RESUMEN
Shiga toxin-producing Escherichia coli (STEC) is a known food pathogen, which main reservoir is the intestine of ruminants. The abundance of different STEC lineages in nature reflect a heterogeneity that is characterised by the differential expression of certain genotypic characteristics, which in turn are influenced by the environmental conditions to which the microorganism is exposed. Bacterial homeostasis and stress response are under the control of the alarmone guanosine tetraphosphate (ppGpp), which intrinsic levels varies across the E. coli species. In the present study, 50 STEC isolates from healthy sheep were evaluated regarding their ppGpp content, cytotoxicity and other relevant genetic and phenotypic characteristics. We found that the level of ppGpp and cytotoxicity varied considerably among the examined strains. Isolates that harboured the stx2 gene were the least cytotoxic and presented the highest levels of ppGpp. All stx2 isolates belonged to phylogroup A, while strains that carried stx1 or both stx1 and stx2 genes pertained to phylogroup B1. All but two stx2 isolates belonged to the stx2b subtype. Strains that belonged to phylogroup B1 displayed on average low levels of ppGpp and high cytotoxicity. Overall, there was a negative correlation between cytotoxicity and ppGpp.
Asunto(s)
Guanosina Pentafosfato/metabolismo , Guanosina Tetrafosfato/metabolismo , Enfermedades de las Ovejas/microbiología , Ovinos/microbiología , Escherichia coli Shiga-Toxigénica/genética , Factores de Virulencia/genética , Animales , Reservorios de Enfermedades , Infecciones por Escherichia coli/microbiología , Variación Genética , Enfermedades de las Ovejas/epidemiología , Toxina Shiga II/metabolismo , Escherichia coli Shiga-Toxigénica/inmunología , Escherichia coli Shiga-Toxigénica/aislamiento & purificaciónRESUMEN
The involvement of anion channels in the mechanism of the acrosome reaction (AR) was investigated. The AR was induced by Ca2+ or by addition of the Ca2+ ionophore A23187. The occurrence of AR was determined by following the release of acrosin from the cells. In order to investigate the role of anion channels in the AR, several anion-channel inhibitors were tested, mainly DIDS (4,4'-diisothiocyanostilbene-2,2'-disulfonic acid). Other blockers, like SITS (4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid), furosemide, probenecid and pyridoxal 5-phosphate, were also tested. We found that DIDS binds covalently to sperm plasma membrane in a time- and concentration-dependent manner. Maximal binding occurs after 2 h with 0.3 mM DIDS. DIDS and SITS inhibit AR in a concentration-dependent manner. The IC50 of DIDS and SITS in the presence of A23187 is 0.15 and 0.22 mM, respectively. Tributyltin chloride (TBTC), an Cl-/OH- exchanger, partially overcomes DIDS inhibition of the AR. HCO3- is required for a maximal acrosin release and Ca(2+)-uptake, in the presence or absence of A23187. It is known that HCO3- activates adenylate cyclase and therefore, increases the intracellular level of cAMP. The inhibition of the AR by DIDS decreases from 95 to 50% when (dibutyryl cyclic AMP (dbcAMP) was added, i.e., HCO3- is no longer required while elevating the level of cAMP in an alternative way. Moreover, we show that the stimulatory effect of HCO3- on Ca(2+)-uptake is completely inhibited by DIDS. We conclude that DIDS inhibits AR by blocking anion channels, including those that transport HCO3- into the cell.
