cDNA Cloning, Expression, and Assembly Characteristics of Mouse Keratin 16*
- From the ‡Cancer Research Campaign Cell Structure Research Group, Department of Anatomy and Physiology and the¶Department of Biochemistry, Medical Sciences Institute/Wellcome Trust Building Complex, University of Dundee, Dow Street, Dundee DD1 5EH, United Kingdom
Abstract
There has been speculation as to the existence of the mouse equivalent of human type I keratin 16 (K16). The function of this keratin is particularly intriguing because, in normal epidermis, it is usually confined to hair follicles and only becomes expressed in the suprabasal intrafollicular regions when the epidermis is traumatized. Previous studies suggested that K16 is highly expressed in the skin of mice carrying a truncated K10 gene. We therefore used the skin of heterozygous and homozygous mice to create a cDNA library, and we report here the successful cloning and sequencing of mouse K16. Recent in vitro studies suggested that filaments formed by human K16 are shorter than those formed by other type I keratins. One hypothesis put forward was that a proline residue in the 1B subdomain of the helical domain was responsible. The data presented here demonstrate that this proline is not conserved between mouse and human, casting doubt on the proposed function of this proline residue in filament assembly. In vitro assembly studies showed that mouse K16 produced long filaments in vitro. Also, in contrast to previous observations, transfection studies of PtK2 cells showed that mouse K16 (without the proline) and also human K16 (with the proline) can incorporate into the endogenous K8/K18 network without detrimental effect. In addition, K16 from both species can form filaments de novo when transfected with human K5 into immortalized human lens epithelial cells, which do not express keratins. These results suggest that reduced assembly capabilities due to unusual sequence characteristics in helix 1B are not the key to the unique function of K16. Rather, these data implicate the tail domain of K16 as the more likely protein domain that determines the unique functions.
Footnotes
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↵* This work was supported by Cancer Research Campaign Grant SP2060/01 (to E. B. L.) and Wellcome Trust Grant 46746 (to R. A. Q.).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.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number(s) AF053235.
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↵§ To whom correspondence should be addressed. Tel.: 44-1382-344805; Fax: 44-1382-224117; E-mail: R.M.PORTER{at}dundee.ac.uk.
- Abbreviations:
- KIF
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keratin intermediate filament
- K
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keratin
- hK16
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human keratin 16
- mK16 mouse keratin 16
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RACE, rapid amplification of cDNA ends
- PAGE
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polyacrylamide gel electrophoresis.
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- Received August 25, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
