Founder vs. non-founder BRCA1/2 pathogenic alleles: the analysis of Belarusian breast and ovarian cancer patients and review of other studies on ethnically homogenous populations.

The spectrum of BRCA1/2 mutations demonstrates significant interethnic variations. We analyzed for the first time the entire BRCA1/2 coding region in 340 Belarusian cancer patients with clinical signs of BRCA1/2-related disease, including 168 women with bilateral and/or early-onset breast cancer (BC), 104 patients with ovarian cancer and 68 subjects with multiple primary malignancies involving BC and/or OC. BRCA1/2 pathogenic alleles were detected in 98 (29%) women, with 67 (68%) of these being represented by founder alleles. Systematic comparison with other relevant studies revealed that the founder effect observed in Belarus is among the highest estimates observed worldwide. These findings are surprising, given that the population of Belarus did not experience geographic or cultural isolation throughout history.

; THE ADVANCEMENTS IN TREATMENT OF HIV-INFECTED PATIENTS WITH HERPETIC INFECTION.

It was found on the base on the study of clinical and immunological parameters of 47 patients with HIV-associated herpes infections (recurrent labial herpes and/or aphthous stomatitis, genital herpes and shingles) that supplement of the basic therapy (valaciclovir 1.0 g daily intake orally 2 times a day for 7-10 days) 6 subcutaneous injections of the drug "Allokin-alpha" in dose 1 mg in a day enables significantly shorten the duration of clinical manifestations of herpes infections, reduce the frequency of relapses and also the duration of the first relapse after treatment. The immunoregulatory effect alokin-alpha used in the treatment of patients with HIV co-infection herpes is installed. Thus, the combination therapy provided a further reduce of CD4+-lymphocytes number at II clinical stage of HIV infection. The concentration of interleukin-8 (IL-8) decreased at I and II stages of immunodeficiency, that statistically weighty different from the values before treatment (P<0,05-0,02). Similarly, the level of IL-10 (P<0.05) decreased. It is important that the impact of treatment on immune parameters match the clinical effect.

Spectrum of APC and MUTYH germ-line mutations in Russian patients with colorectal malignancies.

Distribution of cancer-predisposing mutations demonstrates significant interethnic variations. This study aimed to evaluate patterns of APC and MUTYH germ-line mutations in Russian patients with colorectal malignancies. APC gene defects were identified in 26/38 (68%) subjects with colon polyposis; 8/26 (31%) APC mutations were associated with 2 known mutational hotspots (p.E1309Dfs*4 [n = 5] and p.Q1062fs* [n = 3]), while 6/26 (23%) mutations were novel (p.K73Nfs*6, p.S254Hfs*12, p.S1072Kfs*9, p.E1547Kfs*11, p.L1564X and p.C1263Wfs*22). Biallelic mutations in MUTYH gene were detected in 3/12 (25%) remaining subjects with polyposis and in 6/90 (6.7%) patients with colorectal cancer (CRC) carrying KRAS p.G12C substitution, but not in 231 early-onset CRC cases negative for KRAS p.G12C allele. In addition to known European founder alleles p.Y179C and p.G396D, this study revealed a recurrent character of MUTYH p.R245H germ-line mutation. Besides that, 3 novel pathogenic MUTYH alleles (p.L111P, p.R245S and p.Q293X) were found. Targeted next-generation sequencing of 7 APC/MUTYH mutation-negative DNA samples identified novel potentially pathogenic POLD1 variant (p.L460R) in 1 patient and known low-penetrant cancer-associated allele CHEK2 p.I157T in 3 patients. The analysis of 1120 healthy subjects revealed 15 heterozygous carriers of recurrent MUTYH mutations, thus the expected incidence of MUTYH-associated polyposis in Russia is likely to be 1:23 000.

[Whole exome sequencing in oncology].

Whole exome sequencing (WES) has become a leading tool for genetic analysis right after its invention. This approach permits the detection of mutations spread within coding regions of the entire genome. For cancer patients WES is particularly effective for the search of hereditary cancer mutations and identification of somatically mutated druggable genes. Use of WES already resulted in significant advances in understanding for molecular mechanisms of cancer.

[CHEK2-associated hereditary breast cancer].

