Mycobacterium tuberculosis oriC sequestration by MtrA response regulator.

The regulators of Mycobacterium tuberculosis DNA replication are largely unknown. Here, we demonstrate that in synchronously replicating M. tuberculosis, MtrA access to origin of replication (oriC) is enriched in the post-replication (D) period. The increased oriC binding results from elevated MtrA phosphorylation (MtrA∼P) as evidenced by reduced expression of dnaN, dnaA and increased expression of select cell division targets. Overproduction of gain-of-function MtrAY102C advanced the MtrA oriC access to the C period, reduced dnaA and dnaN expression, interfered with replication synchrony and compromised cell division. Overproduction of wild-type (MtrA+) or phosphorylation-defective MtrAD56N did not promote oriC access in the C period, nor affected cell cycle progression. MtrA interacts with DnaA signaling a possibility that DnaA helps load MtrA on oriC. Therefore, oriC sequestration by MtrA∼P in the D period may normally serve to prevent untimely initiations and that DnaA-MtrA interactions may facilitate regulated oriC replication. Finally, despite the near sequence identity of MtrA in M. smegmatis and M. tuberculosis, the M. smegmatis oriC is not MtrA-target. We conclude that M. tuberculosis oriC has evolved to be regulated by MtrA and that cell cycle progression in this organisms are governed, at least in part, by oscillations in the MtrA∼P levels.

Mycobacterium tuberculosis MtrB sensor kinase interactions with FtsI and Wag31 proteins reveal a role for MtrB distinct from that regulating MtrA activities.

The septal association of Mycobacterium tuberculosis MtrB, the kinase partner of the MtrAB two-component signal transduction system, is necessary for the optimal expression of the MtrA regulon targets, including ripA, fbpB, and ftsI, which are involved in cell division and cell wall synthesis. Here, we show that MtrB, irrespective of its phosphorylation status, interacts with Wag31, whereas only phosphorylation-competent MtrB interacts with FtsI. We provide evidence that FtsI depletion compromises the MtrB septal assembly and MtrA regulon expression; likewise, the absence of MtrB compromises FtsI localization and, possibly, FtsI activity. We conclude from these results that FtsI and MtrB are codependent for their activities and that FtsI functions as a positive modulator of MtrB activation and MtrA regulon expression. In contrast to FtsI, Wag31 depletion does not affect MtrB septal assembly and MtrA regulon expression, whereas the loss of MtrB increased Wag31 localization and the levels of PknA/PknB (PknA/B) serine-threonine protein kinase-mediated Wag31 phosphorylation. Interestingly, we found that FtsI decreased levels of phosphorylated Wag31 (Wag31∼P) and that MtrB interacted with PknA/B. Overall, our results indicate that MtrB interactions with FtsI, Wag31, and PknA/B are required for its optimal localization, MtrA regulon expression, and phosphorylation of Wag31. Our results emphasize a new role for MtrB in cell division and cell wall synthesis distinct from that regulating the MtrA phosphorylation activities.

Septal localization of the Mycobacterium tuberculosis MtrB sensor kinase promotes MtrA regulon expression.

The mechanisms responsible for activation of the MtrAB two-component regulatory signal transduction system, which includes sensor kinase MtrB and response regulator MtrA, are unknown. Here, we show that an MtrB-GFP fusion protein localized to the cell membrane, the septa, and the poles in Mycobacterium tuberculosis and Mycobacterium smegmatis. This localization was independent of MtrB phosphorylation status but dependent upon the assembly of FtsZ, the initiator of cell division. The M. smegmatis mtrB mutant was filamentous, defective for cell division, and contained lysozyme-sensitive cell walls. The mtrB phenotype was complemented by either production of MtrB protein competent for phosphorylation or overproduction of MtrA(Y102C) and MtrA(D13A) mutant proteins exhibiting altered phosphorylation potential, indicating that either MtrB phosphorylation or MtrB independent expression of MtrA regulon genes, including those involved in cell wall processing, are necessary for regulated cell division. In partial support of this observation, we found that the essential cell wall hydrolase ripA is an MtrA target and that the expression of bona fide MtrA targets ripA, fbpB, and dnaA were compromised in the mtrB mutant and partially rescued upon MtrA(Y102C) and MtrA(D13A) overproduction. MtrB septal assembly was compromised upon FtsZ depletion and exposure of cells to mitomycin C, a DNA damaging agent, which interferes with FtsZ ring assembly. Expression of MtrA targets was also compromised under the above conditions, indicating that MtrB septal localization and MtrA regulon expression are linked. We propose that MtrB septal association is a necessary feature of MtrB activation that promotes MtrA phosphorylation and MtrA regulon expression.

