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• W2094663606 abstract "The pancreatic elastase I gene (ELA1) is selectively transcribed to high levels in pancreatic acinar cells. Pancreatic specificity is imparted by a 100-base pair enhancer that activates transcription in β-cells of the islets of Langerhans as well as in acinar cells. Adjacent to the enhancer is a silencer that renders transcription specific to acinar cells by selectively suppressing the inherent β-cell activity of the enhancer. We show that the selective repression of β-cell transcription is due neither to a β-cell specific activity of the silencer nor to selective interference with β-cell-specific transcriptional activators acting on the enhancer. Rather, the silencer is effective in both pancreatic endocrine and acinar cell types against all low and moderate strength enhancers and promoters tested. The silencer appears to act in a binary manner by reducing the probability that a promoter will be active without affecting the rate of transcription from active promoters. We propose that theELA1 silencer is a weak off switch capable of inactivating enhancer/promoter combinations whose strength is below a threshold level but ineffective against stronger enhancer/promoters. The apparent cell-specific effects on the ELA1 enhancer appear due to the ability of the silencer to inactivate the weak β-cell activity of the enhancer but not the stronger acinar cell activity. The pancreatic elastase I gene (ELA1) is selectively transcribed to high levels in pancreatic acinar cells. Pancreatic specificity is imparted by a 100-base pair enhancer that activates transcription in β-cells of the islets of Langerhans as well as in acinar cells. Adjacent to the enhancer is a silencer that renders transcription specific to acinar cells by selectively suppressing the inherent β-cell activity of the enhancer. We show that the selective repression of β-cell transcription is due neither to a β-cell specific activity of the silencer nor to selective interference with β-cell-specific transcriptional activators acting on the enhancer. Rather, the silencer is effective in both pancreatic endocrine and acinar cell types against all low and moderate strength enhancers and promoters tested. The silencer appears to act in a binary manner by reducing the probability that a promoter will be active without affecting the rate of transcription from active promoters. We propose that theELA1 silencer is a weak off switch capable of inactivating enhancer/promoter combinations whose strength is below a threshold level but ineffective against stronger enhancer/promoters. The apparent cell-specific effects on the ELA1 enhancer appear due to the ability of the silencer to inactivate the weak β-cell activity of the enhancer but not the stronger acinar cell activity. base pair(s) human growth hormone kilobase pair(s) Dulbecco's modified Eagle's medium Rous sarcoma virus Rous sarcoma virus herpes simplex virus thymidine kinase histone deacetylase The complementary action of positive and negative transcriptional control is a common regulatory strategy for cell-specific genes (1Ogbourne S. Antalis T.M. Biochem. J. 1998; 331: 1-14Crossref PubMed Scopus (172) Google Scholar). In many instances transcriptional enhancers direct expression to several cell types in an organ, generally due to the action of an organ-specific, but not cell-specific, transcriptional activator or combination of activators. Cell-specific expression is then resolved by the action of a repressor/silencer region that prevents transcription in the inappropriate cell type(s). The basis for cell type restriction by transcriptional silencers is not well understood, although recent evidence indicates repressive chromatin structure may be initiated by targeted recruitment of histone deacetylases or other chromatin-modifying enzymes (for reviews see Refs. 2Ayer D.E. Trends Cell Biol. 1999; 9: 193-198Abstract Full Text Full Text PDF PubMed Scopus (245) Google Scholar and 3Tyler J.K. Cell. 1999; 99: 4443-4446Abstract Full Text Full Text PDF Scopus (203) Google Scholar). Switching between open (accessible) and closed (inaccessible) forms of chromatin (4Jackson M.E. J. Cell Sci. 1991; 100: 1-7Crossref PubMed Google Scholar, 5Kingston R.E. Bunker C.A. Imbalzano A.N. Genes Dev. 1996; 10: 905-920Crossref PubMed Scopus (403) Google Scholar) is consistent with all-or-none mechanisms of transcriptional control by enhancers (6Fiering S. Whitelaw E. Martin D.I. BioEssays. 