Functional pri-miR-34b/c rs4938723 and KRAS 3'UTR rs61764370 SNPs: Novel phenotype modifiers in Li-Fraumeni Syndrome?

PURPOSE: Li-Fraumeni Syndrome (LFS) is a rare cancer predisposing condition caused by germline pathogenic TP53 variants, in which core tumors comprise sarcomas, breast, brain and adrenocortical neoplasms. Clinical manifestations are highly variable in carriers of the Brazilian germline founder variant TP53 p.R337H, possibly due to the influence of modifier genes such as miRNA genes involved in the regulation of the p53 pathway. Herein, we investigated the potential phenotypic effects of two miRNA-related functional SNPs, pri-miR-34b/c rs4938723 and 3'UTR KRAS rs61764370, in a cohort of 273 LFS patients from Southern and Southeastern Brazil. METHODS: The genotyping of selected SNPs was performed by TaqMan® allelic discrimination and subsequently custom TaqMan® genotyping results were confirmed by Sanger sequencing in all SNP-positive LFS patients. RESULTS: Although the KRAS SNP showed no effect as a phenotype modulator, the rs4938723 CC genotype was significantly associated with development of LFS non-core tumors (first tumor diagnosis) in p.R337H carriers (p = 0.039). Non-core tumors were also more frequently diagnosed in carriers of germline TP53 DNA binding domain variants harboring the rs4938723 C variant allele. Previous studies described pri-miR-34b/c rs4938723 C as a risk allele for sporadic occurrence of thyroid and prostate cancers (non-core tumors of the LFS spectrum). CONCLUSION: With this study, we presented additional evidence about the importance of analyzing miRNA genes that could indirectly regulate p53 expression, and, therefore, may modulate the LFS phenotype, such as those of the miR-34 family.

Nisin influence on the expression of Listeria monocytogenes surface proteins.

In this work, a comparative analysis of the peripheral cell component (PCC) proteins of Listeria monocytogenes was carried out. The study was conducted on two set of samples consisting of bacteria treated with sub-lethal concentration of nisin and untreated bacteria as control. PCC proteins were extracted by Tris-Urea-EDTA treatment and then subjected to trypsin digestion and mass spectrometry analysis. The whole cell proteome was analyzed through label-free quantitative proteomics approach. Proteomic analysis was carried out using OrbiTrap Mass Spectrometer coupled to nanoflow liquid chromatography. The treatment with sub-lethal nisin concentration resulted in 62 up regulated and 97 down regulated proteins compared to untreated samples. Using PSORTb 3.0, 19 and 18 surface proteins were detected among the up regulated and down regulated proteins, respectively. Proteins related with increased biofilm formation by L.monocytogenes, such as moonlight proteins of the pyruvate dehydrogenase complex and flagellin-related proteins, were identified as up regulated surface proteins. Proteins associated with virulence of L.monocytogenes, including listeriolysin O, internalin B and actin assembly-inducing protein, were detected among the down regulated proteins. To confirm proteomics data, increased production of biofilm was experimentally confirmed in nisin-treated cells through crystal violet method. BIOLOGICAL SIGNIFICANCE: Proteosurfaceomics can be defined as the "omics" science applied to the proteins of the peripheral cell component (PCC). The surface proteins of Listeria monocytogenes, an important foodborne pathogen were investigated after treatment with nisin, a bacteriocin approved as a natural food preservative by regulatory agencies. Recent cases of nisin tolerance by Listeria spp. were documented, and deeper studies on the molecular process behind the bacterial survival may help in both understanding the development of tolerance process and comparing nisin effect with other antimicrobial compounds.

Cestode strobilation: prediction of developmental genes and pathways.