Mycobacterium tuberculosis origin of replication and the promoter for immunodominant secreted antigen 85B are the targets of MtrA, the essential response regulator.

Efficient proliferation of Mycobacterium tuberculosis (Mtb) inside macrophage requires that the essential response regulator MtrA be optimally phosphorylated. However, the genomic targets of MtrA have not been identified. We show by chromatin immunoprecipitation and DNase I footprinting that the chromosomal origin of replication, oriC, and the promoter for the major secreted immunodominant antigen Ag85B, encoded by fbpB, are MtrA targets. DNase I footprinting analysis revealed that MtrA recognizes two direct repeats of GTCACAgcg-like sequences and that MtrA approximately P, the phosphorylated form of MtrA, binds preferentially to these targets. The oriC contains several MtrA motifs, and replacement of all motifs or of a single select motif with TATATA compromises the ability of oriC plasmids to maintain stable autonomous replication in wild type and MtrA-overproducing strains, indicating that the integrity of the MtrA motif is necessary for oriC replication. The expression of the fbpB gene is found to be down-regulated in Mtb cells upon infection when these cells overproduce wild type MtrA but not when they overproduce a nonphosphorylated MtrA, indicating that MtrA approximately P regulates fbpB expression. We propose that MtrA is a regulator of oriC replication and that the ability of MtrA to affect apparently unrelated targets, i.e. oriC and fbpB, reflects its main role as a coordinator between the proliferative and pathogenic functions of Mtb.

The two-domain LysX protein of Mycobacterium tuberculosis is required for production of lysinylated phosphatidylglycerol and resistance to cationic antimicrobial peptides.

The well-recognized phospholipids (PLs) of Mycobacterium tuberculosis (Mtb) include several acidic species such as phosphatidylglycerol (PG), cardiolipin, phosphatidylinositol and its mannoside derivatives, in addition to a single basic species, phosphatidylethanolamine. Here we demonstrate that an additional basic PL, lysinylated PG (L-PG), is a component of the PLs of Mtb H37Rv and that the lysX gene encoding the two-domain lysyl-transferase (mprF)-lysyl-tRNA synthetase (lysU) protein is responsible for L-PG production. The Mtb lysX mutant is sensitive to cationic antibiotics and peptides, shows increased association with lysosome-associated membrane protein-positive vesicles, and it exhibits altered membrane potential compared to wild type. A lysX complementing strain expressing the intact lysX gene, but not one expressing mprF alone, restored the production of L-PG and rescued the lysX mutant phenotypes, indicating that the expression of both proteins is required for LysX function. The lysX mutant also showed defective growth in mouse and guinea pig lungs and showed reduced pathology relative to wild type, indicating that LysX activity is required for full virulence. Together, our results suggest that LysX-mediated production of L-PG is necessary for the maintenance of optimal membrane integrity and for survival of the pathogen upon infection.

Mycobacterium tuberculosis mtrA merodiploid strains with point mutations in the signal-receiving domain of MtrA exhibit growth defects in nutrient broth.