CHEK2 is classified as a moderate-penetrance gene for hereditary breast cancer (BC). In Russia, CHEK2 mutations hold second position in the list of BC-predisposing gene defects after BRCAl, and include CHEK2 1100deIC, de15395, and IVS2+lG>A gene-inactivating alleles. CHEK2-driven breast carcinomas are generally characterized by poor prognosis and low sensitivity to the conventional therapeutic regimens. CHEK2 testing needs to be incorporated into routine clinical practice owing its overt clinical significance.

Hereditary breast-ovarian cancer syndrome in Russia.

Hereditary breast-ovarian cancer syndrome contributes to as much as 5-7% of breast cancer (BC) and 10-15% of ovarian cancer (OC) incidence. Mutations in the "canonical" genesBRCA1andBRCA2occur in 20-30% of affected pedigrees. In addition toBRCA1andBRCA2 mutations, germ-line lesions in theCHEK2,NBS1, andPALB2genes also contribute to familial BC clustering. The epidemiology of hereditary breast-ovarian cancer in Russia has some specific features. The impact of the "founder" effect is surprisingly remarkable: a single mutation,BRCA15382insC, accounts for the vast majority ofBRCA1defects across the country. In addition, there are two other recurrentBRCA1alleles:BRCA14153delA andBRCA1185delAG. BesidesBRCA1, in Russia breast cancer is often caused by germ-line alterations in theCHEK2andNBS1genes. In contrast toBRCA1andBRCA2, theCHEK2andNBS1heterozygosity does not significantly increase the OC risk. Several Russian breast cancer clinics recently started to investigate the efficacy of cisplatin in the therapy ofBRCA1-related cancers; initial results show a unique sensitivity ofBRCA1-associated tumours to this compound.

[The effects of Celebrex on mammary tumorigenesis and aging in HER2/neu transgenic mice].

It is well known that cyclooxygenase-2 (COX2) plays an important role in the development of many tumors including breast cancer. Our study was concerned with evaluating the effects of the selective COX2 inhibitor, celecoxib, on mammary tumorigenesis and aging in HER2/neu transgenic mice (24). Celecoxib (celebrex) 25 mg/kg was administered 5 times a week from the age of 2 months. Twenty-four intact females were in control. Monitoring kept track of tumor detection time, size, presence of lung metastases, food and water consumption, estral function, body weight and temperature. No significant differences between the two groups were reported as far as life-span, tumor growth rate, size and number of metastases to the lung is concerned. To sum up, celecoxib treatment failed to produce any significant effect on carcinogenesis in HER2/neu transgenic mice.

CHEK2 variants predispose to benign, borderline and low-grade invasive ovarian tumors.

OBJECTIVE: Three founder alleles of the CHEK2 gene have been associated with predisposition to a range of cancer types in Poland. Two founder alleles (1100delC and IVS2 + 1G >A) result in a truncated CHEK2 protein and the other is a missense substitution, leading to the replacement of a threonine with an isoleucine (I157T). METHODS: To establish if these variants play a role in the etiology of ovarian tumors, we genotyped 1108 Polish women with various types of ovarian tumors and 4000 controls for the three CHEK2 variants. We included 539 Polish women with benign ovarian cystadenomas, 122 women with borderline ovarian malignancies and 447 women with invasive ovarian cancer. RESULTS: Positive associations were seen with the CHEK2 I157T missense variant and ovarian cystadenomas (OR = 1.7; P = 0.005), with borderline ovarian cancers (OR = 2.6; P = 0.002) and with low-grade invasive cancers (OR = 2.1; P = 0.04). There was no association with ovarian cancer of high grade (OR = 1.0). The association between the I157T missense variant was then confirmed in a second sample of Russian patients with borderline ovarian cancers (OR = 2.7; P = 0.06). CONCLUSION: These data indicate that CHEK2 variants may predispose to a range of ovarian tumor types of low malignant potential, but not to aggressive cancers.

[Hydrogen peroxide as an inductor of uncultivable state of Vibrio cholerae eltor in an experiment].

