A Sequence-selective Single-strand DNA-binding Protein Regulates Basal Transcription of the Murine Tissue Inhibitor of Metalloproteinases-1 (Timp-1) Gene

Abstract

Tissue inhibitor of metalloproteinases-1 (TIMP-1) is important in maintaining the extracellular proteolytic balance during tissue remodeling processes. To allow homeostatic tissue turnover, the murine Timp-1 gene is expressed by most cells at a low basal level, and during acute remodeling its transcription is activated by a variety of stimuli. A non-consensus AP-1-binding site (5-TGAGTAA-3) that is conserved in mammalian timp-1 genes is a critical element in basal and serum-stimulated transcription. We show here that each strand of this unusual AP-1 site binds a distinct single-stranded DNA-binding protein, although neither strand from a perfect consensus AP-1 site from the human collagenase gene shows similar binding. One of the single-strand binding factors, which we term ssT1, binds to a second upstream Timp-1 region between nucleotides ؊115 and ؊100. Deletion analysis demonstrated that this region is important in basal but not serum-inducible transcription. The ssT1 factor was 52-54 kDa by UV cross-linking of electrophoretic mobility shift assays and Southwestern blot analysis. Its binding to DNA shows sequence selectivity rather than specificity, with 5-CT/ATTN (4 -6)ATC-3 as a favored motif. Multiple ssT1-like activities were found in nuclear extracts from mouse fibroblasts and rat liver and testis, suggesting that these factors may regulate basal Timp-1 transcription in a tissue-specific fashion.Extracellular matrix turnover is highly regulated at multiple levels, a critical one being the balance between the levels of matrix metalloproteinases (MMPs) 1 and their specific inhibitors, the TIMPs (tissue inhibitors of metalloproteinases (1, 2)).

JAC , a direct target of oncogenic transcription factor Jun, is involved in cell transformation and tumorigenesis

“…A TATA box is present 30 bp upstream of the experimentally determined transcriptional start site (data not shown) located at position 3835. Intriguingly, the 5Ј upstream region contains one authentic 5Ј-TGACTCA-3Ј heptamer motif for high-affinity AP-1 binding (2, 6, 7) at position 3478-3484 and, only 14 bp further downstream, a nonconsensus AP-1 binding site 5Ј-TGAGTAA-3Ј, representing a motif that was identified as a functional element in some AP-1-regulated genes (13,14). When the 622-bp JAC cDNA probe was used, three EcoRI fragments hybridizing under stringent conditions were detected by Southern analysis of chicken genomic DNA (Fig.…”

“…Jun-Jun and Jun-Fos dimers bind preferentially with high affinity to enhancer elements with the consensus sequence 5Ј-TGAC͞GTCA-3Ј located in various AP-1-responsive genes (6,7), but nonconsensus binding motifs differing in single nucleotides have also been identified (13,14). There is increasing evidence that cellular transformation induced by the v-Jun protein involves the aberrant expression of specific genes that are normally regulated by endogenous c-Jun as a component of AP-1.…”

Differential expression and localization of TIMP-1 and TIMP-4 in human gliomas

“…Individual TIMPs differ markedly in their pro-MMP interactions and gene regulatory mechanisms. For example, TIMP-1 specifically interacts with pro-gelatinase-B and is subject to tight control at the level of transcription (Phillips et al, 1999), whereas TIMP-2 binds pro-gelatinase A and shows constitutive gene expression (Blavier et al, 1999). Differences also exist with regard to biochemical properties: TIMP-3 is itself an ECM-associated protein but TIMP-1, -2 and -4 are freely diffusible (Leco et al, 1994).…”

The Comparative Role of Activator Protein 1 and Smad Factors in the Regulation of Timp-1 and MMP-1 Gene Expression by Transforming Growth Factor-β1

“…2 Interestingly, the Timp-1 AP1 site is noncanonical (5Ј-TGAGTAA-3Ј), and this is conserved across all species sequenced, whereas the MMP-1 AP1 site is the consensus sequence (5Ј-TGAGTCA-3Ј). The Timp-1 AP1 site has recently been reported to bind a distinct nuclear factor compared with the consensus; this factor, ssT1, is a singlestranded DNA-binding protein of unknown identity or function (23).…”