Thermosensitive vector derived from RP1 plasmid for detection of transposable elements

The involvement of the transposable DNA element of E. coli K12 chromosome in integrative recombination of RP1 plasmid was studied. Using temperature sensitive for replication plasmid RP1ts12--the derivative of RP1 which contains mutated transposon Tnl, it was shown that integration of RP1 into host chromosome and Hfr formation may occur according to a mechanism mediated by chromosome IS-elements. Plasmids that are desintegrated from the chromosome of these Hfrs contain discrete DNA segments (IS-elements) and possess elevated frequency of integration into chromosome of rec+ cells. The latter was used for selection of RP1ts12 recombinants carrying chromosome IS. For identification of IS involved in RP1 integration the number of independent RP1ts 12 recombinants was subjected to restriction and heteroduplex analysis. By analysing recombinants integrated into bacterial chromosome with frequency 5 X 10(-3), a new IS-element of E. coli K12 designated IS111 was discovered. IS111-element is about 1500bp of length, contains Smal, Pst1 and BamH1 restriction endonuclease sites and was found in the same position on the plasmid RP1 in two different orientations. IS-elements that have been revealed in a number of other RP1ts12 recombinants were preliminary identified as IS1-like elements. One recombinants plasmid was found to have an IS5-like elements. The activity of IS-elements inserted into RP1ts12 in recA-dependent integrative recombination was estimated. From the data of absolute and relative RP1ts12 integration frequencies mediated by IS111, IS1- and IS5-like elements a conclusion was made about the absence of E. coli K12 chromosome IS-elements in RP1 plasmid. The Hfr-formation and chromosomal gene transfer by recombinant plasmids RP1ts12: IS111 were studied. The possibility to use insertion RP1ts12 derivatives for the estimation of copies number, mapping and definition of orientation of IS-elements in bacterial chromosome and the possibilities for detection of transposable DNA elements using RP1ts12 in a wide range of gram-negative bacteria are discussed.     

Polymorphism of the angiotensin I-converting enzyme gene in diabetic retinopathy patients and type 2 diabetes mellitus patients with myocardial infarction

The insertion/deletion polymorphism (I/D) of the angiotensin I-converting enzyme gene (ACE) was examined in type I diabetes mellitus patients (DM) with (n = 31), or without (n = 33) retinopathy, and in type 2 DM patients with either myocardial infarction (MI; n = 75), or with (n = 37), or without (n = 178) retinopathy. No association between the ACE gene and retinopathy in type 1 and type 2 DM patients was revealed. In the type 2 DM patients with MI, a statistically significant (P < 0.05) elevation of the D allele frequency (64%) compared to that without MI (55.3%), together with statistically nonsignificant prevalence of the DD homozygotes (41% versus 30.6%) was observed. Our data indicate that the D allele (RR 1.43) and DD genotype (RR 1.75) are risk factors for myocardial infarction in the type 2 DM patients.     

A Search for Association between the Polymorphic Markers of <Emphasis Type="Italic">PON1</Emphasis> and <Emphasis Type="Italic">PON2</Emphasis> Genes and Diabetic Nephropathy in Patients with Type I Diabetes Mellitus

Alleles and genotypes of polymorphic markers of paraoxonase 1 and paraoxonase 2 genes (PON1 and PON2) encoding enzymes of the organism antioxidant defense were compared in type 1 diabetes mellitus patients with or without diabetic nephropathy. The patients with nonoverlapping (“polar”) phenotypes constituted different groups. The first group contained patients with diabetic nephropathy (DN+, n = 62), clinical proteinuria (albuminuria above 300 mg per day), and at least 15-year disease duration. In control group, the patients had no diabetic nephropathy (DN−, n = 68), their albuminuria was below 200 mg per day, and disease duration was at least 20 years. Comparative analysis with exact Fisher’s test revealed no significant differences in frequencies of alleles and genotypes of the PON1 gene polymorphic marker Gln192Arg and of PON2 gene polymorphic markers Ala148Gly and Cys311Ser. Our results suggest that the polymorphic markers studied are not associated with diabetic nephropathy among Russian patients in Moscow.__________Translated from Genetika, Vol. 41, No. 6, 2005, pp. 844–849.Original Russian Text Copyright © 2005 by Voron’ko, Yakunina, Shestakova, Zotova, Chugunova, Shamkhalova, Vikulova, Debabov, Dedov, Nosikov.     