Asunto(s)
Acrosina/análisis , AMP Cíclico/análisis , Canales Iónicos/fisiología , Espermatozoides/fisiología , Ácido 4,4'-Diisotiocianostilbeno-2,2'-Disulfónico , Ácido 4-Acetamido-4'-isotiocianatostilbeno-2,2'-disulfónico/análogos & derivados , Ácido 4-Acetamido-4'-isotiocianatostilbeno-2,2'-disulfónico/metabolismo , Ácido 4-Acetamido-4'-isotiocianatostilbeno-2,2'-disulfónico/farmacología , Animales , Bicarbonatos/farmacología , Bucladesina/farmacología , Calcimicina/farmacología , Calcio/metabolismo , Calcio/farmacología , Bovinos , Membrana Celular/metabolismo , Relación Dosis-Respuesta a Droga , Canales Iónicos/efectos de los fármacos , Masculino , Compuestos Orgánicos de Estaño/farmacología , Espermatozoides/efectos de los fármacosRESUMEN
The addition of both live and ultraviolet-inactivated preparations of human T lymphotropic virus type I (HTLV-I) and HIV to cultures of human peripheral lymphocytes impeded the ability of these cells to respond to phytohaemagglutinin (PHA). This inhibition depended on the concentration of the virus and seemed due, in part at least, to interference with the generation of interleukin-2 (IL-2) activity in the PHA-stimulated cultures. However, the addition of exogenous IL-2 did not effectively restore the lymphocyte proliferative responsiveness of cells which had been co-incubated with these human retroviruses. Exposure to the viruses did not affect expression on co-incubated cells of the Tac antigen, an epitope of the IL-2 receptor, as determined by an indirect immunofluorescence assay. These results suggest that one mechanism through which human retroviruses may be able to impede cellular proliferative responsiveness is interference with the ability of target cells to respond to IL-2, even though IL-2 receptors continue to be expressed under the conditions tested.
Asunto(s)
Deltaretrovirus/inmunología , Interleucina-2/farmacología , Activación de Linfocitos , VIH/inmunología , Humanos , Técnicas In Vitro , Fitohemaglutininas/farmacología , Receptores Inmunológicos/inmunología , Receptores de Interleucina-2 , Linfocitos T/inmunologíaRESUMEN
OBJECTIVE: To describe the rate of development of in vitro HIV resistance to zidovudine (ZDV) and its prognostic implications within the Multicentre Canadian AZT Trial (MCAT). METHODS: HIV-infected subjects in Centers for Disease Control (CDC) stages IIB, III and IVC-2 with CD4 cell counts > 270 x 10(6)/l were treated with ZDV as part of a dose-range study. Participating volunteers underwent prospective clinical and laboratory evaluations at regular intervals. Viral cultures and sensitivity testing were performed every 12 weeks in a predefined subset of 50 volunteers. An isolate was designated ZDV-resistant if it had a median inhibitory concentration (IC50) for ZDV at least 50-fold higher than that of virus isolated from the same subject before initiation of antiviral chemotherapy. The relationship between resistance and subsequent disease progression was studied using the Mantel and Byar method, for which, at each instance of disease progression, 2 x 2 tables classifying progression versus resistance status were constructed. The observed number of progressions was compared with that expected under the null hypothesis using Mantel-Haenszel methods adjusted for baseline CD4:CD8 ratio. RESULTS: The Kaplan-Meier estimate for the cumulative development of in vitro resistance was 64% [95% confidence interval (CI), 41-78] at 180 weeks. Baseline CD4:CD8 ratio was negatively associated (P = 0.10) with the subsequent development of resistance (proportional hazard, 0.44; 95% CI, 0.17-1.10). After adjusting for baseline CD4:CD8 ratio, the numbers of observed and expected progressions following the development of resistance were 15 and 7.6, respectively (P = 0.008). A similar relative risk of progression between resistant and non-resistant states was found in the two CD4:CD8 strata; observed and expected progressions were 4 and 2.3 and 11 and 5.2 in the high and low CD4:CD8 strata, respectively. CONCLUSIONS: In vitro resistance to ZDV developed in 64% of subjects after 180 weeks of ZDV therapy. Lower CD4:CD8 ratio at baseline was associated with faster development of resistance. In addition, the development of resistance was found to be a marker of subsequent disease progression. This association persisted after adjustment for baseline CD4:CD8 ratio. Whether in vitro resistance to ZDV is merely a surrogate marker or a determinant of disease progression remains to be established.
