HMG box proteins bind to four-way DNA junctions in their open conformation

Pohler, J Richard G; Norman, D.; Bramham, Janice; Bianchi, Marco E.; Lilley, David M.J.

doi:10.1093/emboj/17.3.817

Cited by 99 publications

(84 citation statements)

References 49 publications

(78 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Concentrations of this peptide needed for binding were about the same as published values for the HMG1 B-domain when tagged with maltose-binding protein (24 …”

Section: Resultssupporting

confidence: 55%

“…As illustrated in Fig. 3a, the 4WJDNA can take on either an X conformation or a more open crosslike conformation dependent on the presence or absence of Mg 2ϩ ions (24).…”

Section: Resultsmentioning

confidence: 99%

“…HMG1 (24,33) as both proteins bound only the open, extended conformation of 4WJDNA. This Sin1p/Spt2p peptide has a cluster of positively charged amino acids (RGKKR).…”

Section: Sin1p/spt2p Domains Bind 4wjdnamentioning

confidence: 99%

See 2 more Smart Citations

Functional Domains of the Yeast Chromatin Protein Sin1p/Spt2p Can Bind Four-way Junction and Crossing DNA Structures

Novoseler

Hershkovits

Katcoff

2005

Journal of Biological Chemistry

View full text Add to dashboard Cite

Sin1p/Spt2p is a yeast chromatin protein that, when mutated or deleted, alters the transcription of a family of genes presumably by modulating local chromatin structure. In this study, we investigated the ability of different domains of this protein to bind four-way junction DNA (4WJDNA) since 4WJDNA can serve as a model for bent double helical DNA and for the crossed structure formed at the exit and entry of DNA to the nucleosomes. Sequence alignment of Sin1p/Spt2p homologues from 11 different yeast species showed conservation of several domains. We found that three domains of Sin1p/ Spt2p fused to glutathione S-transferase can each bind independently in a structure-specific manner to 4WJDNA as measured in a gel mobility shift assay. A feature common to these domains is a cluster of positively charged amino acids. Modification of this cluster resulted in either abolishment of binding or a change in the binding properties. One of the domains tested clearly bound superhelical DNA, although it failed to induce bending in a circularization assay. Poly-L-lysine, which may be viewed as a cluster of positively charged amino acids, bound 4WJDNA as well. Phenotypic analysis showed that disruption of any of these domains resulted in suppression of a his4-912␦ allele, indicating that each domain has functional significance. We propose that Sin1p/Spt2p is likely to modulate local chromatin structure by binding two strands of double-stranded DNA at their crossover point.Sin1p/Spt2p is a yeast chromatin non-histone protein. While the precise function of the protein is still unknown, it is known to function as a negative transcriptional regulator of a number of genes including SUC2 (1), INO1 (2), and SSA3 (3). Activity of the HO promoter, as measured from an HO promoter driving a lacZ gene (4), is also regulated by SIN1/SPT2. swi1, swi2, and swi3 mutants are unable to transcribe RNA from the HO promoter. However, in sin1/swi double mutants, transcription is restored. Mutants in SPT2 were first identified as suppressors of ty and ␦ insertions in the 5Ј non-coding region of the HIS4 gene (5). As the negative regulation of these genes is overcome by the SW1/SNF chromatin remodeling complex, it was suggested that a function of SIN1p/SPT2p is to somehow maintain chromatin compaction at specific locations in the chromatin.

show abstract

“…Concentrations of this peptide needed for binding were about the same as published values for the HMG1 B-domain when tagged with maltose-binding protein (24 …”

Section: Resultssupporting

confidence: 55%

“…As illustrated in Fig. 3a, the 4WJDNA can take on either an X conformation or a more open crosslike conformation dependent on the presence or absence of Mg 2ϩ ions (24).…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Functional Domains of the Yeast Chromatin Protein Sin1p/Spt2p Can Bind Four-way Junction and Crossing DNA Structures

Novoseler

Hershkovits

Katcoff

2005

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Holliday junctions are also targets for many architectural and regulatory proteins such as histones H1 and H5, topoisomerase IIbeta, HMG proteins, HU, p53, proto-oncogene protein DEK to mention a few (7)(8)(9)(10)(11)(12)(13)(14)(15). The biological role for some of these interactions is still unclear.…”

