The Human Cruciform-binding Protein, CBP, Is Involved in DNA Replication and Associates in Vivo with Mammalian Replication Origins

Novac, Olivia; Álvarez, David; Pearson, Christopher E.; Price, Gerald B.; Zannis‐Hadjopoulos, Maria

doi:10.1074/jbc.m107902200

Cited by 32 publications

(68 citation statements)

References 75 publications

(57 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This suggests that palindromic DNA may adopt a cruciform conformation within the chromatin scaffold. The conformational status of genomic DNA affects many cellular functions through the regulation of DNA replication or transcription (23,32). Thus, the elucidation of an in vivo cruciform structure may lead to a further understanding of the regulation of such cellular functions.…”

Section: Discussionmentioning

confidence: 99%

Cruciform DNA Structure Underlies the Etiology for Palindrome-mediated Human Chromosomal Translocations

Kurahashi

Inagaki

Yamada

et al. 2004

Journal of Biological Chemistry

106

View full text Add to dashboard Cite

There is accumulating evidence to suggest that palindromic AT-rich repeats (PATRRs) represent hot spots of double-strand breakage that lead to recurrent chromosomal translocations in humans. As a mechanism for such rearrangements, we proposed that the PATRR forms a cruciform structure that is the source of genomic instability. To test this hypothesis, we have investigated the tertiary structure of a cloned PATRR. We have observed that a plasmid containing this PATRR undergoesaconformationalchange,causingtemperaturedependent mobility changes upon agarose gel electrophoresis. The mobility shift is observed in physiologic salt concentrations and is most prominent when the plasmid DNA is incubated at room temperature prior to electrophoresis. Analysis using two-dimensional gel electrophoresis indicates that the mobility shift results from the formation of a cruciform structure. S1 nuclease and T7 endonuclease both cut the plasmid into a linear form, also suggesting cruciform formation. Furthermore, anti-cruciform DNA antibody reduces the electrophoretic mobility of the PATRR-containing fragment. Finally, we have directly visualized cruciform extrusions from the plasmid DNA with the size expected of hairpin arms using atomic force microscopy. Our data imply that for human chromosomes, translocation susceptibility is mediated by PATRRs and likely results from their unstable conformation.The constitutional t(11;22)(q23;q11) is the only known recurrent non-Robertsonian translocation in humans. Its recurrent nature implicates a specific genomic structure at the t(11;22) breakpoints. Analyses of numerous independent t(11;22) cases have localized the breakpoints within palindromic AT-rich repeats (PATRRs) 1 on 11q23 and 22q11 (1-4). Most 11;22 translocations show breakpoints at the center of the PATRRs, suggesting that the center of the palindrome is susceptible to double-strand breaks, leading to the translocation (5, 6). Indeed, translocation-specific PCR detects a high frequency of de novo t(11;22)s in normal sperm samples (7).The breakpoints on 22q11 are located within one of the unclonable gaps in the human genome (8, 9). Extensive screening of YAC/BAC/PAC libraries has not been successful in cloning this breakpoint region. However, experimentally derived sequences from numerous t(11;22) junction fragments demonstrate that the 22q11 breakpoints reside within a larger PATRR. The breakpoints of a variety of translocations involving 22q11 cluster within this region, suggesting that the 22q11 PATRR is highly unstable (10 -13). More recently, molecular cloning of translocation breakpoints has demonstrated similar palindromic sequences on partner chromosomes, such as 17q11, 4q35

show abstract

Section: Discussionmentioning

confidence: 99%

Cruciform DNA Structure Underlies the Etiology for Palindrome-mediated Human Chromosomal Translocations

Kurahashi

Inagaki

Yamada

et al. 2004

Journal of Biological Chemistry

106

View full text Add to dashboard Cite

show abstract

“…In vivo crosslinking and chromatin fragmentation Ten plates (15 cm) of cells at 60-70% confluency were washed with pre-warmed PBS and treated with 1% formaldehyde for 10 minutes to crosslink proteins and DNA in vivo (Ritzi et al, 1998;Novac et al, 2001;Novac et al, 2002); they were then washed and scraped into ice-cold PBS, and resuspended in lysis buffer (50 mM HEPES-KOH pH 7.5, 140 mM NaCl, 1% Triton X-100, 2 mM EDTA, Complete protease inhibitors tablet) (Roche Molecular Biochemicals). Following passage through a 21G needle three times, the nuclei were harvested by centrifugation at 2500 g for 5 minutes at 4°C.…”

Section: Western Blot Analysis and Quantificationmentioning

confidence: 99%

Decreased origin usage and initiation of DNA replication in haploinsufficient HCT116 Ku80+/- cells

Sibani

Price

Zannis‐Hadjopoulos

2005

Journal of Cell Science

Self Cite

View full text Add to dashboard Cite

One of the functions of the abundant heterodimeric nuclear protein, Ku (Ku70/Ku80), is its involvement in the initiation of DNA replication through its ability to bind to chromosomal replication origins in a sequence-specific and cell cycle dependent manner. Here, using HCT116 Ku80+/- cells, the effect of Ku80 deficiency on cell cycle progression and origin activation was examined. Western blot analyses revealed a 75% and 36% decrease in the nuclear expression of Ku80 and Ku70, respectively. This was concomitant with a 33% and 40% decrease in chromatin binding of both proteins, respectively. Cell cycle analysis of asynchronous and late G1 synchronized Ku80+/- cells revealed a prolonged G1 phase. Furthermore, these Ku-deficient cells had a 4.5-, 3.4- and 4.3-fold decrease in nascent strand DNA abundance at the lamin B2, β-globin and c-myc replication origins, respectively. Chromatin immunoprecipitation (ChIP) assays showed that the association of Ku80 with the lamin B2, β-globin and c-myc origins was decreased by 1.5-, 2.3- and 2.5-fold, respectively, whereas that of Ku70 was similarly decreased (by 2.1-, 1.5- and 1.7-fold, respectively) in Ku80+/- cells. The results indicate that a deficiency of Ku80 resulted in a prolonged G1 phase, as well as decreased Ku binding to and activation of origins of DNA replication.

show abstract

“…In humans, for example, a cruciform-binding protein from the 14-3-3 family has been found (14,15) as have its yeast counterparts Bmh1p and Bmh2p (16). These proteins have been implicated in initiation of replication (17). RAD51B, another human protein, binds 4WJDNA and has a function in homologous recombinational repair (18).…”

mentioning

confidence: 99%

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

The Human Cruciform-binding Protein, CBP, Is Involved in DNA Replication and Associates in Vivo with Mammalian Replication Origins

Cited by 32 publications

References 75 publications

Cruciform DNA Structure Underlies the Etiology for Palindrome-mediated Human Chromosomal Translocations

Cruciform DNA Structure Underlies the Etiology for Palindrome-mediated Human Chromosomal Translocations

Decreased origin usage and initiation of DNA replication in haploinsufficient HCT116 Ku80+/- cells

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

Contact Info

Product

Resources

About