Asunto(s)
Infecciones por VIH/tratamiento farmacológico , VIH-1/efectos de los fármacos , Zidovudina/uso terapéutico , Adulto , Análisis Mutacional de ADN , Farmacorresistencia Microbiana , Infecciones por VIH/microbiología , VIH-1/genética , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Estudios Prospectivos , Factores de TiempoRESUMEN
OBJECTIVE: Prolonged treatment with antiretroviral drugs results in the selection of HIV-1 variants with mutations conferring resistance to nucleoside and non-nucleoside reverse transcriptase inhibitors (NRTI and NNRTI) or to protease inhibitors (PI). There is serious concern about transmission of resistant viruses to newly infected persons. This study monitored the prevalence of resistant viruses in individuals undergoing primary HIV infection. DESIGN: Resistance testing was performed on 81 individuals infected between 1997 and 1999 by injecting drug use (n =21), sexual (n = 56), or unknown (n = 4) transmission. METHODS: Automated sequencing was used to genotype the reverse transcriptase (RT) and protease regions of virus isolated from patients' plasma. The phenotypic susceptibility of stimulated peripheral blood mononuclear cells to antiretroviral drugs was assayed. Line probe assays detected quasispecies variations in wild-type and mutated RT codons. RESULTS: A high prevalence of PI and RT genotypic variants, associated with high-level resistance to antiretroviral drugs, was observed in individuals newly infected by injecting drug use (PI = 24%, RT = 24%) or sexual transmission (PI = 12%, RT = 22%). The PI mutations, L101, V82A, and L90M, were found in 10.5, 3 and 4% of cases, respectively; whereas for RT, primary mutations at positions T215Y (zidovudine), M184V (lamivudine), T69D/A (zalcitabine), and K103N (multi-NNRTI) were present in 8, 5, 4, and 4% of subjects, respectively. Resistance to NRTI was demonstrated by phenotypic, genotypic, and line probe analyses. Transmission of multidrug (NRTI/NNRTI/PI) resistance in eight subjects (9.9%) was confirmed by showing that source partners possessed viruses of similar genotype. CONCLUSIONS: The transmission of drug-resistant HIV is a serious problem that merits further attention by public health officials as well as virologists and clinicians.
Asunto(s)
Fármacos Anti-VIH/farmacología , Infecciones por VIH/transmisión , VIH-1/efectos de los fármacos , Abuso de Sustancias por Vía Intravenosa/complicaciones , Fármacos Anti-VIH/uso terapéutico , Canadá/epidemiología , Codón , Farmacorresistencia Microbiana/genética , Femenino , Genotipo , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/epidemiología , Proteasa del VIH/genética , Inhibidores de la Proteasa del VIH/farmacología , Inhibidores de la Proteasa del VIH/uso terapéutico , Transcriptasa Inversa del VIH/genética , VIH-1/genética , Humanos , Estudios Longitudinales , Masculino , Mutación , Fenotipo , Inhibidores de la Transcriptasa Inversa/farmacología , Inhibidores de la Transcriptasa Inversa/uso terapéutico , Abuso de Sustancias por Vía Intravenosa/epidemiología , Abuso de Sustancias por Vía Intravenosa/virologíaRESUMEN
We conducted detailed virological evaluations of 16 HIV-1-infected paediatric patients treated with 3TC (lamivudine) monotherapy. High-level phenotypic resistance against this compound (up to 2,500-fold) was seen in virtually all cases, usually within 8-12 weeks of initiation of therapy. This was concomitant with the appearance of the M184V mutation in viral reverse transcriptase, previously shown to be responsiblefor such resistance. Viral burden fell in virtually all cases after commencement of therapy, and remained below baseline in each instance studied, despite a rebound effect and the appearance of drug resistance. Viral isolates from some patients underwent a switch from a non-syncytium-inducing (NSI) to a syncytium-inducing (SI) phenotype during the course of the study, although no relationship was apparent between dose of drug employed, time to development of drug resistance or time of appearance of SI phenotype.
