Metastable Macromolecular Complexes Containing High Mobility Group Nucleosome-binding Chromosomal Proteins in HeLa Nuclei

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The interaction of architectural proteins such as the linker histone H1 and high-mobility-group (HMG) proteins with nucleosomes leads to changes in chromatin structure and histone modifications and alters the cellular transcription profile. The interaction of HMG proteins with chromatin is dynamic. However, it is not clear whether the proteins are constantly and randomly redistributed among all the nucleosomes or whether they preferentially associate with, and turn over at, specific regions in chromatin. To address this question, we examined the genomewide distribution of the nucleosome binding protein HMGN1 and compared it to that of regulatory chromatin marks. We find that HMGN1 is not randomly distributed throughout the genome. Instead, the protein preferentially localizes to DNase I hypersensitive (HS) sites, promoters, functional enhancers, and transcription factor binding sites. Our results suggest that HMGN1 is part of the cellular machinery that modulates transcriptional fidelity by generating, maintaining, or preferentially interacting with specific sites in chromatin.

Section: Discussionsupporting

confidence: 69%

Genomic Profiling of HMGN1 Reveals an Association with Chromatin at Regulatory Regions

Cuddapah

Schones

Cui

et al. 2011

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“…One possibility is that the major chromatin binding sites of each family localize to distinct nuclear compartments or to unique chromatin domains. Another possibility is that each type of HMG protein recruits specific protein partners that target the HMG protein to unique chromatin regions (38,49). Finally, since histone H1 has a tripartite structure (7,13) and binds to nucleosomes through multiple sites (57,61), it is conceivable that each HMG family affects a distinct set of H1 nucleosome binding sites.…”

Section: Discussionmentioning

confidence: 99%

Network of Dynamic Interactions between Histone H1 and High-Mobility-Group Proteins in Chromatin

Catez

Yang

Tracey

et al. 2004

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246

262

Histone H1 and the high-mobility group (HMG) proteins are chromatin binding proteins that regulate gene expression by modulating the compactness of the chromatin fiber and affecting the ability of regulatory factors to access their nucleosomal targets. Histone H1 stabilizes the higher-order chromatin structure and decreases nucleosomal access, while the HMG proteins decrease the compactness of the chromatin fiber and enhance the accessibility of chromatin targets to regulatory factors. Here we show that in living cells, each of the three families of HMG proteins weakens the binding of H1 to nucleosomes by dynamically competing for chromatin binding sites. The HMG families weaken H1 binding synergistically and do not compete among each other, suggesting that they affect distinct H1 binding sites. We suggest that a network of dynamic and competitive interactions involving HMG proteins and H1, and perhaps other structural proteins, constantly modulates nucleosome accessibility and the local structure of the chromatin fiber.The chromatin fiber is a dynamic, malleable structure that is targeted by numerous regulatory factors that modify the histones and the DNA and remodel the structure of the nucleosome. The dynamics of the chromatin fiber reflect the combined action of numerous chromatin modifiers, including architectural proteins such as histone H1 and members of the high-mobility group (HMG) protein superfamily. The interaction of histone H1 with nucleosomes stabilizes the higherorder, compact chromatin structure (57, 61), thereby restricting the ability of regulatory factors, nucleosome remodeling complexes, and histone modifiers to access their chromatin binding sites (17,27,28,30,32,34). Loss of H1 results in both up regulation and down regulation of specific gene expression (2,26,53,56), suggesting that the protein affects the action of both positive and negative transcriptional regulators. Experiments with H1 knockout mice demonstrate the existence of cellular mechanisms that strive to maintain a constant level of H1 and that reduction of H1 beyond a critical point is not compatible with normal embryonic development (21). Thus, factors that modulate the interaction of H1 with nucleosomes may affect a variety of chromatin-related processes and participate in genetic regulatory mechanisms.The HMG superfamily is composed of three families: HMGB, HMGA, and HMGN, each family being characterized by a distinct DNA or chromatin binding motif (12). These non-histones decompact the higher-order chromatin structure and promote the binding of nuclear regulatory factors (1,12,49,58). Footprinting analysis and in vitro binding assays (29,64) showed that the chromatin binding sites of the HMG proteins are similar to those of H1 and suggested that HMG proteins and H1 compete for the same binding sites, and recent photobleaching experiments demonstrated that HMGN proteins affect the binding of H1 to chromatin in living cells (16). Photobleaching techniques like fluorescence recovery after photobleaching (FRAP), which is used...

“…However, the organization of HMGN proteins is highly dynamic in live cells, and their association with any specific nucleosome is temporary (37). It is conceivable that HMGNs are targeted to specific regions by their association with other nuclear proteins, and indeed, biochemical studies suggest that HMGN proteins form multiple metastable complexes with a number of as-yet-unidentified nuclear proteins (29).…”

mentioning

confidence: 99%

Chromosomal Proteins HMGN3a and HMGN3b Regulate the Expression of Glycine Transporter 1

West

Castellini

Duncan

et al. 2004

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HMGN proteins promote chromatin unfolding, enhance access to nucleosomes, and modulate transcription from chromatin templates. It is not known whether they act indiscriminately as general modulators of transcription or whether they regulate specific gene expression. Here, we investigated the role of HMGN3, a recently discovered HMGN family member, in transcription in vivo. We created cell lines overexpressing HMGN3a or its splice variant, HMGN3b, and analyzed their gene expression profiles using microarrays and reverse transcriptase PCR. We found that ectopic expression of HMGN3a alters the expression of approximately 0.8% of genes. Both HMGN3a and HMGN3b upregulate the expression of the glycine transporter 1 gene (Glyt1). Glyt1 encodes a membrane transporter that regulates the glycine concentration in synaptic junctions. Both GLYT1 and HMGN3 are highly expressed in glia cells and the eye, and we show that both proteins are coexpressed in the retina. Chromatin immunoprecipitation assays showed that HMGN3 protein is recruited to a region of the Glyt1 gene encompassing the Glyt1a transcriptional start site. These results suggest that HMGN3 regulates Glyt1 expression and demonstrate that members of the HMGN family can regulate the transcription of specific genes.In eukaryotes, all of the DNA is complexed with histone proteins and packaged into a highly folded, well-ordered, and dynamic structure called chromatin. This packaging modulates the ability of regulatory factors to access their DNA targets and plays a major role in regulating various nuclear activities, including transcription (18,48). Chromatin folding is modulated by numerous nuclear factors including nucleosome remodeling complexes, histone-modifying enzymes, and architectural proteins such as linker histones and HMG proteins. Members of the HMG superfamily interact with chromatin and DNA and affect a wide range of DNA-dependent activities such as transcription, replication, and recombination (9).One of the HMG families, the HMGN family, is comprised of small, basic proteins that bind specifically to nucleosomes (8). HMGN proteins are highly conserved and found only in vertebrates. The two founding members of the family, HMGN1 and HMGN2 (formerly named HMG-14 and HMG-17) (8), have been studied extensively. They contain a highly conserved nucleosome binding domain, a bipartite nuclear localization signal, and a C-terminal chromatin-unfolding domain (12, 47). When incorporated into minichromosomes, HMGN proteins confer a more open chromatin structure that is more sensitive to nucleases and that is transcribed and replicated more efficiently (13,14,35,46,50). Their ability to unfold chromatin also enhances the rate of DNA repair, as recently demonstrated in mice lacking HMGN1 (5).Although HMGN proteins display little or no DNA sequence specificity when binding to nucleosomes (45), several lines of evidence indicate that HMGN binding within the nucleus is nonrandom. Immunofluorescence studies have shown that HMGN proteins are localized in many foci within t...