Multiple silencer elements are involved in regulating the chicken vimentin gene.

Abstract

Vimentin, a member of the intermediate filament protein family, exhibits tissue-as well as developmentspecific expression. Transcription factors that are involved in expression of the chicken vimentin gene have been described and include a cis-acting silencer element (SE3) that is involved in the down-regulation of this gene (F. X. Farrell, C. M. Sax, and Z. E. Zehner, Mol. Cell. Biol. 10:2349Biol. 10: -2358Biol. 10: , 1990. In this study, we report the identification of two additional silencer elements (SE1 and SE2). We show by transfection analysis that all three silencer elements are functionally active and that optimal silencing occurs when multiple (at least two) silencer elements are present. In addition, the previously identified SE3 can be divided into three subregions, each of which is moderately active alone. By (4,22,28,39). For example, induction of vimentin expression has been linked to serum, platelet-derived growth factor, and phorbol esters (23,38,39), in addition to more general transcription factors such as SP-1 (8). In the chicken vimentin gene, differential vimentin expression has been shown to involve several cis elements and trans factors (43-45, 50, 53, 54). However, one of the more interesting aspects of vimentin gene regulation is that it is also down-regulated during development. The primary locale of this negative regulation was determined to be a 40-bp silencer element (SE3) located at -607 to -567 from the start site of transcription (14,44). This element was shown to bind a factor of 95 kDa (14), the silencer element protein (SEP). Upon further analysis, SE3 was observed to contain some sequence similarity to the octamer-binding sequence (4) which has been observed in a negative element in the human c-myc gene (51). However, the data suggest that SEP is a different factor.One important conclusion from this study was that while SE3 could act in a position-and orientation-independent manner, a single copy yielded only a partial reduction in transcription within the native context of the gene and 36% compared with that of the heterologous herpes simplex virus (HSV) thymidine kinase (tk) promoter (ptkCAT) (14). In fact, multiple SE copies (more than two) were (14). Here, we show by transfection analyses that there are at least three SEs active within the chicken vimentin gene. Moreover, all three elements appear to bind the same silencer protein (95 kDa) as measured by gel mobility shift assays (GMSAs), UV cross-linking experiments, and Southwestern (DNA-protein) blots. Transfection data suggest that for complete repression of transcription, multiple (at least two) SE copies are required.The presence of multiple negative elements is not unique to the chicken vimentin gene. Since our initial report, multiple negative elements have also been found in the chicken lysozyme (2, 3), rat glutathione P-transferase (19), human 8-globin (9), interleukin-4 (30), collagen II (42), and hamster (52) and human (41) vimentin genes. Possible mechanistic models are presented to account for the nee...

AP-1, ETS, and transcriptional silencers regulate retinoic acid-dependent induction of keratin 18 in embryonic cells.

“…Silencer activity has been implicated in the regulation of the vimentin gene, another intermediate filament gene (12,17,50,59). However, the expression of vimentin in F9 cells, the absence of apparent sequence homology to the K18 silencers, and the activity of the vimentin silencers in several somatic cell types suggest that the silencer elements of the two types of intermediate filament genes are probably mediated by different factors.…”

Tissue-Restricted Expression of the Cardiac α-Myosin Heavy Chain Gene Is Controlled by a Downstream Repressor Element Containing a Palindrome of Two Ets-Binding Sites

“…Although the mechanism of the PNR element-mediated gene repression remains to be determined, the presence of multiple Ets-binding sites as a palindrome at the downstream position of the ␣-MHC gene and the involvement of the upstream sites would indicate that it might be a requirement for the physical hindrance and/or conformational rigidity needed for the negative control of gene transcription initiation. Similarly, other studies have shown a requirement of multiple sites for silencing the myosin light chain-2, collagen-II, and vimentin gene expression (10,12,53).…”

Promoter-independent regulation of vimentin expression in mammary epithelial cells by val12 ras and TGFβ

“…That the vimentin promoter sequence is functionally conserved is shown by the identification of numerous transcription sites on the human promoter in mouse cells [17,18,52,53] and in the chicken promoter in mouse and human cells [19,32,51]. Transgenic mouse studies also suggested that regulation of the human promoter in mouse cells and tissues parallels the endogenous gene [47].…”

“…Both r3 and r3T cells were used for these analyses, since they have similar levels of vimentin mRNA expression: we compared these cells to NIH3T3, which express 100-fold higher levels of vimentin in the absence of TGFβ ( Figure 3D). The human vimentin promoter has been characterized in a variety of fibroblasts, and the well studied 1.5 kb promoter contains both enhancer and silencer elements [17,19,20]. We initially examined three promoter constructs containing 1500, 411 and 268 nucleotides of promoter sequence fused to a CAT reported gene.…”

“…The regulation of vimentin expression in mesenchymal cells has been well studied, and transcription factor binding sites responsible for serum stimulation and silencing have been described [17][18][19][20][21]. In epithelial cells, vimentin expression is higher in migratory cells, and may contribute to the migratory and invasive phenotype of metastatic cells [21].…”