High mobility group proteins in ram spermatids

The four major high mobility group proteins HMG 1, 2, 14 and 17, HMG 19B and histone H1(0) were identified in the ram testis by their extraction and solubility characteristics and by their electrophoretic mobilities. HMG 14 and 17 were isolated by chromatography and amino acid analysis revealed that they were similar to their calf thymus analogues. A protein, named 2R and co-extracted with HMG 14, was also purified and analysed. Electrophoretic analyses of the proteins extracted by 0.75 M perchloric acid (PCA) or by 0.35 M NaCl from round and non-round spermatids, separated by centrifugal elutriation, showed that the four major HMG proteins disappear from nuclei in the oldest round spermatids, at the time the nuclear content of protein 2R and histone H1(0) increases in spermatids. Ubiquitin and HMG 19B were present in the round and elongating spermatids, but not in elongated spermatids which contained only protamine. The relation was considered between several protein changes and genetic inactivation and structural reorganization of the spermatid chromatin.     

High mobility group proteins and their post-translational modifications

The high mobility group (HMG) proteins, including HMGA, HMGB and HMGN, are abundant and ubiquitous nuclear proteins that bind to DNA, nucleosome and other multi-protein complexes in a dynamic and reversible fashion to regulate DNA processing in the context of chromatin. All HMG proteins, like histone proteins, are subjected to extensive post-translational modifications (PTMs), such as lysine acetylation, arginine/lysine methylation and serine/threonine phosphorylation, to modulate their interactions with DNA and other proteins. There is a growing appreciation for the complex relationship between the PTMs of HMG proteins and their diverse biological activities. Here, we reviewed the identified covalent modifications of HMG proteins, and highlighted how these PTMs affect the functions of HMG proteins in a variety of cellular processes.     

Antigenic determinants of high mobility group chromosomal proteins 1 and 2

The antigenic determinants of nonhistone high mobility group chromosomal proteins 1 (HMG-1) and 2 (HMG-2) were studied with rabbit antisera elicited against HMG-1 and against HMG-2 and monoclonal antibodies elicited by HMG-1. The monoclonal antibodies did not distinguish between the two proteins, suggesting that they have specificity toward a shared determinant. Whereas anti-HMG-1 did not, anti-HMG-2 did distinguish between the proteins, suggesting that the anti-HMG-2 serum contains antibodies against peptides which differ between the proteins. Peptides were generated from HMG-1 and HMG-2 by controlled digestion with trypsin and pepsin. Analysis of the digests by ELISA and by sodium dodecyl sulfate electrophoresis followed by diazobenzyloxymethyl transfer, antibody binding and autoradiography revealed that most of the antibodies are against sequential determinants some of which are smaller than 3000 in molecular weight.     

High mobility group proteins HMG1 and HMG2 do not decrease the melting temperature of DNA

High mobility group proteins 1 and 2 isolated in non-denaturing conditions cannot decrease the temperature of denaturation of DNA. When they are isolated or treated with tricloroacetic acid a hyperchromic peak below the melting temperature of free DNA appears in agreement with previous data ( Javaherian et al. (1979) Nucl . Acids Res. 6, 3569-3580). We show that this is due to light scattering of aggregated protein at submelting temperatures and not to melting of DNA.     

Post-synthetic modifications of nuclear proteins. High mobility group proteins are methylated.

Calf thymus high mobility group proteins 1 and 2 (HMG-1 and HMG-2) were purified to homogeneity from a 0.35 M NaCl extract of chromatin by selective precipitation with trichloroacetic acid followed by ion exchange chromatography on CM-Sephadex. Amino acid analysis of an acid hydrolyzate of HMG-1 and HMG-2 proteins revealed the presence of a ninhydrin-positive peak identified as N^G,N^G-dimethylarginine. Radioactive tracer experiments confirmed the presence of methylated arginine in HMG proteins.     

Immunochemical studies of high mobility group non-histone chromatin proteins HMG 1 and HMG 2

Sera were raised to non-histone chromatin proteins HMG 1 and HMG 2. Immunoperoxidase staining localised these proteins on chromosomes during mitosis and indicated a cell cycle-related variation in these proteins during interphase. Some species differences in HMG 1 and HMG 2 were also observed.     

High mobility group box transcription factor 1 (HBP1) from Lampetra japonica affects cell cycle regulation

High mobility group (HMG) box‐containing protein 1 (HBP1) is a member of the HMG family of chromosomal proteins. Previous studies have shown that human HBP1 exhibits tumor‐suppressor activity. Here, we identified a homologue of HBP1, L‐hbp1, in Lampetra japonica. The L‐hbp1 gene shared high sequence similarity with its homologues in jawed vertebrates, as shown by bioinformatics analyses. L‐hbp1 contains a 1,584‐bp open reading frame that encodes 527 amino acids. A pAdenox‐L‐HBP1 plasmid was constructed and transfected successfully in Raji cells, as revealed by real‐time PCR. The overexpression of L‐HBP1 reduced cell growth rates, inhibited G1 phase progression, decreased cyclin D1 and c‐Myc protein expression, and increased p53 protein expression. Western blot and immunohistochemical assays showed that L‐HBP1 was primarily distributed in the heart, kidney, gill and liver of lamprey. Cell cycle analysis revealed that decreased L‐HBP1 expression in HBP1 morpholino oligonucleotide‐transfected lamprey cells resulted in a decreased fraction of cells in the G1 phase and corresponding increases in the S and G2/M phases. Additionally, treatment of lamprey cardiac cells with pharmacological inhibitors of p38 MAP kinase released the cells from G1 arrest. Together, these results indicated that HBP1 expression in lamprey was correlated with the onset of mitotic arrest in these cells, which have implications for cell cycle regulation.     

High mobility group proteins 1 and 2 are present in simian virus 40 provirions, but not in virions.

Viral particles at the late stages of SV40 morphogenesis were examined for the presence of HMG proteins 1 and 2, by an immunochemical method involving the transfer of proteins from polyacrylamide gels to nitrocellulose membranes. It was found that these proteins are present in SV40 provirions, in which histone H1 is still associated with viral chromatin, but absent in mature SV40 virions.     

High mobility group nonhistone chromosomal proteins also exist in Tetrahymena.

High mobility group (HMG) nonhistone chromosomal proteins have been shown to exist also in the ciliated protozoan Tetrahymena pyriformis. One or two histone-like components were extracted with 0.25 M HCl from the chromatin, in addition to five histone species. These proteins were also extracted selectively with 0.5 M HClO4, 0.35 M NaCl, or 4 mM spermidine, together with H1 histone, and were characterized as HMG proteins on the basis of the following criteria: high mobilities on polyacrylamide gel electrophoresis, relatively low molecular weights, amino acid compositions rich in lysine and glutamic acid, and relative contents in chromatin. This extends the distribution of the HMG proteins to all four eukaryotic kingdoms, and suggests the possibility that they have some universal role in chromatin structure and function.