Allele polymorphism of the dopamine transporter gene in patients with endogenous psychoses: Association with pathological syndromes

A study was made of the association of the allele polymorphism of the 3′VNTR locus of the dopamine transporter (DAT) gene with schizophrenia, schizo-affective psychosis, and affective disorders. Three alleles (440, 480, and 520 nt) were found and the allele and genotype frequencies estimated in all groups. The allele and genotype frequencies in patients with depression significantly differed from those in controls and in patients with bipolar affective psychosis and schizophrenia. The results were correlated with the averaged MMPI profiles of controls and affective patients. In the latter group, 480/480 homozygotes significantly differed from patients with the other genotypes in the mean score on Hypochondria and Hysteria scales. The possible association of the DAT-3′VNTR polymorphism and individual syndromes, which are related to different mechanisms of psychological defense, is discussed.     

Association of the polymorphisms of the <Emphasis Type="Italic">ERBB3</Emphasis> and <Emphasis Type="Italic">SH2B3</Emphasis> genes with type 1 diabetes

To study the association with diabetes mellitus type 1, we analyzed the distribution of allele and genotype frequencies of polymorphic marker rs2292239 of ERBB3 gene, encoding epidermal growth factor receptor type 3 and polymorphic marker rs3184504 of SH2B3 gene, encoding adaptor protein LNK. The study included groups of T1DM patients and unrelated controls of Russian origin. Genotyping was performed using RFLP and real-time amplification methods. No statistically significant association with type 1 diabetes was found for the polymorphic marker rs2292239 of ERBB3, while the analysis of the distribution of allele and genotype frequencies of the polymorphic marker rs3184504 of SH2B3 gene revealed the association with T1DM in the Russian population.     

Polymorphic markers of the NO synthase genes and genetic predisposition to diabetic polyneuropathy in patients with type 1 diabetes mellitus

The allele and genotype frequencies of polymorphic markers of NOS1, NOS2 and NOS3 genes, encoding three types of NO synthases, were compared in type 1 diabetes patients with and without diabetic polyneuropathty. 180 type 1 diabetes patients (T1DM) of Russian or Eastern Slavonic origin, living in Moscow city, were divided into two groups using non-overlapping (polar) phenotypes. 86 patients had overt DPN and T1DM duration in this group was less than 5 years (DPN+ group) and 94 patients had no clinical DPN and T1DM duration was more than 10 years (DPN- group). We have not found the significant differences of allele and genotype frequencies of polymorphic markers (CA)n of NOS1 gene, (CCTTT)n of NOS2 gene, ecNOS4a/4b and Glu298Asp of NOS3 gene that indicates that all these markers are not associated with diabetic polyneuropathty. Only in the case of (CCTTT)n marker of NOS2 gene we have found a tendency for the association of 14 allele with DPN development. The carriers of this allele have the lower risk of DPN in T1DM.     

Identification of the locus associated with diabetic nephropathy in type 1 diabetes mellitus

Polymorphic tetranucleotide microsatellites D3S1512, D3S1744, D3S1550, and D3S232 were used to study the association of chromosome region 3q21-q25 neighboring the angiotensin II receptor type 1 gene (AT2R1) with diabetic nephropathy (DN) in diabetes mellitus type 1 (DM1). Allele and genotype frequencies were compared for DM1 patients with (N = 39) or without (N = 62) DN. Fisher's exact test with Bonferroni's correction revealed significant differences in frequencies of two D3S2326 alleles, one D3S1512 allele, and one allele and one genotype of D3S1550. No significant difference was observed with D3S1744. Thus, region 3q21-q25 proved tightly associated with DN in ethnic Russians with DM1 from Moscow.     

Properties of nitrate reductase from <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> 11dn1 fungi grown under anaerobic conditions

Production of nitrate reductase was studied in 15 species of microscopic fungi grown on a nitrate-containing medium. Experiments were performed with Fusarium oxysporum 11dn1, a fungus capable of producing nitrous oxide as the end product of denitrification. Moreover, a shift from aerobic to anaerobic conditions of growth was accompanied by a sharp increase in the activity of nitrate reductase. Studies of nitrate reductase from the mycelium of Fusarium oxysporum 11dn1, grown under aerobic and anaerobic conditions, showed that this enzyme belongs to molybdenum-containing nitrate reductases. The enzymes under study differed in the molecular weight, temperature optimum, and other properties. Nitrate reductase from the mycelium grown under aerobic conditions was shown to belong to the class of assimilatory enzymes. However, nitrate reductase from the mycelium grown anaerobically had a dissimilatory function. An increase in the activity of dissimilatory nitrate reductase, observed under anaerobic conditions, was associated with de novo synthesis of the enzyme.__________Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 3, 2005, pp. 292–297.Original Russian Text Copyright © 2005 by Morozkina, Kurakov, Nosikov, Sapova, Lvov.