An Open Reading Frame Element Mediates Posttranscriptional Regulation of Tropoelastin and Responsiveness to Transforming Growth Factor β1

Elastin is a resilient connective tissue protein present in the extracellular matrix of most terrestrial vertebrate tissues but, because of its unique elastomeric properties, it is especially abundant in the interstitium of tissues that undergo repeated physical deformations, such as lungs, blood vessels, and skin (43). Elastic fibers are assembled extracellularly and are comprised of elastin and microfibrillar proteins (40). Elastin itself is a polymer of enzymatically cross-linked tropoelastin monomers, the secreted, soluble precursor protein, and constitutes ca. 90% of the mass of elastic fibers. Like other structural extracellular matrix proteins, the bulk of elastin production is limited to a narrow window of development. In most tissues, elastogenesis begins around the time of midgestation, peaks near birth and during early neonatal periods, drops sharply thereafter, and is nearly completely repressed by maturity (46). Because elastin is an extremely durable polymer and essentially does not turn over in healthy tissues (56, 63), fiber function and tissue integrity are not compromised by this limited pattern of production. While transcriptional regulation controls both the turning on and turning off of many developmentally regulated, tissue-specific genes, we determined previously that tropoelastin production is governed by distinct mechanisms acting at different stages of growth (69). Whereas gene transcription controls the induction of tropoelastin expression in utero, a posttranscriptional mechanism mediating rapid decay of the mRNA regulates the dwindling tropoelastin expression during postnatal growth and maintains protein production at undetectable levels in adult tissue (43). In addition to our in vivo studies, regulation of mRNA turnover has been shown to control the repression and reinitiation of tropoelastin expression in a variety of cell models. We reported that vitamin D3 and phorbol ester (phorbol myristate acetate) potently repress tropoelastin expression in rodent and bovine cells by mediating an accelerated decay of its mRNA with no effect on gene transcription (45, 51). Similarly, downregulation of tropoelastin mRNA levels mediated by glucocorticoids or aprotinin or that which occurs in freshly isolated tissue is controlled solely by a reduction in the mRNA half-life (23, 25, 37). In addition, transforming growth factor β1 (TGF-β1) stimulates the low levels of tropoelastin production by adult human and rat fibroblasts from various tissues by increasing the stability of tropoelastin mRNA (24, 30, 36, 71). Thus, modulation of mRNA turnover regulates elastin production in vivo, ex vivo, and in cell-based models, but the precise mechanism controlling transcript decay is not known. The half-life of mRNA transcripts is influenced by poly(A) tail length and by regulatory sequences located in the 5′ or 3′ untranslated regions (UTRs) or within the open reading frame (5, 7, 58), and these elements interact with specific RNA-binding proteins (2, 49). The heterogeneous localization of regulatory elements suggests that mRNA decay is not mediated by a common pathway. Tropoelastin mRNA does not contain any sequences that have been demonstrated or suggested to mediate degradation of other transcripts, such as AU-rich regions and, thus, decay of tropoelastin mRNA may be controlled by unique cis-acting sequences. Typically, the rate at which an mRNA is degraded is determined by the activity of destabilizing sequences and not by stabilization sequences (59), though stabilization sequences have been identified in many transcripts (16, 31, 39, 66). As reported here, we have identified an element in the translated portion of tropoelastin mRNA that specifically binds a cytosolic protein. The level of this binding activity increases as tropoelastin expression declines with age. In addition, binding activity decreases in response to TGF-β1, which, as mentioned above, is known to mediate the stabilization of tropoelastin mRNA. Our findings indicate that the interaction of this cytosolic factor with tropoelastin mRNA element controls elastin production in growing and mature tissues.