2000; 22: 381-387Crossref PubMed Scopus (135) Google Scholar) and presumably silencers as well.Selective transcription of pancreatic genes in exocrineversus endocrine cells requires both positive and negative forms of transcriptional control. For example, the restricted transcription of insulin in pancreatic β-cells (7Nir U. Walker M.D. Rutter W.J. Proc. Natl. Acad. Sci. U. S. A. 1988; 83: 3180-3184Crossref Scopus (86) Google Scholar, 8Boam D.S.W. Clark A.R. Docherty K. J. Biol. Chem. 1990; 265: 8285-8296Abstract Full Text PDF PubMed Google Scholar, 9Goodman P.A. Medina-Martinez O. Fernandez-Mejia C. Mol. Cell. Endocrinol. 1996; 120: 139-146Crossref PubMed Scopus (37) Google Scholar) and elastase in acinar cells (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar) is mediated by the complementary action of enhancers and silencers. β-Cells and acinar cells are constituents of the endocrine and exocrine compartments, respectively, of the compound pancreatic gland. Most of the pancreas is exocrine, organized into acinar structures that secrete a set of digestive hydrolases such as amylase, ribonuclease, and elastase into the intestine. Only approximately 2% of the pancreas is endocrine tissue, organized into islets of Langerhans comprising four cell types, each producing one principal polypeptide hormone. The majority of islet cells are β-cells, which produce insulin. During embryonic pancreatic development mature acinar and islet cells arise from transitional epithelial cells that express both acinar- (e.g. amylase) and islet (e.g. insulin)-specific markers (11Guz Y. Montminy M.R. Stein R. Leonard J. Gamer L.W. Wright C.V.E. Teitelman G. Development. 1995; 121: 11-18Crossref PubMed Google Scholar). Differentiation of these transitional cells leads to silencing of acinar genes in islet cells and vice versa. The elastase I gene (ELA1) is one of the complement of pancreatic genes encoding digestive enzymes that are expressed selectively in the acinar cells. In this report we describe a transcriptional silencer that prevents expression of rat ELA1 in islet cells while permitting expression in acinar cells.Activation and repression of ELA1 gene transcription is mediated through 5′-proximal gene flanking sequences (Fig. 1). A 100-bp,1 three element transcriptional enhancer (−195 to −96 relative to the transcriptional start) contains regulatory information that directs pancreatic expression to both acinar and β-cells of transgenic mice (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar, 12Hammer R.E. Swift G.H. Ornitz D.M. Quaife C.J. Palmiter R.D. Brinster R.L. MacDonald R.J. Mol. Cell. Biol. 1987; 7: 2956-2967Crossref PubMed Scopus (52) Google Scholar, 13Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Mol. Cell. Biol. 1995; 15: 4385-4394Crossref PubMed Scopus (35) Google Scholar, 14Rose S.D. Kruse F. Swift G.H. MacDonald R.J. Hammer R.E. Mol. Cell. Biol. 1994; 14: 2048-2057Crossref PubMed Scopus (34) Google Scholar). The endogenous ELA1 gene as well as transgenes containing as little as 501 bp of 5′-flanking sequences are not expressed in pancreatic islets, however (15Ornitz D.M. Palmiter R.D. Messing A. Hammer R.E. Pinkert C.A. Brinster R.L. Cold Spring Harbor Symp. Quant. Biol. 1985; 50: 399-409Crossref PubMed Google Scholar). The appropriate acinar-specific transcription is imposed by a negative regulatory region located immediately upstream of the enhancer (between −501 and −202) (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar).The acinar and β-cell specific activities of the ELA1enhancer have been assigned to two separate transcriptional elements (Fig. 1), based on their activity in transgenic animals and transfected pancreatic cell lines (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar, 14Rose S.D. Kruse F. Swift G.H. MacDonald R.J. Hammer R.E. Mol. Cell. Biol. 1994; 14: 2048-2057Crossref PubMed Scopus (34) Google Scholar, 16Swift G.H. Liu Y. Rose S.D. Bischof L. Steelman S. Buchberg A.M. Wright C.V.E. MacDonald R.J. Mol. Cell. Biol. 1998; 18: 5109-5120Crossref PubMed Scopus (146) Google Scholar). The 25-bp A element is the sole positively acting transcriptional element for acinar transcription. The latent β-cell specific activity of the three-element enhancer is due to