Preferential Binding of High Mobility Group 1 Protein to UV-damaged DNA

Abstract

Binding of chromosomal high mobility group 1 protein (HMG1) to UV-damaged DNA has been studied with oligonucleotides containing a single dipyrimidine site for formation of UV photolesions. Irradiation of an oligonucleotide with unique TT dinucleotide resulted in generation of cyclobutane pyrimidine dimer with no evidence for induction of (6-4) photoproducts, whereas the analysis of irradiated TC-containing oligonucleotide detected (6-4) photoproducts but not cyclobutane pyrimidine dimers. Mobility shift assays have revealed that HMG1 protein binds preferentially to irradiated TT and TC oligonucleotides. Photoreversal of cyclobutane pyrimidine dimers with DNA photolyase and hydrolysis of the (6-4) photoproducts with hot alkali substantially reduced but did not eliminate binding of HMG1. The protein, therefore, appears to bind the two main types of UV damages in DNA, but some other photolesion(s) contributes to the preferential binding of HMG1 to irradiated DNA. By quantifying gel shift assays and considering the efficiencies of lesion formation, we determined dissociation constants of 1.2 ؎ 0.5 and 4.0 ؎ 1.5 M for irradiated TT and TC oligonucleotides, respectively, and 70 ؎ 20 M for the control non-irradiated probes. Tryptic removal of the acidic COOH-terminal domain of HMG1 significantly affected binding of the protein to both irradiated and intact oligonucleotides. The potential role of HMG1 in recognition of the UV lesions in DNA is discussed.High mobility group proteins 1 and 2 (HMG1 and -2) 1 are abundant chromosomal proteins found in a variety of eukaryotic species. Despite their early identification and biochemical characterization (1) and subsequent implication in a number of cellular events, their functions are still unknown (for reviews see Refs. 2 and 3). The interest in these proteins sharply increased in the early 1990s with the discovery of a new kind of eukaryotic protein domain involved in interactions with DNA (4), which displayed sequence similarity to two homologous repeats of an 80-amino acid sequence in HMG1. This motif called HMG box was found in a wealth of less abundant eukaryotic proteins, mainly general transcription factors and gene-specific transcriptional activators (reviewed in Refs. 3 and 5). A general characteristic of these proteins is their ability to bind bent DNA or to induce bending in DNA independent of sequence or the existence of prebent regions (6). Thus, HMG1 binds preferentially to distorted DNA sites such as synthetic cruciform DNA structures (7,8) and the lesions formed on DNA by the anticancer drug cisplatin (9, 10). The ability of HMG box-containing proteins to mediate bending in linear duplex DNA was demonstrated by several approaches (11-13). In addition, these proteins can constrain negative supercoils in plasmid DNA (14 -16) and can serve as "architectural" elements in the assembly of higher order nucleoprotein complexes (5,11,13,(17)(18)(19).The finding that HMG1 binds more tightly to cisplatin-damaged DNA than to unmodified B form DNA implies that the signal f...

High Mobility Group Box 1 Protein Is Methylated and Transported to Cytoplasm in Clear Cell Renal Cell Carcinoma

“…The general function of acidic tail remains elusive despite its negatively regulations on HMG boxes. It downregulates the binding and bending ability of HMG boxes for linear DNA and distorted DNA although the binding affinity to DNA minicircles is unaffected (Bianchi et al, 1989;Pil et al, 1992;Stros et al, 1994;Pasheva et al, 1998;Lee et al, 2000). Several investigations have shown that the acidic tail binds to HMG boxes in vitro (Ramstein et al, 1999;Jung et al, 2003).…”

Gadd45, a p53-Responsive Stress Protein, Modifies DNA Accessibility on Damaged Chromatin

“…4A to 6) suggests a possible recruitment and therefore concentration of Gadd45 at sites where the nucleosomes have been weakened by either histone posttranslational modifications or DNA damage. While binding of sequence specific transcription factors is generally inhibited on UV-damaged templates (66,73), it is apparently increased for proteins recognizing altered DNA structure (8,25,39,45,63). For example, the high-mobility-group protein HMG1 has recently been shown to bind preferentially to UV-damaged DNA (45).…”

“…While binding of sequence specific transcription factors is generally inhibited on UV-damaged templates (66,73), it is apparently increased for proteins recognizing altered DNA structure (8,25,39,45,63). For example, the high-mobility-group protein HMG1 has recently been shown to bind preferentially to UV-damaged DNA (45). This chromosomal protein has also been shown to bind preferentially to other distorted DNA structures such as synthetic cruciform (4) and lesions formed by the anticancer drug cisplatin (50).…”

“…More than 150 molecular markers, such as p53 status, have been determined in these lines (77). The cytotoxicity data were processed by using COMPARE (45) and programs in the DISCOV-ERY program set (77). The data were correlated to the capacity to induce GADD45 mRNA following exposure to ionizing radiation (2).…”

Overexpression of High-Mobility Group Box 2 Is Associated with Tumor Aggressiveness and Prognosis of Hepatocellular Carcinoma

“…HMGB1 overexpression has been reported in a variety of human cancers, including melanoma (9), pancreatic cancer, prostate cancer (10), colorectal cancer (11), and breast cancer (12). More importantly, HMGB proteins preferentially bind to cis-platinum(II)diaminedichloride (cisplatin)-modified DNA or to misincorporated nucleoside analogues and consequently inhibit nucleotide excision repair, which could be of great value in cancer treatment (13,14). Despite extensive characterization of the diverse roles of HMGB1 in cancer, much less is known of the signaling pathways of HMGB2, especially its relevance in carcinogenesis.…”