Asunto(s)
Fármacos Anti-VIH/farmacología , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , Lamivudine/farmacología , Inhibidores de la Transcriptasa Inversa/farmacología , Fármacos Anti-VIH/uso terapéutico , Niño , Codón , Farmacorresistencia Microbiana/genética , Femenino , Infecciones por VIH/tratamiento farmacológico , Transcriptasa Inversa del VIH/genética , VIH-1/genética , VIH-1/aislamiento & purificación , Humanos , Lamivudine/uso terapéutico , Estudios Longitudinales , Masculino , Mutación , Fenotipo , Inhibidores de la Transcriptasa Inversa/uso terapéutico , Carga ViralRESUMEN
Tumors which are induced in chickens by Rous sarcoma virus (RSV) usually grow progressively for several weeks and then regress. pp60src kinase activity was found to be reduced in cultured tumor cells which derived from regressing sarcomas by 60-75% as compared to progressively growing neoplasms. We further found that a cellular pp89 which co-precipitated with pp60src was largely diminished in tumor cells derived from regressing as compared with progressively growing sarcomas. In contrast, immunoprecipitation analyses from membrane or cytosol fractions did not reveal any significant differences between these 2 types of tumor cells in distribution of pp60src. Moreover, partial proteolysis or isoelectric focussing of labelled immunoprecipitated pp60src did not reveal major differences between the 2 tumor cell types. These data indicate that the lack of pp89 cell binding to pp60src in regressing sarcoma cells correlates well with the diminution of the pp60src kinase activity in these cells, but does not affect the transport of pp60src to the plasma membrane.
Asunto(s)
Pollos/microbiología , Regresión Neoplásica Espontánea/enzimología , Proteínas Proto-Oncogénicas/metabolismo , Sarcoma Aviar/enzimología , Animales , Regresión Neoplásica Espontánea/metabolismo , Proteínas Proto-Oncogénicas pp60(c-src) , Sarcoma Aviar/metabolismoRESUMEN
The widespread use of antiretroviral agents (ARVs) and the growing occurrence of HIV strains resistant to these drugs have given rise to serious concerns regarding the transmission of resistant viruses to newly infected persons. Plasma viral RNA from 80 individuals newly infected between 1997 and 1999 was genotyped by automated sequencing to analyze the profile of viruses resistant to nucleoside and non-nucleoside reverse transcriptase inhibitors (NRTIs and NNRTIs) and to protease inhibitors (PIs). The prevalence of mutations that conferred primary resistance to PIs (L10I, D30Y, V82A, L90M) was 15% of the cohort. RT genotypic variants, associated with high-level resistance to ARVs, were observed in 21% of individuals, including NRTI, NNRTI and multidrug (MDR) resistance in 6, 5, and 10% of cases, respectively. The phenotypic susceptibility of viral isolates to ARVs was also assayed and showed transmission of high-level resistance to ZDV, 3TC, and PIs in those individuals with MDR. The transmission of drug-resistant HIV genotypic variants is a serious problem that merits further attention by public health officials, virologists, and clinicians.
Asunto(s)
Fármacos Anti-VIH/farmacología , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , Fármacos Anti-VIH/uso terapéutico , Farmacorresistencia Microbiana , Femenino , Frecuencia de los Genes , Infecciones por VIH/tratamiento farmacológico , Proteasa del VIH/efectos de los fármacos , Proteasa del VIH/genética , Transcriptasa Inversa del VIH/antagonistas & inhibidores , Transcriptasa Inversa del VIH/genética , VIH-1/enzimología , VIH-1/genética , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Mutación , ARN Viral/análisis , Inhibidores de la Transcriptasa Inversa/farmacología , Inhibidores de la Transcriptasa Inversa/uso terapéuticoRESUMEN
Rous sarcoma virus induced chicken tumors usually grow progressively for several weeks and then regress. pp60src kinase activity was reduced in cultured tumor cells which derived from regressing sarcomas by 60-75% as compared to progressively-growing neoplasms. We have demonstrated the presence of inhibitory activity to pp60src kinase activity in extracts of regressing tumor cells. Such extracts were able to reduced by 60% the level of kinase activity found in progressively-growing tumors, as measured in a IgG phosphorylation assay. Immunoprecipitation analyses of membrane or cytosol fractions did not reveal any significant differences between these two types of tumor cells in distribution of pp60src. The pp60src kinase of progressively-growing tumor cells was more sensitive than that of "regressor" cells to inhibition by P1, P4 di-(adenosine-5') tetraphosphate (Ap4 A) and to heat inactivation. These results suggest that the reduction of pp60src kinase activity in regressing neoplasms might be due to the presence of inhibitors in such cells, but that the basal levels of activity which remain are relatively resistant to further perturbation by heat and/or chemical antagonists.