a 12-bp transcriptional element, the B element. The B element is also active in acinar cells, but in these cells it has a different function, which is to augment the activity of the acinar-specific A element (13Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Mol. Cell. Biol. 1995; 15: 4385-4394Crossref PubMed Scopus (35) Google Scholar). A third element (C) is inactive on its own, but in combination with either the A or the B element augments their activity nearly 10-fold without contributing to or affecting cell specificity (13Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Mol. Cell. Biol. 1995; 15: 4385-4394Crossref PubMed Scopus (35) Google Scholar). In β-cells the activity of the enhancer is mediated by the combination of the B and C elements, whereas in acinar cells all three elements are active. The silencer region (−501 to −202) suppresses the BC element activity in β-cells of mice without detectably altering the activity of the three-element (ABC) enhancer in acinar cells (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar).We set out to determine the regulatory strategy that the silencer uses to suppress selectively the action of the ELA1 enhancer in pancreatic β-cells. For example, the silencer might selectively antagonize the action of the enhancer B element, which is responsible for the β-cell activity. Alternatively, the silencer may be active in β-cells but not in acinar cells; for example, the factors that mediate its activity may be present selectively in β-cells. We report that the action of the silencer is specific neither for the B element nor to β-cells. Instead we present evidence that the apparent selective action of the enhancer is based on its ability to suppress effectively the relatively weak two-element (B + C) enhancer active in β-cells but not the complete enhancer with the three elements (A + B + C) working in concert in acinar cells. We show that the silencer acts in a binary manner to block effectively transcription from a promoter with a weak enhancer.RESULTSInitially we tested two likely mechanisms for the β-cell-specific suppression of the ELA1 enhancer by the silencer. For mechanism 1, the silencer might selectively antagonize the action of the pancreatic transcription factor PDX1, which mediates the action of the B element in islet β-cells (16Swift G.H. Liu Y. Rose S.D. Bischof L. Steelman S. Buchberg A.M. Wright C.V.E. MacDonald R.J. Mol. Cell. Biol. 1998; 18: 5109-5120Crossref PubMed Scopus (146) Google Scholar). For mechanism 2, the silencer may be inactive in acinar cells; for example, the factors that bind to and mediate the activity of the silencer may be absent from acinar cells.To test these mechanisms, it was first necessary to verify that the silencer, active in β-cells of transgenic mice, was also active in transfected pancreatic β-cell lines. RIN1046-38 (hereafter RIN38) is a well differentiated, insulin-synthesizing β-cell line (24Phillipe J. Chick W.L. Habener J.F. J. Clin. Invest. 1987; 79: 351-358Crossref PubMed Scopus (105) Google Scholar). At low passages (as in these experiments) RIN38 cells retain many differentiated characteristics of pancreatic β-cells, including moderate levels of insulin mRNA and glucose-regulated secretion of insulin (25Clark S.A. Burnham B.L. Chick W.L. Endocrinology. 1990; 127: 2779-2788Crossref PubMed Scopus (87) Google Scholar). The β-cell activity of the B element can be mimicked in transgenic animals and cultured cells by linking a pentamer of the B element to the minimal ELA1 promoter spanning nucleotides −92 to +8 (Fig. 1 and Ref. 18Higuchi R. Krummel B. Saiki R.K. Nucleic Acids Res. 1988; 16: 7351-7367Crossref PubMed Scopus (2087) Google Scholar). In animals the minimal ELA1 promoter is inactive, and the addition of the B pentamer activates transcription selectively in β-cells (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar). In transfected RIN38 cells the low activity of the minimal promoter is increased 20-fold by the B element pentamer (Fig. 2, constructs A andB). The B pentamer is also active in a second independently derived and well differentiated β-cell line, βTC3 (Ref. 16Swift G.H. Liu Y. Rose S.D. Bischof L. Steelman S. Buchberg A.M. Wright C.V.E. MacDonald R.J. Mol. Cell. Biol. 1998; 18: 5109-5120Crossref PubMed Scopus (146) Google Scholar and data not shown), but is inactive in non-β-cell lines, such