The influence of hydrogen peroxide on the dynamics of transition into uncultivable state (UCS) and on the reversion of V. cholerae and their subcultures, resistant to hydrogen peroxide, was studied. The transition of the initial cultures in river and distilled water into UCS took place earlier than that in resistant to hydrogen peroxide variants. The capacity for reversion to hydrogen peroxide resistant subcultures preserved, on the average, 2 - 3 times longer. An increase in the level of hydrogen peroxide in uncultivable populations was found to be 2.7 - 4.4 times. Subcultures, resistant to hydrogen peroxide, in the vegetative form had lower characteristics of peroxide concentrations than in uncultivable form (UCF), but somewhat higher than in initial variants. In revertants the concentration of hydrogen peroxide was lower in UCF, but somewhat higher than in vegetative cultures. The dynamics of the formation of UCF by cholera vibrios, with different degree of stability to the action of hydrogen peroxide, the accumulation of hydrogen peroxide in uncultivable populations, the deceleration of transition into uncultivable forms, an accumulation of hydrogen peroxide and an increase in the time of the reversion of clones, resistant to hydrogen peroxide, made it possible to suggest that the accumulation of hydrogen peroxide was possible to make an essential contribution to the formation of UCF of cholera vibrios in an experiment.

TCP34, a nuclear-encoded response regulator-like TPR protein of higher plant chloroplasts.

We describe the identification of a novel chloroplast protein, designated TCP34 (tetratricopeptide-containing chloroplast protein of 34 kDa) due to the presence of three tandemly arranged tetratricopeptide repeat (TPR) arrays. The presence of the genes encoding this protein only in the genomes of higher plants but not in photosynthetic cyanobacterial prokaryotes suggests that TCP34 evolved after the separation of the higher plant lineage. The in vitro translated precursor could be imported into intact spinach chloroplasts and the processed products showed stable association with thylakoid membranes. Using a specific polyclonal antiserum raised against TCP34, three protein variants were detected. Two forms, T(1) and T(2), were associated with the thylakoid membranes and one, S(1), was found released in the stroma. TCP34 protein was not present in etioplasts and appeared only in developing chloroplasts. The ratio of membrane-bound and soluble forms was maximal at the onset of photosynthesis. The high molecular mass thylakoid TCP34 variant was found in association with a transcriptionally active protein/DNA complex (TAC) from chloroplasts and recombinant TCP34 showed specific binding to Spinacia oleracea chloroplast DNA. Two TCP34 forms, T(1) and S(1), were found to be phosphorylated. An as yet unidentified phosphorelay signal may modulate its capability for plastid DNA binding through the phosphorylation state of the putative response regulator-like domain. Based on the structural properties and biochemical analyses, we discuss the putative regulatory function of TCP34 in plastid gene expression.

[Dynamics of the reversible transition of Vibrio cholerae into the uncultivable state in the presence of organic and inorganic microcosmic components].

The dynamics of the transition of V. cholerae into the uncultivable state in distilled, river and tap water, containing organic and inorganic components added, was studied. As additives, potassium nitrate, potassium phosphate, magnesium sulfate, ammonium chloride, lysine, alpha-ketoglutarate, succinic acid, catalase were used. The study of the influence of biotic factors on transition into the uncultivable state was carried out in the presence of one-celled green algae Scenedesmus quadricauda or infusoria Paramecium caudatum. The linear dependence of speed of transition into the uncultivable form on the concentration of cells was noted. The composition of the microcosmic medium was also found to have some influence on the speed of transition into the uncultivable form and on the reversibility of this process. The presence of organic substances, such as peptone solution or destroyed cells of phyto- and zooplankton, in the microcosmic medium prolonged the time of transition into the uncultivable form and produced a positive effect on the capacity of the population to reversion. In respect of live biotic components, no such dependence was found. Inorganic additives prolonged the time of transition into the uncultivable state, but did not promote reversion.

[Interaction of proteins from general transcription complex RNA polymerase II with oligoribonucleotides].

We have analyzed an interaction of the general transcription complex RNA polymerase II proteins (RNA polymerase II, factors TBP, TFIIB, TFIIF, TFIIE and TFIIH) S. cerevisiae with the oligoribonucleotides. With the help of method EMSA was shown that labeled 32P labeled oligoribonucleotide 5'-ACUCUCUUCCGCAUCGC-3' (r-17) binds with the proteins and generates three species of the complexes with the three major shifts. All the three species of the complexes are RNA specific because a total RNA S. cerevisiae was a competitor for all three species but the TATA-containing oligodeoxyribonucleotide (500-fold molar excess) was not a competitor for its. Complexes 32P-r-17 with the proteins belonging to the middle shift are the sequence specific because unlabeled r-17 was a competitor for its binding (100-fold molar excess) but unlabeled UA-rich oligoribonucleotide (5'-AUAUUAUGUUCAAAA-3) was not a competitor for this shift (500-fold molar excess). Complexes belonging to the upper shift are RNA specific probably. We think 32P-r-17 interaction with the proteins belonging to the under shift is nonspecific corresponding to a sorbtion of 32P-r-17 on a protein. The data presented demonstrate that oligoribonucleotide and oligodeoxyribonucleotide don't compete for the binding sites on a basal transcription complex proteins.