Asunto(s)
Virus del Sarcoma Aviar/genética , Proteína Oncogénica pp60(v-src)/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Sarcoma Aviar/enzimología , Animales , Pollos , Cinética , Regresión Neoplásica Espontánea , Sarcoma Aviar/patologíaRESUMEN
The use of zidovudine (ZDV) and other forms of nucleoside therapy, including dideoxyinosine (ddI), to treat HIV-infected individuals has led to both longer survival and improved quality of life. However, ZDV-resistant variants of HIV-1 can be isolated from patients undergoing prolonged therapy with this drug. HIV drug resistance against ZDV, ddI and other nucleosides is attributable to a series of point mutations within the pol gene of HIV-1 that encodes the viral enzyme, reverse transcriptase (RT). This is not surprising, since the virus is known to replicate at high rates in infected individuals; moreover the RT which mediates transcription of proviral DNA from viral genomic RNA is known to be highly error-prone. Thus, mutants of HIV-1, which possess a drug resistance phenotype and genotype, may be expected to emerge under the selective pressure of long-term anti-viral chemotherapy. HIV drug resistance occurs most commonly in individuals with low CD4 counts, who have progressed to more serious forms of disease. Moreover, viruses obtained from patients with AIDS generally display higher levels of resistance, relative to pre-treatment isolates, than do viruses from patients with more limited illness. Although observations of drug resistance can be correlated with disease progression and a weakened immune system, it is still unclear whether a cause and effect relationship exists. Current clinical research is designed to answer this question while testing the notion that combinations of nucleosides and immuno-stimulatory drugs may provide important clinical benefits.
Asunto(s)
VIH-1/efectos de los fármacos , Zidovudina/farmacología , Reacciones Cruzadas , Farmacorresistencia Microbiana/genética , Transcriptasa Inversa del VIH , VIH-1/inmunología , Humanos , Recuento de Leucocitos , ADN Polimerasa Dirigida por ARN/metabolismo , Linfocitos T/inmunología , Replicación Viral/efectos de los fármacosRESUMEN
Shiga toxin-producing Escherichia coli (STEC) is a known food pathogen, which main reservoir is the intestine of ruminants. The abundance of different STEC lineages in nature reflect a heterogeneity that is characterised by the differential expression of certain genotypic characteristics, which in turn are influenced by the environmental conditions to which the microorganism is exposed. Bacterial homeostasis and stress response are under the control of the alarmone guanosine tetraphosphate (ppGpp), which intrinsic levels varies across the E. coli species. In the present study, 50 STEC isolates from healthy sheep were evaluated regarding their ppGpp content, cytotoxicity and other relevant genetic and phenotypic characteristics. We found that the level of ppGpp and cytotoxicity varied considerably among the examined strains. Isolates that harboured the stx2 gene were the least cytotoxic and presented the highest levels of ppGpp. All stx2 isolates belonged to phylogroup A, while strains that carried stx1 or both stx1 and stx2 genes pertained to phylogroup B1. All but two stx2 isolates belonged to the stx2b subtype. Strains that belonged to phylogroup B1 displayed on average low levels of ppGpp and high cytotoxicity. Overall, there was a negative correlation between cytotoxicity and ppGpp.