BACKGROUND: Cestoda is a class of endoparasitic worms in the flatworm phylum (Platyhelminthes). During the course of their evolution cestodes have evolved some interesting aspects, such as their increased reproductive capacity. In this sense, they have serial repetition of their reproductive organs in the adult stage, which is often associated with external segmentation in a developmental process called strobilation. However, the molecular basis of strobilation is poorly understood. To assess this issue, an evolutionary comparative study among strobilated and non-strobilated flatworm species was conducted to identify genes and proteins related to the strobilation process. RESULTS: We compared the genomic content of 10 parasitic platyhelminth species; five from cestode species, representing strobilated parasitic platyhelminths, and five from trematode species, representing non-strobilated parasitic platyhelminths. This dataset was used to identify 1813 genes with orthologues that are present in all cestode (strobilated) species, but absent from at least one trematode (non-strobilated) species. Development-related genes, along with genes of unknown function (UF), were then selected based on their transcriptional profiles, resulting in a total of 34 genes that were differentially expressed between the larval (pre-strobilation) and adult (strobilated) stages in at least one cestode species. These 34 genes were then assumed to be strobilation related; they included 12 encoding proteins of known function, with 6 related to the Wnt, TGF-ß/BMP, or G-protein coupled receptor signaling pathways; and 22 encoding UF proteins. In order to assign function to at least some of the UF genes/proteins, a global gene co-expression analysis was performed for the cestode species Echinococcus multilocularis. This resulted in eight UF genes/proteins being predicted as related to developmental, reproductive, vesicle transport, or signaling processes. CONCLUSIONS: Overall, the described in silico data provided evidence of the involvement of 34 genes/proteins and at least 3 developmental pathways in the cestode strobilation process. These results highlight on the molecular mechanisms and evolution of the cestode strobilation process, and point to several interesting proteins as potential developmental markers and/or targets for the development of novel antihelminthic drugs.

Circulating cells and exosomes in acute myelogenous leukemia and their role in disease progression and survival.

Acute myelogenous leukemia (AML) is an aggressive hematological malignancy associated with high rates of mortality. This incidence is due to the complexity in which the AML cells interact with other healthy human cells. These phenomena create an environment that favors the expansion of leukemic cells, which will affect the patient's prognosis. An important aspect is the ability of AML cells to evade immune responses via targeting and signaling immune cells to suppress anti-tumor responses. Many studies have reported that associations among components in the peripheral bloodstream might modulate leukemic progression because AML survival is a fundamental step for recolonizing bone marrow after allogeneic hematopoietic stem cell (HSC) transplantation or chemotherapy. Therefore, we collected the most important data about components that circulate with leukemic blasts and contribute to their survival and proliferation. We also discuss clinical approaches that could be conducted to more effectively treat the disease.

Proteomic study of Enterococcus durans LAB18S growing on prebiotic oligosaccharides.

This study evaluates the influence of prebiotic carbohydrates, namely fructooligosaccharides (FOS) and galactooligosaccharides (GOS), on the protein expression of Enterococcus durans LAB18S. The strain was cultivated in 10 g L-1 FOS, GOS or glucose (control) and cellular proteins were extracted for mass spectrometry analysis. A total of 771 proteins were identified and 135 E. durans proteins were validated by the Scaffold algorithm. The proteins were functionally categorized according to Gene Ontology terms. Both FOS and GOS were used as carbon source by E. durans LAB18S, upregulating the production of proteins that may be associated with intestinal mucosa adhesion, carbohydrate and nitrogen metabolism, and stress response. Cells grown with GOS showed an increased expression of the cell division protein divIVA, EF-Tu and glyceraldehyde 3-phosphate dehydrogenase that have been associated with epithelial cell adhesion. The use of FOS stimulated the production of proteins related to amino acid metabolism and energy conversion, and ClpX protein, which plays an important role in protein turnover. The results of this study indicate that FOS and GOS can be metabolized by E. durans and stimulate the microorganism to produce proteins related to some desirable characteristics for a probiotic strain.

Supporting data for comparative proteomic analysis of Listeria monocytogenes ATCC 7644 exposed to a sublethal concentration of nisin.