[Hemolytic activity of Vibrio cholerae eltor and V. cholerae O139 toxigenic and nontoxigenic strains]. / Gemoliticheskaia aktivnost' toksikogennykh i netoksikogennykh shtammov Vibrio cholerae eltor i V.cholerae O139 serogruppy.

A total of 20 ctx- and 16 ctx+ V. cholerae eltor strains, 20 ctx- and 22 ctx+ V. cholerae O139 strains were under study. Hemolytic activity was tested in modified Greig test with sheep, guinea pig and rabbit red blood cells. The comparative study of the hemolytic properties of V. cholerae O1 and O139 under different conditions of cultivation demonstrated their capacity of lysing sheep red blood cells (SRBC) irrespective of the presence of toxigenic properties. A wider spectrum of lytic activity of ctx- strains in Greig test with respect to red blood cells of different animals and the capacity of lysing SRBC, most resistant to the action of toxin, may be due to a considerably greater content of Hly+ clones in their population.

[Role of iron ions in the production of hemolysin by toxigenic and nontoxigenic Vibrio cholerae of different serogroups]. / Rol' ionov zheleza v produktsii gemolizina toksigennymi i netoksigennymi kholernymi vibrionami razlichnykh serogrupp.

The hemolytic activity of ctx- and ctx+ V. cholerae, serogroups eltor and O39, in a medium free of FeCl3 was studied. During the cultivation in this medium, the strains of both V. cholerae serogroups proved to be capable of lysing sheep red blood cells in the Graig test, irrespective of the presence of ctx genes. The cultivation of V. cholerae ctx+ strains of both serogroups under such conditions facilitated the production of hemolysin with the same spectrum of lytic activity as hemolysin produced by ctx- strains.

Involvement of the SppA1 peptidase in acclimation to saturating light intensities in Synechocystis sp. strain PCC 6803.

The sll1703 gene, encoding an Arabidopsis homologue of the thylakoid membrane-associated SppA peptidase, was inactivated by interposon mutagenesis in Synechocystis sp. strain PCC 6803. Upon acclimation from a light intensity of 50 to 150 microE m(-2) s(-1), the mutant preserved most of its phycobilisome content, whereas the wild-type strain developed a bleaching phenotype due to the loss of about 40% of its phycobiliproteins. Using in vivo and in vitro experiments, we demonstrate that the DeltasppA1 strain does not undergo the cleavage of the L(R)(33) and L(CM)(99) linker proteins that develops in the wild type exposed to increasing light intensities. We conclude that a major contribution to light acclimation under a moderate light regime in cyanobacteria originates from an SppA1-mediated cleavage of phycobilisome linker proteins. Together with changes in gene expression of the major phycobiliproteins, it contributes an additional mechanism aimed at reducing the content in phycobilisome antennae upon acclimation to a higher light intensity.

[Interaction of Escherichia coli RNA polymerase with eukaryotic TATA-binding protein]. / Vzaimodeistvie RNK-polimerazy Escherichia coli s TATA-sviazyvaiushchim belkom éukariot.

Interaction with eukaryotic TATA-binding protein (TBP) was analyzed for natural Escherichia coli RNA polymerase or the recombinant holoenzyme, minimal enzyme, or its sigma subunit. Upon preincubation of full-sized RNA polymerase with TBP and further incubation with a constant amount of 32P-labeled phosphamide derivative of a TATA-containing oligodeoxyribonucleotide, the yield of the holoenzyme-oligonucleotide covalent complex decreased with increasing TBP concentration. This was considered as indirect evidence for complexing of RNA polymerase with TBP. In gel retardation assays, the holoenzyme, but neither minimal enzyme nor the sigma subunit, interacted with TPB, since the labeled probe formed complexes with both proteins in the reaction mixture combining TBP with the minimal enzyme or the sigma subunit. It was assumed that E. coli RNA polymerase is functionally similar to eukaryotic RNA polymerase II, and that the complete ensemble of all subunits is essential for the specific function of the holoenzyme.