RESUMEN
The Pst system is a high-affinity inorganic phosphate transporter found in many bacterial species. Streptococcus mutans, the etiological agent of tooth decay, carries a single copy of the pst operon composed of six cistrons (pstS, pstC1, pstC, pstB, smu.1134 and phoU). Here, we show that deletion of pstS, encoding the phosphate-binding protein, reduces phosphate uptake and impairs cell growth, which can be restored upon enrichment of the medium with high concentrations of inorganic phosphate. The relevance of Pst for growth was also demonstrated in the wild-type strain treated with an anti-PstS antibody. Nevertheless, a reduced ability to bind to saliva-coated surfaces was observed, along with the reduction of extracellular polysaccharide production, although no difference on pH acidification was observed between mutant and wild-type strains. Taken together, the present data indicate that the S. mutans Pst system participates in phosphate uptake, cell growth and expression of virulence-associated traits.
Asunto(s)
Adhesión Bacteriana/fisiología , Proteínas de Transporte de Fosfato/fisiología , Streptococcus mutans/fisiología , Transportadoras de Casetes de Unión a ATP/análisis , Adenosina Trifosfatasas/análisis , Proteínas Bacterianas/análisis , Película Dental/metabolismo , Técnicas de Inactivación de Genes , Silenciador del Gen , Genes Bacterianos/genética , Humanos , Concentración de Iones de Hidrógeno , Proteínas de Transporte de Membrana/análisis , Mutación/genética , Operón/genética , Proteínas de Transporte de Fosfato/genética , Proteínas de Unión a Fosfato/análisis , Fosfatos/análisis , Polisacáridos Bacterianos/metabolismo , Análisis de Secuencia de ADN , Streptococcus mutans/genética , Streptococcus mutans/metabolismo , Factores de Transcripción/análisis , Virulencia/genéticaRESUMEN
The effect of thioacetamide (TA), an hepatotoxic and hepatocarcinogenic compound, on the expression and activity of the cytosolic enzyme glutathione-S-transferase (GST) was studied in rat liver. Four h following the administration of 14C-labeled thioacetamide (50 mg/Kg), several subunits of GST were found to be radioactively labeled. A single sublethal dose of TA (250 mg/Kg) decreased by three-fold the expression of class alpha GST at 24-48 h of treatment, but did not significantly affect the transcription of class mu GST. The activity of the enzyme toward 1-chloro-2,4-dinitrobenzene was mildly inhibited (66% of the control) by a 24 h TA treatment and gradually increased thereafter. It is proposed that the covalent binding of TA or its derivative to the GST subunits does not affect the activity of the enzyme. Nevertheless, GST activity inhibition is due to the deleterious effect of TA on GST transcription.
Asunto(s)
Glutatión Transferasa/metabolismo , Hígado/efectos de los fármacos , Hígado/enzimología , Tioacetamida/farmacología , África del Norte , Secuencia de Aminoácidos , Animales , Radioisótopos de Carbono , Cromatografía en Gel , Citosol/enzimología , Electroforesis en Gel de Poliacrilamida , Femenino , Glutatión Transferasa/antagonistas & inhibidores , Glutatión Transferasa/genética , Datos de Secuencia Molecular , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Alineación de Secuencia , Tioacetamida/metabolismo , Factores de Tiempo , Transcripción Genética/efectos de los fármacosRESUMEN
Starving of Escherichia coli cells for inorganic orthophosphate (Pi) results in the accumulation of spoT-dependent ppGpp and induces the expression of genes of the PHO regulon. In a delta relA delta spoT strain that is unable to accumulate ppGpp the expression of two genes (phoA and pstS) that belong to the PHO regulon is impaired, even in constitutive mutants. The transcription of phoA and of pstS is not affected in the ppGpp0 strain, and therefore this impairment is due to a post-transcriptional defect. Conversely, overexpression of ppGpp inhibits transcription of these two PHO regulon genes. In phoB mutants, the accumulation of ppGpp during Pi starvation is diminished, suggesting that PhoB or one of the PHO products is involved in the control of ppGpp accumulation. We propose that the presence of ppGpp in the cell, but not its accumulation as a result of the starvation stress, is important for the expression of the PHO genes.
