Quantification of global transcription patterns in prokaryotes using spotted microarrays

We describe an analysis, applicable to any spotted microarray dataset produced using genomic DNA as a reference, that quantifies prokaryotic levels of mRNA on a genome-wide scale. Applying this to Mycobacterium tuberculosis, we validate the technique, show a correlation between level of expression and biological importance, define the complement of invariant genes and analyze absolute levels of expression by functional class to develop ways of understanding an organism's biology without comparison to another growth condition.     

Inositol monophosphate phosphatase genes of <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis>

Background  

Mycobacteria use inositol in phosphatidylinositol, for anchoring lipoarabinomannan (LAM), lipomannan (LM) and phosphatidylinosotol mannosides (PIMs) in the cell envelope, and for the production of mycothiol, which maintains the redox balance of the cell. Inositol is synthesized by conversion of glucose-6-phosphate to inositol-1-phosphate, followed by dephosphorylation by inositol monophosphate phosphatases (IMPases) to form myo-inositol. To gain insight into how Mycobacterium tuberculosis synthesises inositol we carried out genetic analysis of the four IMPase homologues that are present in the Mycobacterium tuberculosis genome.     

Evaluation of The Effect of Letrozole in the Ovarian Hyperstimulation Syndrome Prevention in Participants at Risk of Treatment with Ovulation-Stimulating Drugs:A Randomized Controlled Trial

Background:This study aims to evaluate the effect of Letrozole (LE) in reducing ovarian hyperstimulation syndrome (OHSS) in high-risk participants with polycystic ovary syndrome (PCOS) treated with In vitro fertilization (IVF).Methods:This study was a randomized clinical trial in which participants were randomly divided into two groups (n= 25 per group). Based on GnRH-antagonist protocol, recombinant follicle-stimulating hormone 150 units/day subcutaneously and human menopausal gonadotropin 75 units/ day intramuscularly used from day 2 of the menstrual cycle. In the study group, Letrozole 5 mg daily was added simultaneously with gonadotropin during the first five days of the IVF cycle and in the control group placebo was added.Results:There were statistically significant differences among the groups in terms of Estradiol level on Trigger Day (p= 0.04). The total days of stimulation and cumulative Gonadotropin dose were significantly lower in the Letrozole group (p= 0.00). There were no significant differences between the groups in terms of the number of oocytes retrieved, numbers of implanted embryos, and clinical pregnancy rates (p-value> 0.05). There was only one moderate case in the intervention group and 9 moderate symptoms in the control group (p= 0.04).DiscussionAdministration of Letrozole with GnRH antagonist protocol, conventional protocol in PCOS cases in IVF cycle, had a significant effect on reducing the incidence of OHSS. So, if the future studies prove LE co-administration may lessen the incidence of OHSS, LE will be a highly potent drug for preventing OHSS in PCOS cases.Key Words: In vitro fertilization, Infertility, Letrozole, Ovarian hyperstimulation syndrome     

Nucleoside-catabolizing Enzymes in Mycoplasma-infected Tumor Cell Cultures Compromise the Cytostatic Activity of the Anticancer Drug Gemcitabine

The intracellular metabolism and cytostatic activity of the anticancer drug gemcitabine (2′,2′-difluoro-2′-deoxycytidine; dFdC) was severely compromised in Mycoplasma hyorhinis-infected tumor cell cultures. Pronounced deamination of dFdC to its less cytostatic metabolite 2′,2′-difluoro-2′-deoxyuridine was observed, both in cell extracts and spent culture medium (i.e. tumor cell-free but mycoplasma-containing) of mycoplasma-infected tumor cells. This indicates that the decreased antiproliferative activity of dFdC in such cells is attributed to a mycoplasma cytidine deaminase causing rapid drug catabolism. Indeed, the cytostatic activity of gemcitabine could be restored by the co-administration of tetrahydrouridine (a potent cytidine deaminase inhibitor). Additionally, mycoplasma-derived pyrimidine nucleoside phosphorylase (PyNP) activity indirectly potentiated deamination of dFdC: the natural pyrimidine nucleosides uridine, 2′-deoxyuridine and thymidine inhibited mycoplasma-associated dFdC deamination but were efficiently catabolized (removed) by mycoplasma PyNP. The markedly lower anabolism and related cytostatic activity of dFdC in mycoplasma-infected tumor cells was therefore also (partially) restored by a specific TP/PyNP inhibitor (TPI), or by exogenous thymidine. Consequently, no effect on the cytostatic activity of dFdC was observed in tumor cell cultures infected with a PyNP-deficient Mycoplasma pneumoniae strain. Because it has been reported that some commensal mycoplasma species (including M. hyorhinis) preferentially colonize tumor tissue in cancer patients, our findings suggest that the presence of mycoplasmas in the tumor microenvironment could be a limiting factor for the anticancer efficiency of dFdC-based chemotherapy. Accordingly, a significantly decreased antitumor effect of dFdC was observed in mice bearing M. hyorhinis-infected murine mammary FM3A tumors compared with uninfected tumors.     