mentioning

confidence: 99%

Effect of DNA Supercoiling on the Geometry of Holliday Junctions

2006

View full text Add to dashboard Cite

Unusual DNA conformations including cruciforms play an important role in gene regulation and various DNA transactions. Cruciforms are also the models for Holliday junctions, the transient DNA conformations critically involved in DNA homologous and site-specific recombination, repair and replication. Although the conformations of immobile Holliday junctions in linear DNA molecules have been analyzed with use of various techniques, the role of DNA supercoiling has not been studied systematically. We utilized Atomic Force Microscopy (AFM) to visualize cruciform geometry in plasmid DNA with different superhelical densities at various ionic conditions. Both folded and unfolded conformations of the cruciform were identified, and the data showed that DNA supercoiling shifts the equilibrium between folded and unfolded conformations of the cruciform towards the folded one. In topoisomers with low superhelical density the population of folded conformation is 50 to 80 %, depending on ionic strength of the buffer and a type of cation added, whereas in the sample with high superhelical density, this population is as high as 98-100%. The time-lapse studies in aqueous solutions allowed us to observe the conformational transition of the cruciform directly. The time-dependent dynamics of the cruciform correlates with the structural changes revealed by the ensemble-averaged analysis of dry samples. Altogether, the data obtained show directly that DNA supercoiling is the major factor determining the Holliday junction conformation.DNA cruciforms play an important role in regulation of biological processes involving DNA. These structures are formed by inverted repeats and require DNA supercoiling for their stable existence (1). Inverted repeats are associated with origin of replication (1-3). Cruciforms are suggested to be involved in the regulation of gene expression (1,4), and may also play role in nucleosome structure and function (5). In addition, the cruciform is inherent model for Holliday junction, an intermediate in homologous and site-specific recombination (reviewed in (6) (reviewed in (6,14,16)). According to these studies, it can be summarized that the Holiday junction adopts two distinct conformations: folded and unfolded. Unfolded conformation, in which adjacent arms are nearly perpendicular to each other and the structure, has a 4-fold symmetry, and exists at low concentration of metal ions. Folded conformation (or stacked, X-type), in which four arms undergo pairwise coaxial stacking, could be modeled with two duplexes exchanging strands at the junction point. This conformation is stabilized by high ionic strength divalent or polyvalent cations in particular (e.g. Mg2+ cations at the concentration more then 100 μM). Sodium cations also shift equilibrium distribution towards folded conformation though much higher concentrations of cations are required (17). Also, folded conformation of cruciform can be parallel or antiparallel. Synthetic immobile DNA junction exists in antiparallel geometry and no parallel g...

show abstract

“…Like 59 protein, some HMG proteins bind cruciform DNA (reviewed in Refs. [25][26][27][28]. The surface of 59 protein has a high density of hydrophobic and basic residues that may be involved in DNA and protein binding, but the binding sites are not obvious from the structure.…”

mentioning

confidence: 99%

Mutations of Bacteriophage T4 59 Helicase Loader Defective in Binding Fork DNA and in Interactions with T4 32 Single-stranded DNA-binding Protein

Jones

Green

Stephens

et al. 2004

Journal of Biological Chemistry

View full text Add to dashboard Cite

Bacteriophage T4 gene 59 protein greatly stimulates the loading of the T4 gene 41 helicase in vitro and is required for recombination and recombination-dependent DNA replication in vivo. 59 protein binds preferentially to forked DNA and interacts directly with the T4 41 helicase and gene 32 single-stranded DNA-binding protein. The helicase loader is an almost completely ␣-helical, two-domain protein, whose N-terminal domain has strong structural similarity to the DNA-binding domains of high mobility group proteins. We have previously speculated that this high mobility group-like region may bind the duplex ahead of the fork, with the C-terminal domain providing separate binding sites for the fork arms and at least part of the docking area for the helicase and 32 protein. Here, we characterize several mutants of 59 protein in an initial effort to test this model. We find that the I87A mutation, at the position where the fork arms would separate in the model, is defective in binding fork DNA. As a consequence, it is defective in stimulating both unwinding by the helicase and replication by the T4 system. 59 protein with a deletion of the two C-terminal residues, Lys 216 and Tyr 217 , binds fork DNA normally. In contrast to the wild type, the deletion protein fails to promote binding of 32 protein on short fork DNA. However, it binds 32 protein in the absence of DNA. The deletion is also somewhat defective in stimulating unwinding of fork DNA by the helicase and replication by the T4 system. We suggest that the absence of the two terminal residues may alter the configuration of the lagging strand fork arm on the surface of the C-terminal domain, so that it is a poorer docking site for the helicase and 32 protein.Bacteriophage T4 DNA replication begins by synthesis from one of several origins in the early stage of infection, but replication from forks created on recombination intermediates becomes the predominant replication initiation mechanism at later times (1, 2). Recombination-dependent replication is made possible by the terminally redundant and circularly permuted arrangement of the 168-kb linear T4 DNA genome. Thus, the end of one molecule can invade the homologous internal region of another molecule. Phage T4 encodes the replication proteins that are needed for both modes of replication (reviewed in Refs. 3 and 4).In the T4 replication system, gene 41 helicase unwinds the duplex ahead of the leading strand T4 DNA polymerase and associates with the gene 61 primase to enable it to make the pentamer primers that initiate each lagging strand fragment. In vitro, the helicase by itself loads slowly on replication forks, but its loading is greatly stimulated by the gene 59 helicase loading protein (5). In vivo, the helicase loading protein is essential for recombination and recombination-dependent replication (1, 2). Both 41 helicase and its 59 loader are needed for polar branch migration on joint molecules formed by the T4 UvsX recombinase and gene 32 single-stranded DNA-binding protein (6), and the helicase, 59 l...

show abstract

HMG box proteins bind to four-way DNA junctions in their open conformation

Cited by 99 publications

References 49 publications

Functional Domains of the Yeast Chromatin Protein Sin1p/Spt2p Can Bind Four-way Junction and Crossing DNA Structures

Functional Domains of the Yeast Chromatin Protein Sin1p/Spt2p Can Bind Four-way Junction and Crossing DNA Structures

Effect of DNA Supercoiling on the Geometry of Holliday Junctions

Mutations of Bacteriophage T4 59 Helicase Loader Defective in Binding Fork DNA and in Interactions with T4 32 Single-stranded DNA-binding Protein

Contact Info

Product

Resources

About