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GANT 58 Indications that HMGA proteins are involved
Indications that HMGA proteins are involved in regulating the structure and function of large chromatin domains come from the early work of Saitoh and Laemmli [143]. Employing immunolocalization techniques, these workers demonstrated that HMGA1 proteins are co-localized with both histone H1 and the enzyme topoisomerase II at sites called scaffold attachment regions (i.e., SARs), A/T-rich DNA sequences that constitute the structural backbone of metaphase chromosomes. SARs are thought to be cis-acting DNA elements located at the GANT 58 of large loops of gene-containing DNA and have been postulated to be involved in the dynamic regulation of both the chromatin structure and transcriptional activity the DNA loop or “domain”. Support for the idea that HMGA proteins participate in regulating the activity of such “domains” comes from experiments with in vitro transcription systems developed to search for proteins capable of antagonizing histone H1-mediated repression of transcription from SAR containing plasmid DNA templates [178]. Employing such systems, HMGA1 was identified as the nuclear protein that was able to out-compete histone H1 for binding to SAR elements and, as a consequence, induce gene transcription from previously repressed plasmid templates. Based on these and other data, a model has been proposed in which H1 and HMGA1 proteins compete for binding to SAR elements located at the base of large chromatin domains with the winner of this competition determining both the condensation state of the domain and the transcriptional capacity of the genes it contains [178]. It was further suggested that such domain regulation was likely to transpire during embryonic development and other situations where large-scale alterations in chromatin structure and gene expression occur. Several lines of evidence support the HMGA1:H1competition model for determining the structure and activity of large chromatin domains. These include the fact that HMGA1 proteins are greatly enriched in, and histone H1-depleted from, the transcriptionally active subfraction of chromatin isolated from cells [178]. Additional support for the model comes from in vitro experiments demonstrating that artificial multi-AT-hook (MATH) proteins (synthetic mimics of HMGA proteins) that bind tightly to SAR sequences are potent inhibitors of chromosome condensation when added to mitotic extracts of Xenopus oocytes [154]. Apoptosis is another biological process in which HMGA proteins influence global chromatin structure. Apoptosis, or programmed cell death, is characterized by chronological alterations of mitochondrial and plasma membrane function followed by marked changes in nuclear morphology, chromatin condensation, DNA fragmentation and other dramatic changes in cellular phenotype [16,105]. But to understand the role HMGA proteins play in apoptosis, it is first necessary to consider the complex roles these proteins are thought to play in development and in cancer. Intracellular concentrations of HMGA proteins are maximal during embryogenesis but drop to very low, often nearly undetectable, levels in normal differentiated somatic cells [27,179]. Most “immortalized” pre-cancerous cells and nearly all overtly cancerous cells, on the other hand, constitutively overexpress HMGA proteins with increasing concentrations being correlated with tumor progression, increasing metastatic potential and poor patient prognosis [58,131]. Overexpression of HMGA proteins in normal cells is toxic and induces apoptosis. Fedele et al. [50] have demonstrated, for example, that forced overexpression of HMGA1b in normal rat thyroid epithelial cells does not transform cells but rather induces apoptosis as a result of deregulation of S phase DNA synthesis, delaying entry of cells into the G2/M phase of the cell cycle and activation of the caspase-3 death pathway. The molecular mechanisms by which overexpression deregulates cell cycle progression are undoubtedly complex but include HMGA mediated aberrant induction of transcription of genes such as cyclin A [156] and AP-1 [162].