as pancreatic acinar effectively, NIH3T3, and Rat2 fibroblast lines (16Swift G.H. Liu Y. Rose S.D. Bischof L. Steelman S. Buchberg A.M. Wright C.V.E. MacDonald R.J. Mol. Cell. Biol. 1998; 18: 5109-5120Crossref PubMed Scopus (146) Google Scholar).The 300-bp ELA1 silencer region (−501 to −202) reduced the activity of the B pentamer in transfected RIN38 cells (Fig. 2 C), consistent with its ability to suppress the action of the ELA1 enhancer in β-cells of transgenic mice. The silencer decreased the activity of the B pentamer 12–15-fold, so that the residual activity was approximately that of the minimal promoter alone. The silencer also was effective in reverse orientation (Fig. 2 D). However, the silencer region was not effective when moved to the other side of the hGH reporter gene; this position is 2.2 kb downstream and 2.8 kb upstream of the promoter in the circular plasmid (Fig. 2, E and F). Thus, the action of the silencer is independent of its orientation but affected by distance from the promoter.The ELA1 Silencer Is Not Specific for the ELA1 Minimal Promoter or the B ElementTo test whether the specificity of the silencer may be due to selective interference with the islet-specific activity of the B element (mechanism 1), we measured the effects of the silencer on the activity of a variety of other promoters and enhancers, both cellular and viral, containing a wide variety of transcriptional elements. First, the activity of the silencer is not restricted to constructs containing the cognate ELA1 minimal promoter, because it also repressed the activity of the B pentamer coupled to thehsp70 minimal promoter (Fig. 3, compare constructs C andD). Moreover, the silencer inhibited the activity of the 49-bp H-2 Kb cellular enhancer driving either the hsp70 or the ELA1 minimal promoter (85 and 95% reduction, respectively; Fig. 3, compare E with F andH with I). The H-2 Kb enhancer has an exceedingly broad cell-type expression pattern (26Kimura A. Israel A. Le Bail O. Kourilsky P. Cell. 1986; 44: 261-272Abstract Full Text PDF PubMed Scopus (240) Google Scholar) and therefore is likely driven by ubiquitous factors. The silencer region also reduced the activity of the HSV tk promoter by 95% (Fig. 3, J andK) and the SV40 enhancer/promoter by 80% (Fig. 3,L and M). The viral HSV tk promoter contains binding sites for common factors and is active in a wide variety of cell lines (27Jones K.A. Yamamoto K.R. Tjian R. Cell. 1985; 42: 559-572Abstract Full Text PDF PubMed Scopus (391) Google Scholar). The SV40 early enhancer/promoter is also active in a variety of cell types, although individual elements within the enhancer display a unique pattern of cell-specific activity (28Schirm S. Jiricny J. Schaffner W. Genes Dev. 1987; 1: 65-74Crossref PubMed Scopus (111) Google Scholar, 29Ondek B. Shepard A. Herr W. EMBO J. 1987; 6: 1017-1025Crossref PubMed Scopus (95) Google Scholar). These results demonstrated that the silencer region can interfere with the activity of a wide variety of heterologous promoters and enhancers. Consequently, its ability to repress selectively the ELA1enhancer in transgenic mouse islets appears not due to an effect specific to the B element or to PDX1, which binds and mediates the activity of the B element (mechanism 1).Figure 2Repression of B element activity in transfected RIN1046-38 β-cells. The activity of each test gene and the effects of the silencer were measured by transient transfection in RIN38 cells as described under “Experimental Procedures.” The gene fragments in each fusion construct are discussed in the text and are identified as follows:EIp, the ELA1 minimal promoter; B, the B element of the ELA1 enhancer; Sil, theELA1 silencer region in the normal orientation (−501/−202); and liS, the silencer region in the reverse orientation (−202/−501); hGH, the 2.2-kb human growth hormone reporter gene. The nucleotide numberings are relative to the start site of transcription of the gene from which each fragment was derived. Expression was measured by assaying hGH production as described under “Experimental Procedures.” The level of activity of each construct is expressed as a percent of the activity of 5B.EIp.hGH (construct B) after correction for relative transfection efficiencies by monitoring the activity of a cotransfected RSV.mCAT plasmid.View Large Image Figure ViewerDownload Hi-res image Download (PPT)Figure 