The genetic and biochemical aspects of the essential Mycobacteriumtuberculosis MtrAB two-component regulatory signal transduction (2CRS) system have not been extensively investigated. We show by bacterial two-hybrid assay that the response regulator (RR) MtrA and the sensor kinase MtrB interact. We further demonstrate that divalent metal ions [Mg²+, Ca²+ or both] promote MtrB kinase autophosphorylation activity, but only Mg²+ promotes phosphotransfer to MtrA. Replacement of the conserved aspartic acid residues at positions 13 and 56 with alanine (D13A), glutamine (D56E) or asparagine (D56N) prevented MtrA phosphorylation, indicating that these residues are important for phosphorylation. The MtrA(D56E) and MtrA(D13A) proteins bound to the promoter of fbpB, the gene encoding antigen 85B protein, efficiently in the absence of phosphorylation, whereas MtrA(D56N) did not. We also show that M.tuberculosismtrA merodiploids overproducing MtrA(D13A), unlike cells overproducing wild-type MtrA, grow poorly in nutrient broth and show reduced expression of fbpB. These latter findings are reminiscent of a phenotype associated with MtrA overproduction during intramacrophage growth. Our results suggest that MtrA(D13A) behaves like a constitutively active response regulator and that further characterization of mtrA merodiploid strains will provide valuable clues to the MtrAB system.

Ontario's COVID-19 Modelling Consensus Table: mobilizing scientific expertise to support pandemic response.

SETTING: COVID-19 has highlighted the need for credible epidemiological models to inform pandemic policy. Traditional mechanisms of commissioning research are ill-suited to guide policy during a rapidly evolving pandemic. At the same time, contracting with a single centre of expertise has been criticized for failing to reflect challenges inherent in specific modelling approaches. INTERVENTION: This report describes an alternative approach to mobilizing scientific expertise. Ontario's COVID-19 Modelling Consensus Table (MCT) was created in March 2020 to enable rapid communication of credible estimates of the impact of COVID-19 and to accelerate learning on how the disease is spreading and what could slow its transmission. The MCT is a partnership between the province and academic modellers and consists of multiple groups of experts, health system leaders, and senior decision-makers. Armed with Ministry of Health data, the MCT meets once per week to share results from modelling exercises, generate consensus judgements of the likely future impact of COVID-19, and discuss decision-makers' priorities. OUTCOMES: The MCT has enabled swift access to data for participants, a structure for developing consensus estimates and communicating these to decision-makers, credible models to inform health system planning, and increased transparency in public reporting of COVID-19 data. It has also facilitated the rapid publication of research findings and its incorporation into government policy. IMPLICATIONS: The MCT approach is one way to quickly draw on scientific advice outside of government and public health agencies. Beyond speed, this approach allows for nimbleness as experts from different organizations can be added as needed. It also shows how universities and research institutes have a role to play in crisis situations, and how this expertise can be marshalled to inform policy while respecting academic freedom and confidentiality.

RéSUMé: LIEU: La COVID-19 a mis en évidence le besoin de modèles épidémiologiques crédibles pour éclairer la politique pandémique. Les mécanismes habituels pour commander des travaux de recherche sont peu propices à orienter les politiques lors d'une pandémie qui évolue rapidement. En même temps, la passation de contrats avec un seul centre d'expertise est critiquée, car elle ne tient pas compte des difficultés inhérentes de certaines approches de modélisation. INTERVENTION: Le présent rapport décrit une approche de rechange pour mobiliser le savoir scientifique. L'Ontario a créé en mars 2020 une Table de concertation sur la modélisation (TCM) qui permet de communiquer de façon rapide et fiable les estimations des effets de la COVID-19 et d'apprendre plus vite comment la maladie se propage et ce qui pourrait en ralentir la transmission. La TCM, un partenariat entre les modélisateurs de la province et des milieux universitaires, est composée de plusieurs groupes d'experts, de dirigeants du système de santé et de décideurs de haut niveau. Armée des données du ministère de la Santé, la TCM se réunit une fois par semaine pour partager les résultats d'exercices de modélisation, générer des jugements consensuels sur les futurs effets probables de la COVID-19 et discuter des priorités des décideurs. RéSULTATS: La TCM rend possible un accès rapide aux données pour les participants, une structure pour élaborer des estimations consensuelles et les communiquer aux décideurs, des modèles fiables pour éclairer la planification du système de santé, ainsi qu'une transparence accrue dans la communication des données sur la COVID-19 au public. Elle facilite aussi la publication rapide des résultats de recherche et leur intégration dans la politique gouvernementale. CONSéQUENCES: L'approche de la TCM est un moyen d'obtenir rapidement des conseils scientifiques à l'extérieur du gouvernement et des organismes de santé publique. Au-delà de sa rapidité, cette approche offre une grande souplesse, car des experts de différents organismes peuvent être ajoutés au besoin. Elle montre aussi que les universités et les établissements de recherche ont un rôle à jouer dans les situations de crise, et qu'il est possible de mobiliser leurs compétences pour éclairer les politiques tout en respectant la liberté et la confidentialité des milieux de la recherche et de l'enseignement.

