Characterization of a “silencer” in yeast: A DNA sequence with properties opposite to those of a transcriptional enhancer

Gene regulation involves long-range communication between silencers, enhancers, and promoters. In Saccharomyces cerevisiae, silencers flank transcriptionally repressed genes to mediate regional silencing. Silencers recruit the Sir proteins, which then spread along chromatin to encompass the entire silenced domain. In this report we have employed a boundary trap assay, an enhancer activity assay, chromatin immunoprecipitations, and chromosome conformation capture analyses to demonstrate that the two HMR silencer elements are in close proximity and functionally communicate with one another in vivo. We further show that silencing is necessary for these long-range interactions, and we present models for Sir-mediated silencing based upon these results.Gene activation and gene repression are central to the proper development and differentiation of organisms. DNA elements such as promoters, enhancers, and silencers play a central role in eukaryotic gene regulation. These elements are separated from each other by several kilobase pairs of DNA but are able to communicate with one another to regulate the activation or repression of genes. The exact mechanism by which distally located elements communicate with one another is not clear and is one of the key questions in gene regulation. Long-range communication between distantly located elements in chromosomes is thought to occur by one of two principal mechanisms (8). One class of models postulate that a signal emanating from a distal regulatory element spreads along the DNA fiber until it encounters a proximal regulatory element. A second class of models postulate that distal and proximal regulatory elements interact with one another directly, with the intervening DNA forming a loop. Both mechanisms must function within the context of the global chromosome structure, which appears to be composed of large chromosome loops that attach to a proteinaceous superstructure (11). The nucleus appears to be divided further into distinct chromatin compartments, with heterochromatic domains being present in regions near the nuclear periphery while euchromatic domains are found mainly in the interior of the nucleus, although a significant portion of euchromatin is located near nuclear pores.It has been suggested that the functionally and structurally defined chromatin domains may be coincident (36). Enhancers and locus control regions (LCRs) are long-range regulatory elements that activate promoters in a distance-and orientation-independent manner, and recent studies indicate that enhancers and LCRs often cluster together in three-dimensional space to form an "active chromatin hub" (23,49,58,59).Similarly, in yeast the promoters and terminators of genes are in close proximity to one another (3, 48) and tethered to the nuclear pore (10, 52). The consequence of this spatial organization is that the DNA between these regulatory elements is looped out. It is thought that the formation of these nuclear substructures aids in transcription activation.Silencers are negative regulatory element...

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“…does not recruit Sir proteins in the absence of HMR-E and are consistent with previous functional results (1,7,50).…”

Section: Resultssupporting

confidence: 93%

Long-Range Communication between the Silencers of HMR

Valenzuela

Dhillon

Dubey

et al. 2008

Molecular and Cellular Biology

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“…First, the fragments behaved similarly to one another. Second, rather than enhancing the activity of a downstream promoter, the fragments negatively regulated both the homologous HPV-6 E6 and a heterologous SV40 early promoter in a position-and orientation-independent manner, thus fulfilling the definition of a silencer (Brand et al, 1985). Silencer activity was detected in HeLa cells, C33A cells and primary human keratinocytes.…”

Section: Introductionmentioning

confidence: 92%

Detection of silencer activity in the long control regions of human papillomavirus type 6 isolated from both benign and malignant lesions

Farr

Pattison

Youn

et al. 1995

Journal of General Virology

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Human papillomavirus type 6 (HPV-6) DNA is the predominant HPV type found in condyloma acuminata: it is rarely found in carcinomas. We have previously reported cloning and characterizing an HPV-6 from a vulvar condyloma (HPV6-W50) and an HPV-6 from a vulvar carcinoma (HPV6-T70). The E5, E6 and E7 proteins encoded by the two genomes were identical, however, the two genomes differed in the long control region (LCR). Cloning of the entire LCR into the enhancerless plasmid pSVEcat showed that the two LCRs had comparable enhancer activity. Since the major differences between the two LCRs resided in the 5' end of the LCR, upstream of the L1 polyadenylation signal, we subcloned the two LCRs to analyse more closely their effect on cat gene expression. The data indicated that LCR subclones of the two genomes had comparable chloramphenicol acetyltransferase (CAT) activity. A negative regulatory region was detectable when the test plasmids were transfected into HeLa and C33A cells and in primary keratinocytes. A decrease in CAT activity was also detected when the SV40 early promoter was replaced with the putative HPV-6 E6 promoter. The negative regulatory region functioned in a position-and orientation-independent manner, thus fulfilling the definition of a silencer.

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“…Silencers were initially defined as sequence elements which are capable of repressing promoter activity in an orientation-and position-independent fashion, in the context of a native or a heterologous promoter [80]. However, in practice, a silencer is now generally considered to be a short, specific sequence of nucleotides which are located in the 5h upstream promoter region of a given gene.…”

Section: Silencers and Transcriptional Repressionmentioning

confidence: 99%

Transcriptional control and the role of silencers in transcriptional regulation in eukaryotes

Ogbourne

Antalis

1998

Biochemical Journal

227

171

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Mechanisms controlling transcription and its regulation are fundamental to our understanding of molecular biology and, ultimately, cellular biology. Our knowledge of transcription initiation and integral factors such as RNA polymerase is considerable, and more recently our understanding of the involvement of enhancers and complexes such as holoenzyme and mediator has increased dramatically. However, an understanding of transcriptional repression is also essential for a complete understanding of promoter structure and the regulation of gene expression. Transcriptional repression in eukaryotes is achieved through ‘silencers ’, of which there are two types, namely ‘silencer elements ’ and ‘negative regulatory elements ’ (NREs). Silencer elements are classical, position-independent elements that direct an active repression mechanism, and NREs are position-dependent elements that direct a passive repression mechanism. In addition, ‘repressors ’ are DNA-binding trasncription factors that interact directly with silencers. A review of the recent literature reveals that it is the silencer itself and its context within a given promoter, rather than the interacting repressor, that determines the mechanism of repression. Silencers form an intrinsic part of many eukaryotic promoters and, consequently, knowledge of their interactive role with enchancers and other transcriptional elements is essential for our understanding of gene regulation in eukaryotes.

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Characterization of a “silencer” in yeast: A DNA sequence with properties opposite to those of a transcriptional enhancer

Cited by 510 publications

References 34 publications

Long-Range Communication between the Silencers of HMR

Long-Range Communication between the Silencers of HMR

Detection of silencer activity in the long control regions of human papillomavirus type 6 isolated from both benign and malignant lesions

Transcriptional control and the role of silencers in transcriptional regulation in eukaryotes

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