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High Mobility Group A (HMGA) is a family of architectural nuclear factors which play an important role in neoplastic transformation. HMGA proteins are multifunctional factors that associate both with DNA and nuclear proteins that have been involved in several nuclear processes including transcription. HMGA localization is exclusively nuclear but, to date, the mechanism of nuclear import for these proteins remains unknown. Here, we report the identification and characterization of a nuclear localization signal (NLS) for HMGA2, a member of the HMGA family. The NLS overlaps with the second of the three AT-hooks, the DNA-binding domains characteristic for this group of proteins. The functionality of this NLS was demonstrated by its ability to target a heterologous beta-galactosidase/green fluorescent protein fusion protein to the nucleus. Mutations to alanine of basic residues within the second AT-hook resulted in inhibition of HMGA2 nuclear localization and impairment of its function in activating the cyclin A promoter. In addition, HMGA2 was shown to directly interact with the nuclear import receptor importin-alpha2 via the second AT-hook. HMGA proteins are overexpressed and rearranged in a variety of tumors; our findings can thus help elucidating their role in neoplastic transformation.

The HMGI-C protein is a nuclear phosphoprotein expressed at high levels in transformed cells. The cDNA encoding the mouse protein has been isolated and the sequence of the encoded protein shows that it is related to the HMGY and I proteins, proteins which bind in the minor groove of DNA containing stretches of A and T. The HMGI-C protein has three short highly basic domains, an acidic C-terminal domain, and potential CDC2/p34 and casein kinase II phosphorylation sites. Analysis of mRNA levels demonstrate that the HMGI-C gene is not expressed in a variety of mouse tissues but is expressed in Lewis lung carcinoma cells.

High mobility group I proteins (HMGI, HMGY and HMGI-C) are a family of low molecular mass non-histone nuclear proteins which constitute an important component of the active chromatin structure. Two members of this family, HMGI and HMGY, have been demonstrated to contribute to the transcriptional regulation of several promoters by interacting with the DNA and with different transcription factors. On the contrary, very little is known about the third member, HMGI-C, which plays an important role during embryonic growth and in the process of cell transformation, its gene being rearranged in a large number of mesenchimal tumors. In this paper we show for the first time that HMGI-C is also able to function as architectural factor, enhancing the activity of a transcription factor, NF-kappaB, through the PRDII element of the beta-interferon enhancer. Moreover we show that this enhancement is absolutely dependent on the binding of HMGI-C to its target sequence. The demonstration that HMGI-C is able to modulate transcription is thus an important initial step in the identification of genes regulated by this factor.

This paper describes the characterisation of a novel chicken homeobox gene, Prh, whose encoded homeodomain sequence differs significantly from those of other factors which have been described. As expected, a portion of the encoded protein, containing the homeodomain, is capable of sequence-specific DNA-binding. Outside the homeodomain, Prh, possesses an N-terminal region extremely rich in proline residues and a C-terminal acidic portion, either of which may function as transcription regulatory domains. Since, among the chicken tissues tested, its transcription is restricted to haematopoietic cells, lung and liver, it may function in tissue-specific patterns of gene regulation. Human and murine Prh homologues have also been identified; so it is likely that such genes are a general feature of vertebrate genomes.

A method is described for the rapid purification of high quality lambda DNA. The method can be used from either liquid or plate lysates and on a small scale or a large scale. It relies on the preadsobtion of all polyanions present in the lysate to an "insoluble" anion-exchange matrix (DEAE or TEAE). Phage particles are then disrupted by combined treatment with EDTA/proteinase K and the resulting DNA is precipitated by the addition of the cationic detergent cetyl (or hexadecyl)-trimethyl ammonium bromide-CTAB ("soluble" anion-exchange matrix). The precipitated CTAB-DNA complex is then exchanged to Na-DNA and ethanol precipitated. The resultant purified DNA is suitable for enzymatic reactions and provides a high quality template for dideoxy-sequence analysis.