Extracellular matrix-modulated expression of human cell surface glycoproteins A42 and J143. Intrinsic and extrinsic signals determine antigenic phenotype.

We have used serologic, biochemical, and genetic methods to characterize two stage-specific human differentiation antigens of neural and melanocytic cells: A42 (57,000 Mr glycoprotein) and J143 (140,000/30,000 Mr glycoprotein). The genes determining A42 and J143 cell surface expression in rodent-human hybrids were chromosomally mapped, and the respective human chromosomes were introduced into rodent cells derived from distinct differentiation lineages. Serologic analysis of the resulting hybrid clones has permitted the identification of two types of regulatory signals determining A42 and J143 expression. First, both antigens are expressed in hybrids constructed with antigen-positive human cells and also in certain hybrids constructed with antigen-negative human cells, indicating that intrinsic signals provided by the differentiation program of the rodent fusion partner induce antigen expression. Second, a series of human-mouse neuroblastoma hybrids, which are A42- or J143- when cultured on plastic surfaces, can be induced to express the antigens when cultured on substrates coated with extracellular matrix (ECM) produced by bovine corneal endothelial cells or fibronectin. This induction of antigen expression by extrinsic, ECM-derived signals is accompanied in the neuroblastoma hybrids by increased substrate adhesiveness and cell spreading and by characteristic changes in cell morphology. A similar program of phenotypic changes is also seen in spontaneous variants of human neuroblastoma and Ewing's sarcoma cells and in ECM-induced Ewing's sarcoma cells. These findings suggest that ECM-derived signals have a role analogous to mitogens and soluble differentiation factors in modulating differentiation phenotypes and tissue-specific patterns of cell surface antigen expression.

Synchronous replication initiation in novel Mycobacterium tuberculosis dnaA cold-sensitive mutants.

The genetic aspects of oriC replication initiation in Mycobacterium tuberculosis are largely unknown. A two-step genetic screen was utilized for isolating M. tuberculosis dnaA cold-sensitive (cos) mutants. First, a resident plasmid expressing functional dnaA integrated at the attB locus in dnaA null background was exchanged with an incoming plasmid bearing a mutagenized dnaA gene. Next, the mutants that were defective for growth at 30 degrees C, a non-permissive temperature, but resumed growth and DNA synthesis when shifted to 37 degrees C, a permissive temperature, were subsequently selected. Nucleotide sequencing analysis located mutations to different regions of the dnaA gene. Modulation of the growth temperatures led to synchronized DNA synthesis. The dnaA expression under synchronized DNA replication conditions continued to increase during the replication period, but decreased thereafter reflecting autoregulation. The dnaAcos mutants at 30 degrees C were elongated suggesting that they may possibly be blocked during the cell division. The DnaA115 protein is defective in its ability to interact with ATP at 30 degrees C, but not at 37 degrees C. Our results suggest that the optimal cell cycle progression and replication initiation in M. tuberculosis requires that the dnaA promoter remains active during the replication period and that the DnaA protein is able to interact with ATP.

Synthesis and biological evaluation of piperazinyl heterocyclic antagonists of the gonadotropin releasing hormone (GnRH) receptor.

Antagonism of the gonadotropin releasing hormone (GnRH) receptor has resulted in positive clinical results in reproductive tissue disorders such as endometriosis and prostate cancer. Following the recent discovery of orally active GnRH antagonists based on a 4-piperazinylbenzimidazole template, we sought to investigate the properties of heterocyclic isosteres of the benzimidazole template. We report here the synthesis and biological activity of eight novel scaffolds, including imidazopyridines, benzothiazoles and benzoxazoles. The 2-(4-tert-butylphenyl)-8-(piperazin-1-yl)imidazo[1,2-a]pyridine ring system was shown to have nanomolar binding potency at the human and rat GnRH receptors as well as functional antagonism in vitro. Additional structure-activity relationships within this series are reported along with a pharmacokinetic comparison to the benzimidazole-based lead molecule.