3Repression of heterologous promoters and enhancers in RIN38 β-cells. The silencer is not specific to the action of the B element of the ELA1enhancer. The gene fragments in each fusion construct are as follows:hsp, the minimal heat shock promoter of hsp70;H-2K b e, the enhancer of the major histocompatibility complex class I H-2K b gene;HSV tkp, the herpes simplex virus thymidine kinase promoter; and SV40p/e, the promoter and enhancer of early gene transcription of simian virus 40. Levels of activity are expressed as a percentage of the activity of 5B.EIp.hGH (construct A). All other designations are as described in the legend for Fig. 2.View Large Image Figure ViewerDownload Hi-res image Download (PPT)The Silencer Is Also Active in Pancreatic Acinar Cell LinesTo test whether the silencer is active in β- but not acinar cells (mechanism 2), we took advantage of the observation that the heterologous enhancers and promoters it can suppress in the RIN38 β-cell line (Fig. 3) are also active in a wide variety of other cell types. 266-6 cells are derived from a mouse pancreatic acinar adenocarcinoma induced by an SV40 T-antigen transgene (15Ornitz D.M. Palmiter R.D. Messing A. Hammer R.E. Pinkert C.A. Brinster R.L. Cold Spring Harbor Symp. Quant. Biol. 1985; 50: 399-409Crossref PubMed Google Scholar); they retain properties of differentiated acinar cells, including the expression of cell-specific genes for the digestive enzymes elastase I, trypsinogen I, and amylase (30Swift G.H. Kruse F. MacDonald R.J. Hammer R.E. Genes Dev. 1989; 3: 687-696Crossref PubMed Scopus (44) Google Scholar). Strikingly, the silencer region inhibited the activity of the cellular H-2 Kb enhancer driving either thehsp70 or the ELA1 minimal promoter in transiently transfected 266-6 acinar cells (Fig. 4, compare C with D and F withG). The silencer also suppressed the activity of the viral HSV tk promoter in this acinar cell line (Fig. 4, compareH and I). When the activity of the silencer with these identical enhancer and promoter constructs are compared for the acinar and β-cell lines, the silencer was at least as effective in the acinar cells. These results suggest that the differential action of the silencer in islet versus acinar cells of mice is due neither to cell-specific activity (mechanism 2) nor to specificity toward the B element (mechanism 1).Figure 4Repression of heterologous promoters and enhancers in 266-6 acinar cells. These data show that the silencer is also active in an acinar cell line derived from transformed pancreatic acinar cells. EIe represents the ELA1gene enhancer. The designations for all other fusion genes are the same as described in the legends for Figs. 1 and 2. Levels of activity are expressed as a percentage of the activity of the ELA1enhancer/promoter construct (A).View Large Image Figure ViewerDownload Hi-res image Download (PPT)Far Upstream Elements Relieve Repression in Acinar but Not Endocrine CellsAlthough the silencer had no apparent effect on the ELA1 gene enhancer in acinar cells of transgenic mice (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar), its ability to repress heterologous promoters in the 266-6 acinar cell line (Fig. 4) led us to test whether it was effective against theELA1 enhancer in these cultured cells. The silencer did indeed repress the activity of the ELA1 enhancer about 90% (Fig. 5, compare B andC). The residual activity in the presence of the silencer was still at least 10-fold greater than that of the ELA1minimal promoter without the enhancer (Fig. 5; compare A andC). Therefore, the silencer appears unable to suppress the activity of the enhancer in the acinar tumor cells as effectively as it does the B element multimer in insulinoma cells. This significant residual acinar activity may account in part for the continued expression of the identical silencer construct (as C of Fig. 5) in the pancreatic acinar cells of transgenic mice (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar). Due to the difficulty in comparing expression levels of different constructs in transgenic animals (e.g. Ref. 30Swift G.H. Kruse F. MacDonald R.J. Hammer R.E. Genes Dev. 1989; 3: 687-696Crossref PubMed Scopus (44) Google Scholar), quantitative effects of the silencer in acinar cells in vivo may not be evident.Figure 5A distant upstream region relieves repression of the ELA1 enhancer in acinar cells