[Dependence of the expression of the biological features of Vibrio cholerae on the conditions of their cultivation]. / Zavisimost' ékspressii biologicheskikh svoistv kholernykh vibrionov ot uslovii ikh kul'tivirovaniia.

The biological activity of toxigenic and non-toxigenic V. cholerae supernatants was found to depend on the cultivation medium. The use of iron-free tryptone medium made it possible to obtain supernatants of toxigenic V. cholerae with haemolytic activity and destructive action on passaged cell cultures. In the experimental infection of suckling rabbits the influence of the cultivation conditions of V. cholerae on the character and expression of their pathogenic properties was determined. The dissemination of V. cholerae into the internal organs of rabbits after their infection with both toxigenic and non-toxigenic strains correlated neither with the cultivation conditions of these strains, nor with the character of changes in the intestine of the infected animals.

[Morphology and enzyme activity of nonculturable forms of Vibrio cholerae]. / Morfologiia i fermentativnaia aktivnost' nekul'tiviruemykh form cholernykh vibrionov.

The transition of V. cholerae into the uncultivable state under experimental conditions was accompanied by gradual changes in their morphology, motility and metabolic activity. The vibrios took the oval form, lost their flagellum, motility and enzyme activity on diagnostic media. Dehydrogenase activity tested by reduction of triphenyl tetrazolium chloride, increased at the initial stages and dropped to the initial level or even lower by the end of the observation period (10 months). Similar dynamics was noted when the cytoplasmatic marker enzyme--ATPase activity was studied. Glucose catabolism in the uncultivable forms shifted towards glycolysis. During 1-2 months ctx and tcp genes could be detected in these forms by the PCR. The dynamics of the biological properties under study made it possible to find out the existence of 3 functionally different stages in the development of an uncultivable population.

Identification and characterization of SppA, a novel light-inducible chloroplast protease complex associated with thylakoid membranes.

A new component of the chloroplast proteolytic machinery from Arabidopsis thaliana was identified as a SppA-type protease. The sequence of the mature protein, deduced from a full-length cDNA, displays 22% identity to the serine-type protease IV (SppA) from Escherichia coli and 27% identity to Synechocystis SppA1 (sll1703) but lacks the putative transmembrane spanning segments predicted from the E. coli sequence. The N-terminal sequence exhibits typical features of a cleavable chloroplast stroma-targeting sequence. The chloroplast localization of SppA was confirmed by in organello import experiments using an in vitro expression system and by immunodetection with antigen-specific antisera. Subfractionation of intact chloroplasts demonstrated that SppA is associated exclusively with thylakoid membranes, predominantly stroma lamellae, and is a part of some high molecular mass complex of about 270 kDa that exhibits proteolytic activity. Treatments with chaotropic salts and proteases showed that SppA is largely exposed to the stroma but that it behaves as an intrinsic membrane protein that may have an unusual monotopic arrangement in the thylakoids. We demonstrate that SppA is a light-inducible protease and discuss its possible involvement in the light-dependent degradation of antenna and photosystem II complexes that both involve serine-type proteases.

[clpP2 gene encoding peptidase in cyanobacteria Synechocystis sp. PCC 6803 controls the sensitivity of cells to photoinhibition]. / Gen clpP2, kodiruiushchii peptidazu u tsianobakterii Synechocystis sp. PCC 6803, kontroliruet chuvstvitel'nost' kletok k fotoingibirovaniiu.

A homozygous insertion mutant with the inactivated clpP2 gene, which encodes the proteolytic subunit of ATP-dependent peptidase, was obtained in the unicellular cyanobacterium Synechocystis sp. PCC 6803. The mutant cannot grow under photoautotrophic conditions, but cells grown under heterotrophic conditions in a glucose-containing medium have active photosystems I and II (PS I and PS II). The loss of capacity for photoautotrophic growth is determined by a high sensitivity of mutant cells to the inactivating effect of light. Their incubation under light with an intensity above 10 microE m-2 s-1 inhibits cell growth in culture and causes degradation of photosynthetic pigments. It is proposed that the ClpP2 peptidase is involved in the protection of Synechocystis 6803 cells from photoinhibition.