CE with anti-mouse C- but not N-terminal OSAP Ab according to immunohistochemical analysis. Subcellular localization of CESP-1 to the mitochondria was demonstrated in cultured human CE. The N-terminal of CESP-1, possessing a mitochondrial targeting sequence, may be processed after the protein is imported into the mitochondria.

CONCLUSIONS. CESP-1 was distributed in the corneal epithelium, the CE and cultured human CE, as well as the brain, testis, and ovary. CESP-1 was localized in the mitochondria of cultured human CE. These findings may provide some clues about the physiological function of CESP-1.






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