but not in β-cells. These transfection data show the activity of the silencer in ELA1 gene constructs in 266-6 acinar cells (top panel) and RIN38 β-cells (bottom panel). The dotted lines in the figure represent a deletion of the silencer region from −501 to −202 and a fusion of the boundary at −502 to the boundary of either the ELA1enhancer or the B element multimer. Levels of activity are expressed as a percent of the activity of the ELA1 enhancer/promoter (construct A) in 266-6 cells and as a percentage of the activity of 5B.EIp.hGH (construct G) in RIN38 cells. All the designations for the gene fragments are described in the legends for previous figures.View Large Image Figure ViewerDownload Hi-res image Download (PPT)To determine whether additional regulatory elements contribute to the control of the endogenous ELA1 gene, we examined the regulatory properties of the region upstream of the silencer. When an additional 4 kb of ELA1 5′-flanking DNA was included with the silencer, enhancer and promoter, the repression by the silencer in 266-6 acinar cells was lost (Fig. 5, compare C andD). However, in the absence of the silencer (Fig. 5 E) the upstream region did not increase the expression from the enhancer. Therefore, the upstream region may interfere directly with the silencer rather than indirectly by increasing the strength of the enhancer and thereby overcoming the influence of the silencer.In contrast to the effects in transfected 266-6 acinar cells, the upstream region could not overcome the action of the silencer in RIN38 β-cells (Fig. 5, compare H and I). Moreover, this upstream region suppressed the activity of the B element in RIN38 cells in the absence of the −501 to −202 silencer region (Fig. 5 J), suggesting the presence of additional silencing activity in the upstream region that is not manifested in 266-6 acinar cells (Fig. 5 D). In summary, the region 4 kb upstream of the silencer contains two classes of regulatory elements that help confer cell type-specific expression, additional negative control elements that suppress β-cell expression and others that interfere with the action of the silencer in acinar cells. The overall effect of the upstream region is to reinforce expression in acinar cells and to suppress it in β-cells.The Silencer Is Ineffective with Strong Enhancer/Promoter RegionsAn apparent discrepancy remains for the selective action of the 300-bp silencer in animals compared with cultured cells.In vitro the silencer is neither specific for theELA1 B element, because it acts on heterologous promoters and enhancers (Fig. 3), nor for β-cells, because it also acts in acinar cells (Figs. 4 and 5). Although other regulatory sequences in the endogenous locus may complement the action of the silencer (Fig. 5), in transgenic animals the 300-bp silencer effectively silenced theELA1 enhancer in islet cells while acinar activity was maintained (10Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Genes Dev. 1993; 7: 774-786Crossref PubMed Scopus (36) Google Scholar). There are at least three possible explanations for this discrepancy. First, the RIN38 β-cell and 266-6 acinar cell lines may not accurately reproduce the differential activity of the silencer in β-cells and acinar cells in situ. Because islet and acinar cells have a common developmental origin, retro-differentiation of the acinar cell line may acquire some endocrine properties, including β-cell proteins for the silencer. Second, the silencer may also decrease the activity of the ELA1 enhancer in the acinar cells of animals but not completely; because of the difficulty of accurately quantifying transgenic expression in mice, this effect may have been overlooked.A third possibility is a previously unsuspected basis for the differential action of a silencer. In transfected RIN38 cells the silencer is able to repress the B element pentamer activity completely to the level of the ELA1 minimal promoter (Fig. 2), whereas in acinar cells it is unable to repress the ELA1 enhancer completely to the level of the same minimal promoter (Fig. 5). The post-repression activity of the −500EIhGH construct in acinar cells is at least 10-fold greater than that of the ELA1 minimal promoter (Fig. 5, construct C). Thus, the silencer is only partially effective on the three-element ELA1 enhancer but is much more effective on the synthetic B element multimer, which is