Quantifying the shift in social contact patterns in response to non-pharmaceutical interventions.

Social contact mixing plays a critical role in influencing the transmission routes of infectious diseases. Moreover, quantifying social contact mixing patterns and their variations in a rapidly evolving pandemic intervened by changing public health measures is key for retroactive evaluation and proactive assessment of the effectiveness of different age- and setting-specific interventions. Contact mixing patterns have been used to inform COVID-19 pandemic public health decision-making; but a rigorously justified methodology to identify setting-specific contact mixing patterns and their variations in a rapidly developing pandemic, which can be informed by readily available data, is in great demand and has not yet been established. Here we fill in this critical gap by developing and utilizing a novel methodology, integrating social contact patterns derived from empirical data with a disease transmission model, that enables the usage of age-stratified incidence data to infer age-specific susceptibility, daily contact mixing patterns in workplace, household, school and community settings; and transmission acquired in these settings under different physical distancing measures. We demonstrated the utility of this methodology by performing an analysis of the COVID-19 epidemic in Ontario, Canada. We quantified the age- and setting (household, workplace, community, and school)-specific mixing patterns and their evolution during the escalation of public health interventions in Ontario, Canada. We estimated a reduction in the average individual contact rate from 12.27 to 6.58 contacts per day, with an increase in household contacts, following the implementation of control measures. We also estimated increasing trends by age in both the susceptibility to infection by SARS-CoV-2 and the proportion of symptomatic individuals diagnosed. Inferring the age- and setting-specific social contact mixing and key age-stratified epidemiological parameters, in the presence of evolving control measures, is critical to inform decision- and policy-making for the current COVID-19 pandemic.

Small molecule antagonists of the gonadotropin-releasing hormone (GnRH) receptor: structure-activity relationships of small heterocyclic groups appended to the 2-phenyl-4-piperazinyl-benzimidazole template.

A previous report described the serum LH suppression pharmacology of the 2-phenyl-4-piperazinyl-benzimidazole N-ethyluracil GnRH receptor antagonist 1 following oral administration in rats. A series of small heterocycles were appended to the 2-(4-tert-butylphenyl)-4-piperazinyl-benzimidazole template in place of the N-ethyluracil. Two imidazole analogues, 32 and 41, were shown to possess substantial in vitro potency at the target receptor (hGnRH IC(50) = 7 and 18 nM, respectively) and aqueous solubility (55 and 100 microg/mL at pH 7.4, respectively). Both compounds had high oral bioavailability in rats and 32 was further examined in an orchidectomized rat model for serum LH suppression based on increased volume of distribution over 41. Serum LH levels trended lower in orchidectomized rats following oral administration of 32.

Facilitation of dissociation reaction of nucleotides bound to Mycobacterium tuberculosis DnaA.

Acidic phospholipids have been shown to promote dissociation of bound nucleotides from Mycobacterium tuberculosis DnaA (DnaA(TB)) purified under denaturing conditions [Yamamoto et al., (2002) Modulation of Mycobacterium tuberculosis DnaA protein-adenine-nucleotide interactions by acidic phospholipids. Biochem. J., 363, 305-311]. In the present study, we show that a majority of DnaA(TB) in non-overproducing cells of M. tuberculosis is membrane associated. Estimation of phospholipid phosphorus following chloroform: methanol extraction of soluble DnaA(TB) purified under native conditions (nDnaA(TB)) confirmed the association with phospholipids. nDnaA(TB) exhibited weak ATPase activity, and rapidly exchanged ATP for bound ADP in the absence of any added phospholipids. We suggest that the outcome of intra-cellular DnaA(TB)-nucleotide interactions, hence DnaA(TB) activity, is influenced by phospholipids.

Amelioration of arsenic-induced oxidative stress in CHO cells by Ixora coccinea flower extract.