Here we provide the LC-MS/MS data from a comparative analysis of Listeria monocytogenes ATCC 7644 treated and non-treated with a sublethal concentration of nisin (10(-3) mg/mL). Protein samples were analyzed by multidimensional protein identification technology (MudPIT) approach, in an off-line configuration. The raw MS/MS data allowed the detection of 49,591 spectra which resulted in 576 protein identifications. After Scaffold validation, 179 proteins were identified with high confidence. A label-free quantitative analysis based of normalized spectral abundance factor (NSAF) was used and 13 proteins were found differentially expressed between nisin-treated and non-treated cells. Gene ontology analysis of differentially expressed proteins revealed that most of them are correlated to metabolic process, oxidative stress response mechanisms and molecular binding. A detailed analysis and discussion of these data may be found in Miyamoto et al. [1].

Comparative proteomic analysis of Listeria monocytogenes ATCC 7644 exposed to a sublethal concentration of nisin.

Listeria monocytogenes infections have been frequently reported in many food poisoning outbreaks around the world. In this work, the protein repertoires of L. monocytogenes ATCC 7644 cells treated or not with a 10(-3)mg/mL nisin sublethal concentration, established by antimicrobial susceptibility tests, were analyzed by LC-MS/MS. Overall, 179 proteins were identified, 9 of them more abundant in nisin-treated samples, and 4 more abundant in non-treated control samples. In nisin treated cells, proteins associated to oxidative stress response showed higher abundance. Also, the higher abundance of an enzyme related to the production of cell membrane lipids upon nisin exposure is suggestive of both a failure in conventional cell division mechanism and the activation of an alternative L-form mediated division mechanism. Finally, flagellar and motility proteins' overexpression upon nisin exposure is indicative of increased bacterial motility in response to the bacteriocin. Taken together, these results provide new insights on nisin effects on L. monocytogenes cells and on how this bacterium may overcome a bacteriocin-containing environment. BIOLOGICAL SIGNIFICANCE: The antimicrobial mechanism of nisin on target bacterial cells has been extensively studied since discovery of this bacteriocin. The nisin pore-forming mechanism is mediated by its binding to the pyrophosphate portion of membrane lipid II [1], but some evidences point out to alternative mechanisms. Results from assays with mutacin 1140 hybrids [2] showed that the portion of nisin that is not involved with lipid II binding could damage the bacterial cell, independently of pore formation [3,4]. Moreover, there are insufficient data to explain how nisin affects the bacterial survival. In this scenario, proteomics is an interesting approach, as a comparison between treated and untreated cells may provide insights of both antimicrobial mechanisms of action and bacterial response mechanisms [5].

Prevalência de resistência constitutiva e induzível à clindamicina em cocos Gram positivos isolados de pacientes hospitalizados em Santa Maria, RS / Prevalence of clindamycin constitutive and inducible resistance in Gram positive cocci isolated from hospitalized patients in Santa Maria, RS

O objetivo deste estudo foi determinar a prevalência de resistência constitutiva e induzível à clindamicina em pacientes hospitalizados . Foram analisados 63 isolados de diversos sítios infecciosos de pacientes internados em um hospital privado de Santa Maria - RS, Brasil. Para determinação dos mecanismos de resistência foi utilizado o método de difusão em disco (D-teste). Foi verificado que três cepas de Staphylococcus aureus (4,8%) apresentaram-se positivas no D-teste, ou seja, exibiram resistência induzível à clindamicina (MLSBi), possivelmente pela presença do gene em (A), em microrganismos dessa espécie. Ademais, foi observado a presença de resistência constitutiva (MLSBc)em 22 amostras, caracterizando, possivelmente, a presença do gene em (B), em Streptococcus sp e em (C), em Staphylococcus sp. A prevalência do mecanismo de resistência, mediado por bomba de efluxo, em 19 amostras foi caracterizada, provavelmente pela presença do gene mrsA. A introdução de mudanças, na rotina do laboratório, através da inclusão do D-teste deve ser encorajada. A pesquisa desses mecanismos de resistência são de elevada importância na escolha e eficácia terapêutica, pois representam, na prática, uma maior segurança para os pacientes e evitam a troca desnecessária de medicação e, consequentemente, a falha terapêutica.