Transketolase Is a Major Protein in the Mouse Cornea*
- Christina M. Sax‡§,
- Csaba Salamon‡,
- W. Todd Kays‡,
- Jing Guo¶,
- Fushin X. Yu¶,
- R. Andrew Cuthbertson‡ and
- Joram Piatigorsky‡
- From the ‡ Laboratory of Molecular and Developmental Biology, National Eye Institute, National Institutes of Health, MSC 2730, Bethesda, Maryland 20892-2730 and
- ¶ The Schepens Eye Research Institute, Harvard Medical School, Boston, Massachusetts 02114-2508
- §To whom correspondence should be addressed. Tel.: 301-402-4342; Fax: 301-402-0781.
Abstract
Earlier experiments in this laboratory identified a highly expressed 65-68-kDa protein in both mouse and human corneas (Cuthbertson, R. A., Tomarev, S. I., and Piatigorsky J. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 4004-4008). Here, we demonstrate that this protein is transketolase (TKT; EC 2.2.1.1), an enzyme in the nonoxidative branch of the pentose-phosphate pathway, based on peptide and cDNA isolation and sequence analysis of mouse cornea protein and RNA samples, respectively. While expressed at low levels in a number of tissues, the 2.1-kilobase TKT mRNA was expressed at a 50-fold higher level in the adult mouse cornea. The area of most abundant expression was localized to the cornea epithelial cell layer by in situ hybridization. Western blot analysis confirmed TKT protein abundance in the cornea and indicated that TKT may comprise as much as 10% of the total soluble protein of the adult mouse cornea. Soluble cornea extracts exhibited a correspondingly high level of TKT enzymatic activity. TKT expression increased progressively through cornea maturation, as shown by Northern blot, in situ hybridization, Western blot, and enzymatic analyses. TKT mRNA and protein were expressed at low levels in the cornea prior to eye opening, while markedly increased levels were observed after eye opening. Taken together, these observations suggest that TKT may be a cornea enzyme-crystallin, and suggest that the crystallin paradigm and concept of gene sharing, once thought to be restricted to the lens, apply to other transparent ocular tissues.
Footnotes
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↵* These studies were supported in part by National Eye Institute Grant EY10869 (to F. X. Y.). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- TKT
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transketolase
- HCP
-
human cornea protein
- MCP
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mouse cornea protein
- oligo
-
oligonucleotide
- PBS
-
phosphate-buffered saline
- PCR
-
polymerase chain reaction
- PP
-
pentose-phosphate
- RT
-
reverse transcriptase.
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↵2 C. M. Sax and J. Piatigorsky, unpublished data.
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↵3 W. T. Kays and J. Piatigorsky, unpublished data.
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- Received June 14, 1996.
- Revision received September 30, 1996.
- © 1996 by The American Society for Biochemistry and Molecular Biology, Inc.