Asunto(s)
Proteínas Bacterianas/genética , Proteínas de Escherichia coli , Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Guanosina Tetrafosfato/fisiología , Proteínas de Unión Periplasmáticas , Regulón/genética , Fosfatasa Alcalina/biosíntesis , Fosfatasa Alcalina/genética , Proteínas Bacterianas/biosíntesis , Proteínas Portadoras/biosíntesis , Proteínas Portadoras/genética , Proteínas de la Membrana/biosíntesis , Proteínas de la Membrana/genética , Proteínas de Unión a Fosfato , Fosfatos/metabolismo , Fosfatos/farmacología , Transcripción GenéticaRESUMEN
The genes encoding alkaline phosphatase (phoA) and the inducible inorganic phosphate transport system Pst (pstS,C,A,B,U) belong to the PHO regulon. Mutants of Escherichia coli lacking the global regulatory protein integration host factor (IHF) show an increased level of alkaline phosphatase and a decreased level of Pst. IHF binds weakly but specifically to a DNA fragment containing the promoter region of the pst operon but does not bind to a fragment that includes the promoter region of phoA. It is proposed that IHF is a positive regulator of the pst operon and as such controls indirectly the expression of phoA.
Asunto(s)
Fosfatasa Alcalina/genética , Proteínas Bacterianas/fisiología , Proteínas Portadoras/genética , Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Secuencia de Bases , Sitios de Unión , Factores de Integración del Huésped , Datos de Secuencia Molecular , Proteínas de Unión a Fosfato , Regiones Promotoras GenéticasRESUMEN
The pst operon of Escherichia coli, which encodes the phosphate-specific transport system, is composed of five genes, pstS, pstC, pstA, pstB and phoU, whose transcription is induced by phosphate starvation. A phosphate-regulated promoter located upstream of the most proximal gene ( pstS) controls the transcription of the entire operon. Though the full-length pst mRNA could be detected by an improved RT-PCR protocol, Northern analysis using several pst-specific probes failed to reveal this transcript. Instead, smaller but distinct pst mRNA species were evident. Primer-extension experiments localized the 5' ends of pst mRNAs within the operon. The data suggest that the full-length mRNA is rapidly processed post-transcriptionally.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/genética , Proteínas Bacterianas/genética , Escherichia coli/genética , Operón/genética , Transcripción Genética , Fosfatasa Alcalina/metabolismo , Northern Blotting , Cartilla de ADN/química , Familia de Multigenes/genética , Fosfatos/metabolismo , Regiones Promotoras Genéticas , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Cells of Escherichia coli which enter a phase of starvation for Pi induce the synthesis of the nucleotide guanosine 3',5'-bispyrophosphate (ppGpp). This induction is relA independent but depends on the spoT gene product. A mutant unable to produce ppGpp is impaired in the expression of two genes which belong to the pho regulon, a defect which is dependent on the product of spoT. We suggest that ppGpp is essential for the proper induction of the genes which belong to the pho regulon.
Asunto(s)
Escherichia coli/metabolismo , Guanosina Tetrafosfato/biosíntesis , Fosfatos/deficiencia , Regulón/genética , Fosfatasa Alcalina/biosíntesis , Proteínas Bacterianas/biosíntesis , Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Pirofosfatasas/biosíntesisRESUMEN
We have shown that the spermicidal agent benzalkonium chloride can exert a direct inhibitory effect on the viral reverse transcriptase activity of human immunodeficiency virus type 1 (HIV-1) when utilized at concentrations of 0.05% and higher. Exposure of HIV-1 to this disinfectant at concentrations of more than 0.05% was able to completely destroy viral infectivity, as assessed on susceptible target cells. We have further shown that HIV-1, which is present in both seminal and genital secretions, can be inactivated in such fluids by direct exposure to benzalkonium chloride.