Histone proteins and the nucleosomes they form with DNA are the fundamental building blocks of eukaryotic chromatin. A diverse array of post-translational modifications that often occur on tail domains of these proteins has been well documented. Although the function of these highly conserved modifications has remained elusive, converging biochemical and genetic evidence suggests functions in several chromatin-based processes. We propose that distinct histone modifications, on one or more tails, act sequentially or in combination to form a 'histone code' that is, read by other proteins to bring about distinct downstream events.

The language of covalent histone modifications.

Genetically identical cells exposed to the same environmental conditions can show significant variation in molecular content and marked differences in phenotypic characteristics. This variability is linked to stochasticity in gene expression, which is generally viewed as having detrimental effects on cellular function with potential implications for disease. However, stochasticity in gene expression can also be advantageous. It can provide the flexibility needed by cells to adapt to fluctuating environments or respond to sudden stresses, and a mechanism by which population heterogeneity can be established during cellular differentiation and development.

/pdf/stochasticity-in-gene-expression-from-theories-to-phenotypes-32urfuz0k3.pdf

Stochasticity in gene expression: from theories to phenotypes

https://www.cell.com/cell/pdf/S0092-8674(00)80205-6.pdf

Core structure of GP41 from the HIV envelope glycoprotein

The state of chromatin (the packaging of DNA in eukaryotes) has long been recognized to have major effects on levels of gene expression, and numerous chromatin-altering strategies-including ATP-dependent remodeling and histone modification-are employed in the cell to bring about transcriptional regulation. Of these, histone acetylation is one of the best characterized, as recent years have seen the identification and further study of many histone acetyltransferase (HAT) proteins and their associated complexes. Interestingly, most of these proteins were previously shown to have coactivator or other transcription-related functions. Confirmed and putative HAT proteins have been identified from various organisms from yeast to humans, and they include Gcn5-related N-acetyltransferase (GNAT) superfamily members Gcn5, PCAF, Elp3, Hpa2, and Hat1: MYST proteins Sas2, Sas3, Esa1, MOF, Tip60, MOZ, MORF, and HBO1; global coactivators p300 and CREB-binding protein; nuclear receptor coactivators SRC-1, ACTR, and TIF2; TATA-binding protein-associated factor TAF(II)250 and its homologs; and subunits of RNA polymerase III general factor TFIIIC. The acetylation and transcriptional functions of these HATs and the native complexes containing them (such as yeast SAGA, NuA4, and possibly analogous human complexes) are discussed. In addition, some of these HATs are also known to modify certain nonhistone transcription-related proteins, including high-mobility-group chromatin proteins, activators such as p53, coactivators, and general factors. Thus, we also detail these known factor acetyltransferase (FAT) substrates and the demonstrated or potential roles of their acetylation in transcriptional processes.

Acetylation of Histones and Transcription-Related Factors

Developmental progression is driven by specific spatiotemporal domains of gene expression, which give rise to stereotypically patterned embryos even in the presence of environmental and genetic variation. Views of how transcription factors regulate gene expression are changing owing to recent genome-wide studies of transcription factor binding and RNA expression. Such studies reveal patterns that, at first glance, seem to contrast with the robustness of the developmental processes they encode. Here, we review our current knowledge of transcription factor function from genomic and genetic studies and discuss how different strategies, including extensive cooperative regulation (both direct and indirect), progressive priming of regulatory elements, and the integration of activities from multiple enhancers, confer specificity and robustness to transcriptional regulation during development.

http://materiais.dbio.uevora.pt/BD/Crescimento/TFs_2012.pdf

Transcription factors: from enhancer binding to developmental control.