Chronic exposure to inorganic arsenic creates various health problems. Ixora coccinea flower extract was investigated for its ability to protect against arsenic-induced cytotoxicity and genotoxicity in CHO cell line. MTT assay confirmed the efficacy of the extract in ameliorating arsenic-induced cytotoxicity. The value (48 mM) of 24 h inhibitory concentration (IC50) of sodium arsenate for CHO cells was obtained by MTT assay. Various free radical scavenging assays like DPPH, ABTS and nitric oxide scavenging assay confirmed antioxidant activity of the Ixora coccinea flower extract. Pretreatment of the extract significantly inhibited the arsenic-induced DNA damage (p < 0.01) in CHO cells. The extract administration significantly (p < 0.01) inhibited the intracellular ROS and depolarization of mitochondrial membrane induced by sodium arsenate. Ixora coccinea flower extract reduced oxidative stress in cells. Antioxidant enzymes like catalase and SOD activity was restored significantly (p < 0.01) in pretreated CHO cells. Ixora coccinea flower extract also exhibited the anti-apoptotic potential by decreasing the percentage apoptotic index (p < 0.01). These results may expand the applications of Ixora coccinea flowers as an alternative food with antioxidant properties and protective functions against arsenic (iAs) induced toxicological effects.

MtrA Response Regulator Controls Cell Division and Cell Wall Metabolism and Affects Susceptibility of Mycobacteria to the First Line Antituberculosis Drugs.

The biological processes regulated by the essential response regulator MtrA and the growth conditions promoting its activation in Mycobacterium tuberculosis, a slow grower and pathogen, are largely unknown. Here, using a gain-of-function mutant, MtrAY 102C, which functions in the absence of the cognate MtrB sensor kinase, we show that the MtrA regulon includes several genes involved in the processes of cell division and cell wall metabolism. The expression of selected MtrA targets and intracellular MtrA levels were compromised under replication arrest induced by genetic manipulation and under stress conditions caused by toxic radicals. The loss of the mtrA gene in M. smegmatis, a rapid grower and non-pathogen, produced filamentous cells with branches and bulges, indicating defects in cell division and cell shape. The ΔmtrA mutant was sensitized to rifampicin and vancomycin and became more resistant to isoniazid, the first line antituberculosis drug. Our data are consistent with the proposal that MtrA controls the optimal cell division, cell wall integrity, and susceptibility to some antimycobacterial drugs.

The spectrum of myelodysplastic syndromes post-solid organ transplantation: a single institutional experience.

An increased incidence of acute myeloid leukemia (AML) has recently been documented in patients post-solid organ transplantation but the incidence and types of myelodysplastic syndromes (MDS) occurring in this patient population are not known. We identified 5 patients (3M, 2F, age 48-64 years) who developed MDS ranging from 1.8 to 25 years (median 4.2 years) post-solid organ transplantation, only 2 patients had received azathioprine. The cumulative incidence of MDS in heart and lung transplant recipients at 15 years was 0.5% and 1.8%, respectively, which is markedly higher compared to the general population. Low-risk types of MDS predominated, 3 of 5 patients are alive (median 3.9 years) since diagnosis. Deletions of chromosome 20q, which have not been previously reported in post-transplant MDS/AML, were identified in 3 cases. Our findings expand the morphologic and cytogenetic spectrum of MDS occurring post-solid organ transplantation and suggest that mechanisms beside azathioprine toxicity might be important in disease pathogenesis.

CUTLL1, a novel human T-cell lymphoma cell line with t(7;9) rearrangement, aberrant NOTCH1 activation and high sensitivity to gamma-secretase inhibitors.

Activating mutations in NOTCH1 are present in over 50% of human T-cell lymphoblastic leukemia (T-ALL) samples and inhibition of NOTCH1 signaling with gamma-secretase inhibitors (GSI) has emerged as a potential therapeutic strategy for the treatment of this disease. Here, we report a new human T-cell lymphoma line CUTLL1, which expresses high levels of activated NOTCH1 and is extremely sensitive to gamma-secretase inhibitors treatment. CUTLL1 cells harbor a t(7;9)(q34;q34) translocation which induces the expression of a TCRB-NOTCH1 fusion transcript encoding a membrane-bound truncated form of the NOTCH1 receptor. GSI treatment of CUTLL1 cells blocked NOTCH1 processing and caused rapid clearance of activated intracellular NOTCH1. Loss of NOTCH1 activity induced a gene expression signature characterized by the downregulation of NOTCH1 target genes such as HES1 and NOTCH3. In contrast with most human T-ALL cell lines with activating mutations in NOTCH1, CUTLL1 cells showed a robust cellular phenotype upon GSI treatment characterized by G1 cell cycle arrest and increased apoptosis. These results show that the CUTLL1 cell line has a strong dependence on NOTCH1 signaling for proliferation and survival and supports that T-ALL patients whose tumors harbor t(7;9) should be included in clinical trials testing the therapeutic efficacy NOTCH1 inhibition with GSIs.