a much less potent enhancer. This raises the possibility that the effectiveness of the silencer may depend on the strength of the enhancer with which it is paired. Thus, the 300-bp silencer may be able to suppress the action of a weak enhancer completely but only partially suppress a stronger enhancer.To test whether the effectiveness of the silencer depends on the strength of the paired enhancer/promoter, we measured the ability of the silencer to repress the activity of the potent RSV and CMV promoters in both acinar and β-cell lines. The silencer had no significant effect on the activity of these extremely strong promoters in either cell type (Fig. 6), consistent with the notion that the effectiveness of the silencer is dependent on the strength of the positively activating control sequences it antagonizes. The ability to affect weak but not strong promoters is also a property of a silencer from the chicken lysozyme gene (31Baniahmad A. Muller M. Steiner C. Renkawitz R. EMBO J. 1987; 6: 2297-2303Crossref PubMed Scopus (155) Google Scholar). Because only the B and C elements of the ELA1 enhancer are active in β-cells, whereas all three elements are active in acinar cells (13Kruse F. Rose S.D. Swift G.H. Hammer R.E. MacDonald R.J. Mol. Cell. Biol. 1995; 15: 4385-4394Crossref PubMed Scopus (35) Google Scholar, 32Kruse F. Komro C.T. Michnoff C.H. MacDonald R.J. Mol. Cell. Biol. 1988; 8: 893-902Crossref PubMed Scopus (31) Google Scholar), the enhancer may be more susceptible to the silencer in β-cells.Figure 6Lack of repression of the potent RSV and CMV promoters in both acinar and islet cells. RSV and CMV LTR represent the long terminal repeats containing the promoter/enhancer region of the early genes of the Rous sarcoma virus and the cytomegalovirus, respectively. The activity of the various constructs are expressed relative to the activity of theELA1 promoter (EIp.hGH), which was arbitrarily designated as 1.View Large Image Figure ViewerDownload Hi-res image Download (PPT)Binary Action of the SilencerMany enhancers appear to affect transcription by increasing the probability of a promoter being active without increasing the rate of initiation at the promoter (6Fie" @default.
• W2094663606 created "2016-06-24" @default.
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• W2094663606 creator A5053609372 @default.
• W2094663606 creator A5061637831 @default.
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• W2094663606 date "2000-12-01" @default.
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• W2094663606 title "A Binary Mechanism for the Selective Action of a Pancreatic β-Cell Transcriptional Silencer" @default.
• W2094663606 cites W122840240 @default.
• W2094663606 cites W126863029 @default.
• W2094663606 cites W1480830771 @default.
• W2094663606 cites W1775713904 @default.
• W2094663606 cites W1846058178 @default.
• W2094663606 cites W1912127524 @default.
• W2094663606 cites W1954861259 @default.
• W2094663606 cites W1964858195 @default.
• W2094663606 cites W1965372133 @default.
• W2094663606 cites W1972042181 @default.
• W2094663606 cites W1974582630 @default.
• W2094663606 cites W1980137584 @default.
• W2094663606 cites W1981765089 @default.
• W2094663606 cites W1985300178 @default.
• W2094663606 cites W2007918998 @default.
• W2094663606 cites W2013220929 @default.
• W2094663606 cites W2018403838 @default.
• W2094663606 cites W2026548183 @default.
• W2094663606 cites W2028214195 @default.
• W2094663606 cites W2030211226 @default.
• W2094663606 cites W2034279963 @default.
• W2094663606 cites W2036017389 @default.
• W2094663606 cites W2037164528 @default.
• W2094663606 cites W205061201 @default.
• W2094663606 cites W2056799965 @default.
• W2094663606 cites W2057073066 @default.
• W2094663606 cites W2057564306 @default.
• W2094663606 cites W2062408930 @default.
• W2094663606 cites W2072881211 @default.
• W2094663606 cites W2087181226 @default.
• W2094663606 cites W2090286338 @default.
• W2094663606 cites W2090610011 @default.
• W2094663606 cites W2095115211 @default.
• W2094663606 cites W2097948853 @default.
• W2094663606 cites W2105922555 @default.
• W2094663606 cites W2106900373 @default.
• W2094663606 cites W2119323082 @default.
• W2094663606 cites W2129754384 @default.
• W2094663606 cites W2138096293 @default.
• W2094663606 cites W2145337373 @default.
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