Localizing confined epileptic foci in patients with an unclear focus or presumed multifocality using a component-based EEG-fMRI method

Precise localization of epileptic foci is an unavoidable prerequisite in epilepsy surgery. Simultaneous EEG-fMRI recording has recently created new horizons to locate foci in patients with epilepsy and, in comparison with single-modality methods, has yielded more promising results although it is still subject to limitations such as lack of access to information between interictal events. This study assesses its potential added value in the presurgical evaluation of patients with complex source localization. Adult candidates considered ineligible for surgery on account of an unclear focus and/or presumed multifocality on the basis of EEG underwent EEG-fMRI. Adopting a component-based approach, this study attempts to identify the neural behavior of the epileptic generators and detect the components-of-interest which will later be used as input in the GLM model, substituting the classical linear regressor. Twenty-eight sets interictal epileptiform discharges (IED) from nine patients were analyzed. In eight patients, at least one BOLD response was significant, positive and topographically related to the IEDs. These patients were rejected for surgery because of an unclear focus in four, presumed multifocality in three, and a combination of the two conditions in two. Component-based EEG-fMRI improved localization in five out of six patients with unclear foci. In patients with presumed multifocality, component-based EEG-fMRI advocated one of the foci in five patients and confirmed multifocality in one of the patients. In seven patients, component-based EEG-fMRI opened new prospects for surgery and in two of these patients, intracranial EEG supported the EEG-fMRI results. In these complex cases, component-based EEG-fMRI either improved source localization or corroborated a negative decision regarding surgical candidacy. As supported by the statistical findings, the developed EEG-fMRI method leads to a more realistic estimation of localization compared to the conventional EEG-fMRI approach, making it a tool of high value in pre-surgical evaluation of patients with refractory epilepsy. To ensure proper implementation, we have included guidelines for the application of component-based EEG-fMRI in clinical practice.     

Investigating the function of the putative mycolic acid methyltransferase UmaA: divergence between the Mycobacterium smegmatis and Mycobacterium tuberculosis proteins

Mycolic acids are major and specific lipid components of the cell envelope of mycobacteria that include the causative agents of tuberculosis and leprosy, Mycobacterium tuberculosis and Mycobacterium leprae, respectively. Subtle structural variations that are known to be crucial for both their virulence and the permeability of their cell envelope occur in mycolic acids. Among these are the introduction of cyclopropyl groups and methyl branches by mycolic acid S-adenosylmethionine-dependent methyltransferases (MA-MTs). While the functions of seven of the M. tuberculosis MA-MTs have been either established or strongly presumed nothing is known of the roles of the remaining umaA gene product and those of M. smegmatis MA-MTs. Mutants of the M. tuberculosis umaA gene and its putative M. smegmatis orthologue, MSMEG0913, were created. The lipid extracts of the resulting mutants were analyzed in detail using a combination of analytical techniques such as matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and proton nuclear magnetic resonance spectroscopy, and chemical degradation methods. The M. smegmatis mutants no longer synthesized subtypes of mycolates containing a methyl branch adjacent to either trans cyclopropyl group or trans double bond at the "proximal" position of both alpha- and epoxy-mycolates. Complementation with MSMEG0913, but not with umaA, fully restored the wild-type phenotype in M. smegmatis. Consistently, no modification was observed in the structures of mycolic acids produced by the M. tuberculosis umaA mutant. These data proved that despite their synteny and high similarity umaA and MSMEG0913 are not functionally orthologous.     

Anatomical responses of leaves of Black Locust (<Emphasis Type="Italic">Robinia pseudoacacia</Emphasis> L.) to urban pollutant gases and climatic factors

This study investigates responses in the leaf anatomy of Black Locust (Robinia pseudoacacia L.) to the atmospheric pollutants, SO₂, NO₂ and O₃ and climate in Tehran. The anatomical variables studied include thickness of the leaf lamina and of its main constituent tissues and the length and density of stomata. We present evidence that, in response to urban air pollution, the spongy mesophyll layer is thinner, the upper cuticle of the leaf thicker and stomatal density and the ratio of palisade parenchyma to spongy parenchyma are increased. Similar responses were also detected in relation to a climatic gradient. Stomatal density and thickness of the leaf lamina and of its mesophyll layer were all higher under warmer drier conditions. This overlap in anatomical response to two very different suites of environmental variables may reflect a functional overlap between mechanisms designed to restrict water loss in dry climates and those that minimize the uptake of toxic gases in polluted habitats.     

K‐Lysine acetyltransferase 2a regulates a hippocampal gene expression network linked to memory formation

Neuronal histone acetylation has been linked to memory consolidation, and targeting histone acetylation has emerged as a promising therapeutic strategy for neuropsychiatric diseases. However, the role of histone‐modifying enzymes in the adult brain is still far from being understood. Here we use RNA sequencing to screen the levels of all known histone acetyltransferases (HATs) in the hippocampal CA1 region and find that K‐acetyltransferase 2a (Kat2a)—a HAT that has not been studied for its role in memory function so far—shows highest expression. Mice that lack Kat2a show impaired hippocampal synaptic plasticity and long‐term memory consolidation. We furthermore show that Kat2a regulates a highly interconnected hippocampal gene expression network linked to neuroactive receptor signaling via a mechanism that involves nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB). In conclusion, our data establish Kat2a as a novel and essential regulator of hippocampal memory consolidation.     

The Mycobacterium tuberculosis ino1 gene is essential for growth and virulence

Inositol is utilized by Mycobacterium tuberculosis in the production of its major thiol and of essential cell wall lipoglycans. We have constructed a mutant lacking the gene encoding inositol-1-phosphate synthase (ino1), which catalyses the first committed step in inositol synthesis. This mutant is only viable in the presence of extremely high levels of inositol. Mutant bacteria cultured in inositol-free medium for four weeks showed a reduction in levels of mycothiol, but phosphatidylinositol mannoside, lipomannan and lipoarabinomannan levels were not altered. The ino1 mutant was attenuated in resting macrophages and in SCID mice. We used site-directed mutagenesis to alter four putative active site residues; all four alterations resulted in a loss of activity, and we demonstrated that a D310N mutation caused loss of the active site Zn2+ ion and a conformational change in the NAD+ cofactor.