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The HMGIY non-histone proteins play important roles as architectural transcription factors that regulate gene transcription in mammalian cells and also act as host-supplied cofactors necessary for retroviral integration. The genes coding for the HMGIY proteins are proto-oncogenes, and their aberrant or over-expression is correlated with both neoplastic transformation and metastatic progression in a wide variety of tumors. Here, we report the first complete sequence of the murine Hmgiy (a.k.a. Hmga1) gene and provide a detailed comparison of this with the sequence and organization of the human HMGIY gene, including an analysis of its promoter region with the previously unreported 5' upstream region of the human gene. These analyses reveal a remarkable degree of overall sequence conservation in both the protein coding and promoter regions of the murine and human genes, including conservation of the c-Myc binding site that has been demonstrated to regulate murine Hmgiy transcription (Wood et al., 2000. Mol. Cell. Biol. 20, 5490-5502). The promoters of both genes contain other conserved transcription factor binding sites that may also represent important cis-regulatory elements. Two exons present in the 5' untranslated region of the human gene, however, are missing from the murine gene, suggesting that these two closely related mammalian species regulate transcription of their Hmgiy genes in an individualistic manner.

Histone H1, HMG-1 and HMG-I(Y) are mammalian nuclear proteins possessing distinctive DNA-binding domain structures that share the common property of preferentially binding to four-way junction (4H) DNA, an in vitro mimic of the in vivo genetic recombination intermediate known as the Holliday junction. Nevertheless, these three proteins bind to 4H DNA in vitro with very different affinities and in a mutually exclusive manner. To investigate the molecular basis for these distinctive binding characteristics, we employed base pair resolution hydroxyl radical footprinting to determine the precise sites of nucleotide interactions of both HMG-1 and histone H1 on 4H DNA and compared these contacts with those previously described for HMG-I(Y) on the same substrate. Each of these proteins had a unique binding pattern on 4H DNA and yet shared certain common nucleotide contacts on the arms of the 4H DNA molecule near the branch point. Both the HMG-I(Y) and HMG-1 proteins made specific contacts across the 4H DNA branch point, as well as interacting at discrete sites on the arms, whereas the globular domain of histone H1 bound exclusively to the arms of the 4H DNA substrate without contacting nucleotides at the crossover region. Experiments employing the chemical cleavage reagent 1, 10-orthophenanthroline copper(II) attached to the C-terminal end of a site-specifically mutagenized HMG-I(Y) protein molecule demonstrated that this protein binds to 4H DNA in a distinctly polar, direction-specific manner. Together these results provide an attractive molecular explanation for the observed mutually exclusive 4H DNA-binding characteristics of these proteins and also allow for critical assessment of proposed models for their interaction with 4H DNA substrates. The results also have important implications concerning the possible in vivo roles of HMG-I(Y), histone H1 and HMG-1 in biological processes such as genetic recombination and retroviral integration.

Integration of mycobacteriophage L5 requires the mycobacterial integration host factor (mIHF) in vitro. mIHF is a 105-residue heat-stable polypeptide that is not obviously related to HU or any other small DNA-binding proteins. mIHF is most abundant just prior to entry into stationary phase and is essential for the viability of Mycobacterium smegmatis.

We evaluated the effect of 2 years' therapy with an angiotensin converting enzyme inhibitor (captopril) in 16 patients who had severe reflux nephropathy and microalbuminuria. During the period of therapy, microalbuminuria decreased, glomerular filtration rate measured by diethylenetriamine pentaacetate scan, serum creatinine, and blood pressure remained stable. We suggest the captopril was useful in reducing microalbuminuria and may have slowed the progression of renal damage in our patients.

Lysogenization of mycobacteriophage L5 involves integration of the phage genome into the Mycobacterium smegmatis chromosome. Integration occurs by a site-specific recombination event between a phage attachment site, attP, and a bacterial attachment site, attB, which is catalyzed by the phage-encoded integrase protein. DNase I footprinting reveals that L5 integrase binds to two types of sites within attP which span an unexpectedly large region of 413 bp: seven arm-type sites (P1 to P7) each of which correspond to a consensus sequence 5'-TGCaaCtcYy, and core-type sites at the points of strand exchange. Mutational analyses indicate that not all of the arm-type sites are required for integration, and that the P3 site and the rightmost pair of sites (P6 and P7) are dispensable for integration. We show that a 252 bp segment of attP DNA is sufficient for efficient integrative recombination and that int can be provided in trans for simple and efficient transformation of the mycobacteria.

Bacterial integration host factors (IHFs) play central roles in the cellular processes of recombination, DNA replication, transcription, and bacterial pathogenesis. We describe here a novel mycobacterial IHF (mIHF) of Mycobacterium smegmatis and Mycobacterium tuberculosis that stimulates integration of mycobacteriophage L5. mIHF is the product of a single gene and is unrelated at the sequence level to other integration host factors. By itself, mIHF does not bind preferentially to attP DNA, although it significantly alters the pattern of integrase (Int) binding, promoting the formation of specific integrase-mIHF-attP intasome complexes.

The aim of this study was to investigate the relationship between plasma insulin levels, peripheral insulin sensitivity and androgen secretion in ten patients with polycystic ovary syndrome and in six obese women as compared with six normal-weight control subjects. During a euglycemic-hyper-insulinemic clamp no significant change of testosterone, androstenedione or dehydroepiandrosterone sulfate plasma levels was observed in the two groups of patients or in the control subjects; insulin sensitivity was clearly reduced and was similar in polycystic ovary patients and in obese women, in spite of the different plasma androgen levels. A long-term (5 months) androgen suppression with the gonadotropin-releasing hormone agonist leuprolide was not able to improve significantly the insulin sensitivity. These results demonstrate that the short-term hyperinsulinemia achieved with the clamp technique does not affect androgen secretion and that insulin resistance, measured with the same technique, is not influenced by long-term suppression of plasma androgen levels in polycystic ovary syndrome.