Gene expression profiling and its practice in drug development.

The availability of sequenced genomes of human and many experimental animals necessitated the development of new technologies and powerful computational tools that are capable of exploiting these genomic data and ask intriguing questions about complex nature of biological processes. This gave impetus for developing whole genome approaches that can produce functional information of genes in the form of expression profiles and unscramble the relationships between variation in gene expression and the resulting physiological outcome. These profiles represent genetic fingerprints or catalogue of genes that characterize the cell or tissue being studied and provide a basis from which to begin an investigation of the underlying biology. Among the most powerful and versatile tools are high-density DNA microarrays to analyze the expression patterns of large numbers of genes across different tissues or within the same tissue under a variety of experimental conditions or even between species. The wide spread use of microarray technologies is generating large sets of data that is stimulating the development of better analytical tools so that functions can be predicted for novel genes. In this review, the authors discuss how these profiles are being used at various stages of the drug discovery process and help in the identification of new drug targets, predict the function of novel genes, and understand individual variability in response to drugs.

An Intrathermocline Eddy and a tropical cyclone in the Bay of Bengal.

The Bay of Bengal, subjected to monsoonal forcing and tropical cyclones, displays a complex field of ocean eddies. On 5 December 2013 a sub-surface vortex or Intrathermocline Eddy (ITE) composed of water characteristic of the Andaman Sea was observed within the thermocline of the western Bay of Bengal. We propose that the ITE was the product of Tropical Cyclone Lehar interaction on 27 November 2013 with a westward propagating surface eddy from the eastern Bay of Bengal. While Lehar's interaction with the ocean initially removes heat from the upper layers of the eddy, air-sea flux is limited as the deeper portions of the eddy was subducted into the stratified thermocline, inhibiting further interaction with the atmosphere. The ITE core from 30 to 150 m is thus isolated from local air-sea fluxes by strong stratification at the mixed layer base, and its periphery is stable to shear instability, suggestive of longevity and the ability to carry water far distances with minimal modification.

Loss of PRDM1/BLIMP-1 function contributes to poor prognosis of activated B-cell-like diffuse large B-cell lymphoma.

PRDM1/BLIMP-1, a master regulator of plasma-cell differentiation, is frequently inactivated in activated B-cell-like (ABC) diffuse large B-cell lymphoma (DLBCL) patients. Little is known about its genetic aberrations and relevant clinical implications. A large series of patients with de novo DLBCL was effectively evaluated for PRDM1/BLIMP-1 deletion, mutation, and protein expression. BLIMP-1 expression was frequently associated with the ABC phenotype and plasmablastic morphologic subtype of DLBCL, yet 63% of the ABC-DLBCL patients were negative for BLIMP-1 protein expression. In these patients, loss of BLIMP-1 was associated with Myc overexpression and decreased expression of p53 pathway molecules. In addition, homozygous PRDM1 deletions and PRDM1 mutations within exons 1 and 2, which encode for domains crucial for transcriptional repression, were found to show a poor prognostic impact in patients with ABC-DLBCL but not in those with germinal center B-cell-like DLBCL (GCB-DLBCL). Gene expression profiling revealed that loss of PRDM1/BLIMP-1 expression correlated with a decreased plasma-cell differentiation signature and upregulation of genes involved in B-cell receptor signaling and tumor-cell proliferation. In conclusion, these results provide novel clinical and biological insight into the tumor-suppressive role of PRDM1/BLIMP-1 in ABC-DLBCL patients and suggest that loss of PRDM1/BLIMP-1 function contributes to the overall poor prognosis of ABC-DLBCL patients.