Identification and Characterization of a New Latent Transforming Growth Factor-β-binding Protein, LTBP-4*
- From the Departments of ‡Virology and§Medical Genetics, The Haartman Institute and the¶Department of Dermatology and Venereology, University of Helsinki, Helsinki FIN-00014, Finland
Abstract
Transforming growth factor βs (TGF-βs) are secreted by most cell types as latent high molecular weight complexes consisting of TGF-β and its latency associated peptide (LAP) propeptide dimers, covalently linked to latent TGF-β-binding proteins (LTBPs). Currently, three different LTBPs are known (LTBPs 1, 2, and 3), all with highly similar protein domain structure consisting of epidermal growth factor-like and 8-Cys repeats. The 3rd 8-Cys repeat of LTBP-1 mediates its association with TGF-β1·LAP. By using an expressed sequence tag homologous to the 3rd 8-Cys repeat of human LTBP-1 as a probe, a novel cDNA similar to known LTBPs was cloned from human heart cDNA library. This cDNA was named LTBP-4 and found to exist in at least four different forms, generated by alternative splicing at the amino terminus and at the central epidermal growth factor repeat domain. One of the alternative amino-terminal forms contained an RGD sequence, indicating possible cell-surface interactions with integrins. LTBP-4 gene was localized to chromosomal position 19q13.1–19q13.2. The major LTBP-4 mRNA form is about 5.1 kilobase pairs in size and is predominantly expressed in the heart, aorta, uterus, and small intestine. Immunoblotting analysis indicated that LTBP-4 was secreted from cultured human lung fibroblasts both in a free form and in a disulfide bound complex with a TGF-β·LAP-like protein. Both LTBP-4 forms were also found to be deposited in the extracellular matrix. The matrix-associated LTBP-4 was susceptible to proteolytic release with plasmin. LTBP-4 is a new member of the growing LTBP-fibrillin family of proteins and offers an alternative means for the secretion and targeted matrix deposition of TGF-βs or related proteins.
Footnotes
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↵* This work was supported by The Academy of Finland, Sigrid Juselius Foundation, Finnish Cultural Foundation, Finnish Cancer Organizations, Maud Kuistila Foundation, Novo Nordisk Foundation, Paulo Foundation, Biocentrum Helsinki, Helsinki University Hospital, and the University of Helsinki.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) (LTBP-4S), (LTBP-4L partial cDNA, (LTBP-4ΔEGF), and (LTBP-4Δ2EGF).
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↵‖ To whom correspondence should be addressed: The Haartman Institute, Dept. of Virology, University of Helsinki, P.O. Box 21 (Haartmaninkatu 3), FIN-00014 University of Helsinki, Helsinki, Finland. Tel.: 358-9-1912-6476; Fax: 358-9-1912-6475.
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↵1 The abbreviations used are: TGF-β, transforming growth factor-β; LAP, latency associated peptide; LTBP-1, LTBP-2, LTBP-3 and LTBP-4, latent TGF-β-binding proteins 1, 2, 3, and 4; EGF, epidermal growth factor; ECM, extracellular matrix; 8-Cys repeat, a conserved protein sequence motif containing eight cysteines found in LTBPs and fibrillins, also called LT domain; LTBP-4S, LTBP-4L, LTBP-4Δ2EGF and LTBP-4ΔEGF different alternatively spliced forms of LTBP-4; CHO, Chinese hamster ovary; PAGE, polyacrylamide gel electrophoresis; ORF, open reading frame; EST, expressed sequence tag; Ab, antibody.
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↵3 J. Saharinen and J. Keski-Oja, manuscript in preparation.
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↵4 J. Saharinen, C. Koski, and J. Keski-Oja, manuscript in preparation.
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- Received December 12, 1997.
- Revision received March 31, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
