Molecular Characteristics of the Novel Intermediate Filament Protein Paranemin
SEQUENCE REVEALS EAP-300 AND IFAPa-400 ARE HIGHLY HOMOLOGOUS TO PARANEMIN*
- Philip M. Hemken,
- Robert M. Bellin,
- Suzanne W. Sernett,
- Bruno Becker‡,
- Ted W. Huiatt and
- Richard M. Robson§
- From the Muscle Biology Group, Departments of Biochemistry and Biophysics and of Animal Science, Iowa State University, Ames, Iowa 50011-3260
Abstract
Paranemin was initially found to copurify with the intermediate filament (IF) proteins vimentin and desmin from embryonic chick skeletal muscle and was described as an IF-associated protein (IFAP). We have purified paranemin from embryonic chick skeletal muscle, prepared antibodies, and demonstrated that they label at the Z-lines of both adult avian and porcine cardiac and skeletal muscle myofibrils. We determined the cDNA sequence of paranemin by immunoscreening a λgt22A cDNA library from embryonic chick skeletal muscle. Northern blot analysis revealed a single transcript of 5.3 kilobases, which is much smaller than predicted from the size of paranemin (280 kDa) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The derived amino acid sequence of paranemin (1,606 residues; 178,161 kDa) contains the conserved IF rod domain (308 amino acids), which has highest homology to the rod domains of nestin and tanabin. Thus, paranemin is an IF protein rather than an IFAP. Sequence analysis also revealed that the partial cDNA sequences of two proteins, namely EAP-300 and IFAPa-400, are almost identical to regions of the cDNA sequence of paranemin. The complete paranemin cDNA was expressed in a cell line (SW13) with, and without, detectable cytoplasmic IFs. Antibody labeling of these cells suggests that paranemin does not form IFs by itself, but rather is incorporated into heteropolymeric IFs with vimentin.
Footnotes
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↵* This work was supported in part by grants from the United States Department of Agriculture National Research Initiative Competitive Grants Program Award 96-35206-3744 and the American Heart Association, Iowa Affiliate. This is Paper J-16888 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA 50011, Projects 3444, 3349 and 2127, and supported by the Hatch Act and State of Iowa funds.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) U59287.
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↵‡ Present address: Dept. of Neurobiology HPM, ETH-Hoenggerberg, Zurich, CH-8093 Switzerland.
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↵§ To whom correspondence should be addressed: Muscle Biology Group, 3110 Molecular Biology Bldg., Iowa State University, Ames, IA 50011-3260. Tel.: 515-294-5036; Fax: 515-294-0453; E-mail:rmrobson{at}iastate.edu.
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↵1 The abbreviations used are: IF, intermediate filament; IFAP, intermediate filament-associated protein; PAGE, polyacrylamide gel electrophoresis; kb, kilobase(s); RACE, rapid amplification of cDNA ends; PCR, polymerase chain reaction; bp, base pair(s).
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↵2 P. M. Hemken and R. M. Robson, unpublished observations.
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- Received July 28, 1997.
- Revision received September 16, 1997.
