Transforming growth factor betas (TGF-betas) are secreted by most cell types as latent high molecular weight complexes consisting of TGF-beta and its latency associated peptide (LAP) propeptide dimers, covalently linked to latent TGF-beta-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-beta1.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-beta. 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-betas or related proteins.