We have purified distinct complexes of nine to 12 proteins [referred to as BRG1-associated factors (BAFs)] from several mammalian cell lines using an antibody to the SWI2-SNF2 homolog BRG1. Microsequencing revealed that the 47 kDa BAF is identical to INI1. Previously INI1 has been shown to interact with and activate human immunodeficiency virus integrase and to be homologous to the yeast SNF5 gene. A group of BAF47-associated proteins were affinity purified with antibodies against INI1/BAF47 and were found to be identical to those co-purified with BRG1, strongly indicating that this group of proteins associates tightly and is likely to be the mammalian equivalent of the yeast SWI-SNF complex. Complexes containing BRG1 can disrupt nucleosomes and facilitate the binding of GAL4-VP16 to a nucleosomal template similar to the yeast SWI-SNF complex. Purification of the complex from several cell lines demonstrates that it is heterogeneous with respect to subunit composition. The two SWI-SNF2 homologs, BRG1 and hbrm, were found in separate complexes. Certain cell lines completely lack BRG1 and hbrm, indicating that they are not essential for cell viability and that the mammalian SWI-SNF complex may be tailored to the needs of a differentiated cell type.

Purification and biochemical heterogeneity of the mammalian SWI-SNF complex.

EUKARYOTIC transcriptional activators mediate transcriptional induction through stabilization of the preinitiation complex1–4, probably through direct interactions with basal transcription factors5–7. In vitro studies on the role of an activator in the maintenance of on-going transcription (reinitiation) have been contradictory, suggesting that, after formation of a preinitiation complex, an activator may8 or may not9 be necessary for transcription to be maintained. We have developed a means of regulating transcription in living cells through the use of both homodimeric and heterodimerizing synthetic ligands that allow the ligand-dependent association and disassociation of a transcriptional activation domain with a promoter. Here we report that maintaining the transcription of endogenous genes in vivo, in both yeast and human cells, requires the continuous presence of the activation domain. The use of synthetic ligands as a transcriptional on–off switch represents a powerful means of controlling the transcription in vitro and in vivo for both experimental and therapeutic purposes.

Dimeric ligands define a role for transcriptional activation domains in reinitiation

Transcriptional control is generally thought to operate as a binary switch, a behavior that might explain observations such as monoallelic gene expression, stochastic phenotypic changes and bimodal gene activation kinetics. By measuring the activity of the single-copy GAL1 promoter in single cells, we found that changes in the activities of either the transcriptional activator, Gal4 (by simple recruitment with synthetic ligands), or the transcriptional repressor, Mig1, generated graded (non-binary) changes in gene expression that were proportional to signal intensity. However, in the context of the endogenous glucose-responsive signaling pathway, these transcription factors formed part of a binary transcriptional response. Genetic studies demonstrated that this binary response resulted from regulation of a second repressor, Gal80, whereas regulation of Mig1 by a distinct signaling pathway generated graded changes in GAL1 promoter activity. Surprisingly, isogenetic cells can respond to glucose with either binary or graded changes in gene expression, depending on growth conditions. Our studies demonstrate that a given promoter can adapt either binary or graded behavior, and identify the Mig1 and Gal80 genes as necessary for binary versus graded behavior of the Gal1 promoter.

/pdf/cell-signaling-can-direct-either-binary-or-graded-qkf7bw4f67.pdf

Cell signaling can direct either binary or graded transcriptional responses

Chromatin presents a significant obstacle to transcription, but two means of overcoming its repressive effects, histone acetylation and the activities of the Swi-Snf complex, have been proposed. Histone acetylation and Swi-Snf activity have been shown to be crucial for transcriptional induction and to facilitate binding of transcription factors to DNA. By regulating the activity of the Swi-Snf complex in vivo, we found that active transcription requires continuous Swi-Snf function, demonstrating a role for this complex beyond the induction of transcription. Despite the presumably generalized packaging of genes into chromatin, previous studies have indicated that the transcriptional requirements for the histone acetyltransferase, Gcn5, and the Swi-Snf complex are limited to a handful of genes. However, inactivating Swi-Snf function in cells also lacking GCN5 revealed defects in transcription of several genes previously thought to be SWI-SNF- and GCN5-independent. These findings suggest that chromatin remodeling plays a widespread role in gene expression and that these two chromatin remodeling activities perform independent and overlapping functions during transcriptional activation.

Stephen R. Biggar

Papers

Purification and biochemical heterogeneity of the mammalian SWI-SNF complex.

Dimeric ligands define a role for transcriptional activation domains in reinitiation

Cell signaling can direct either binary or graded transcriptional responses

Continuous and widespread roles for the Swi-Snf complex in transcription.

Small molecule-dependent genetic selection in stochastic nanodroplets as a means of detecting protein-ligand interactions on a large scale