The high mobility group box: the ultimate utility player of a cell

Abstract

High mobility group (HMG) box proteins are abundant and ubiquitous DNA binding proteins with a remarkable array of functions throughout the cell. The structure of the HMG box DNA binding domain and general mechanisms of DNA binding and bending have been known for more than a decade. However, new mechanisms that regulate HMG box protein intracellular translocation, and by which HMG box proteins recognize DNA with and without sequence specificity, have only recently been uncovered. This review focuses primarily on the Sry-like HMG box family, HMGB1, and mitochondrial transcription factor A. For these proteins, structural and biochemical studies have shown that HMG box protein modularity, interactions with other DNA binding proteins and cellular receptors, and post-translational modifications are key regulators of their diverse functions.

Figure 1. Alignments of single- and dual-domain high mobility group (HMG)-box proteins

(a) Sequence alignment of the HMG domains of selected single HMG box proteins: structure specific recognition protein 1 (SSRP1), sex-determining region Y (SRY) [80], and the SRY-like box (Sox) family of HMGB proteins [81]. Primary site (1°, red font) intercalating residues and secondary site (‘2°’, green font) intercalating/hydrogen bonding residues are indicated. Conserved residues are shown in blue font. Green cylinders below the sequence alignment illustrate the regions of the proteins that form the alpha helices of the HMG box. (b) Sequence alignment of the HMG domains of selected dual HMG box proteins including mitochondrial transcription factor A (TFAM); the Saccharomyces cerevisiae homologue of TFAM, ABF2 (*numbering here begins after the 42 or 35 amino acid residue mitochondrial localization sequence for TFAM or ABF2, respectively); the Drosophila melanogaster dorsal repressor DSP1, and HMGB1-4. Primary and secondary intercalating residues and conserved residues are indicated as in (a). The cysteine residues that have been shown to form an intramolecular disulfide bond in HMGB1 are shown in magenta. The green unbroken cylinders below the sequence alignment indicate the amino acids that form the helical structures comprising the HMG boxes of TFAM, when bound to DNA. The broken green cylinder represents the linker region of TFAM that forms an alpha helix upon binding to DNA.

Figure 2. Structures of high mobility group (HMG)-box-DNA complexes

(a) Non-sequence-specific HMGD bound to an unmodified DNA decamer (PDB ID 1QRV) [33]. (b) Sequence-specific Sox4 bound to a 16 base pair DNA oligomer from the Lama 1 gene (PDB ID 3U2B) [41]. The structure is orientated as if it was superimposed onto the HMG box of HMGD in panel (a). (c) Dual HMG box Sex determining region Y-HMGB1 Box B chimera (Sry.B) bound to 16 base pair DNA oligomer containing the cognate Sry binding site (PDB ID 2GZK) [29]. SRY is in green and the HMGB1 box B is in pink. (d) Native dual HMG box mitochondrial transcription factor A (TFAM) bound to a 28 base pair segment of the human mitochondrial light strand promoter (PDB ID 3TMM) [31]. The numbering for TFAM is based on the residual protein after loss of the 42 amino acid residue long mitochondrial localization sequence, indicated by a *. The structure is oriented as if box A was superimposed onto the SRY HMG box from the SRY.B chimera in panel (c). The non-polar intercalating residues are shown in red, and polar residues at the 1° or 2° intercalation sites are shown in cyan.

Figure 3. Specificity of DNA recognition

(a) Sox17-DNA complex. The primary intercalating residue is shown in red, and the residue at the secondary intercalating site is shown in cyan. The bases ‘XY’, for which nucleotide interdependence has been observed, are shown in black, and the primary recognition site sequence is colored orange. (b) Two dimensional representation of the amino acids that contact the DNA in the crystal structures of Sox4 (PDB ID 3U2B) and Sox17 (PDB ID 3F27). The primary intercalating residue is Met 67 (red), and Sox17 amino acids that differ in DNA contacts from the Sox4 structure are indicated in green. Amino acid numbering is based on the Sox4 structure. Hydrogen bonds to the bases are indicated with blue lines and van der Waal’s contacts are shown as broken beige lines. The figure was generated using the program NUCPLOT [82]. (c) Sox2 (green) and the POU domains of Oct1 [51] (PDB ID 1GTO). The protein-protein interaction interface is circled in red. The DNA is colored as in (a).

Figure 4. High mobility group (HMG) boxes in nucleosome remodeling

The diagram summarizes and combines findings from S. cerevisiae and other eukaryotes. HMGB proteins in metazoans (yellow) and NHP6A in S. cerevisiae (red), in their individual cellular contexts can bind to nucleosomal DNA and assist ATP-dependent chromatin remodelers (grey) in loosening DNA and providing access to other DNA dependent machineries. Related SWI/SNF (in S. cerevisiae) and BAF (in metazoans) complexes harbor an HMG box containing subunit BAF57 (orange) that promotes nucleosome remodeling, in some cases requiring binding of calcium calmodulin (CaC in black). The histone chaperone FACT has three subunits in S. cerevisiae (Spt16, Pob3, NHP6A), of which NHP6A promotes the association of the FACT complex with nucleosomes to facilitate remodeling.

Figure 5. High mobility group box 1 (HMGB1) interactions in the cell

The diagram summarizes and combines findings from studies of post-translational modifications and cysteine oxidation of HMGB1 that correlate with different functions and localization of HMGB1. The DNA-bound nuclear form of HMGB1 (yellow) is generally reduced, but phosphorylation (green ‘P’), methylation (cyan ‘Me’) and acetylation (blue ‘Ac’) can regulate DNA (in blue) binding activity. The cytoplasmic and extracellular forms of HMGB1 are generally more highly acetylated and oxidized than the nuclear form. Cytoplasmic HMGB1 can compete with Bcl-2 (grey) for binding to Beclin1 (blue), which promotes autophagy. Release of HMGB1 to the extracellular space, where it can act on receptors of the same as well as neighboring cells, results in different responses, depending on the receptor; receptor for advanced glyclation end products (RAGE, purple) promotes autophagy, whereas toll-like receptor 4 (TLR4, green) promotes cytokine release, and oxidized forms of HMBG1 can promote apoptosis. Reduced Cys 22, Cys 44 and Cys 106 are indicated by a black ‘C’. The oxidized Cys 22 and Cys 44 form a disulfide bond indicated by red ‘C-C’ and the oxidized C 106 sulfinic acid is represented by a black ‘C’ that has a red star outline. The specific placement of the marks is not meant to indicate where they are located in HMGB1.