The Signature Amino Acid Residue Serine 31 of HIV-1C Tat Potentiates an Activated Phenotype in Endothelial Cells.

The natural cysteine to serine variation at position 31 of Tat in HIV-1C disrupts the dicysteine motif attenuating the chemokine function of Tat. We ask if there exists a trade-off in terms of a gain of function for HIV-1C Tat due to this natural variation. We constructed two Tat-expression vectors encoding Tat proteins discordant for the serine 31 residue (CS-Tat vs. CC-Tat), expressed the proteins in Jurkat cells under doxycycline control, and performed the whole transcriptome analysis to compare the early events of Tat-induced host gene expression. Our analysis delineated a significant enrichment of pathways and gene ontologies associated with the angiogenic signaling events in CS-Tat stable cells. Subsequently, we validated and compared angiogenic signaling events induced by CS- vs. CC-Tat using human umbilical vein endothelial cells (HUVEC) and the human cerebral microvascular endothelial cell line (hCMEC/D3). CS-Tat significantly enhanced the production of CCL2 from HUVEC and induced an activated phenotype in endothelial cells conferring on them enhanced migration, invasion, and in vitro morphogenesis potential. The ability of CS-Tat to induce the activated phenotype in endothelial cells could be of significance, especially in the context of HIV-associated cardiovascular and neuronal disorders. The findings from the present study are likely to help appreciate the functional significance of the SAR (signature amino acid residues) influencing the unique biological properties.

Genome-wide analysis of mammary gland shows modulation of transcriptome landscape with alternative splice variants in Staphylococcus aureus mastitis in mice.

Epidemiological mapping shows Staphylococcus aureus to be the leading mastitis causing pathogen in India with diverse genetic lineages circulating in the dairy cattle population. We previously reported that endemic clonal strains of S. aureus isolated from subclinical mastitis lead to specific alteration of epigenetic modulators resulting in deviating immune response in intramammary infection mouse model. However, the extent of transcriptome modulation and associated alternative splicing in S. aureus mastitis is poorly understood. Hence, to gain a deeper insight of the extent of modulation of transcriptome landscape, we expanded the study here using high throughput, paired-end RNA sequencing analysis of the mouse mammary gland inoculated with three strains of S. aureus (SA1, SA2, and SA3) possessing specific genotype, virulence and enterotoxin traits. Overall, we detected 35,878 transcripts in S. aureus inoculated mammary gland, 23% more than those annotated in the reference genome. Expression of 20,756 transcripts was > 1 fragment per kilobase of transcript per million mapped fragments and 25.95% of multi-exonic genes were alternatively spliced. We noted Alternative Splicing (AS) events for > 100 immune-related genes. S. aureus infection quantitatively altered AS events in mice mammary gland. Collectively, the majority of differentially expressed significant genes clustered into immune-associated, cell adhesion and metabolic process categories. We observed AS events for 379 transcripts of genes putatively encoding several splicing associated proteins and transcription factors besides inflammatory mediators. The present analysis provides new insights into global transcriptome landscape and AS events in host-defense related genes in response to S. aureus intramammary infection, suggesting the need for studies focusing on multi-target and/or network therapeutics approach to combat mastitis.

Publisher Correction: Co-expression network analysis of toxin-antitoxin loci in Mycobacterium tuberculosis reveals key modulators of cellular stress.

A correction to this article has been published and is linked from the HTML and PDF version of this paper. The error has been fixed in the paper.

Draft Genome Sequence of Rhizoctonia solani Anastomosis Group 1 Subgroup 1A Strain 1802/KB Isolated from Rice.

Sheath blight, caused by Rhizoctonia solani anastomosis group 1 subgroup 1A (AG1-1A), is one of the most devastating rice diseases worldwide. Here, we report the draft genome sequence of R. solani AG1-1A strain 1802/KB isolated from a popular Malaysian rice variety. To the best of our knowledge, this is the second reported representative genome from AG1-1A.

Co-expression network analysis of toxin-antitoxin loci in Mycobacterium tuberculosis reveals key modulators of cellular stress.

Research on toxin-antitoxin loci (TA loci) is gaining impetus due to their ubiquitous presence in bacterial genomes and their observed roles in stress survival, persistence and drug tolerance. The present study investigates the expression profile of all the seventy-nine TA loci found in Mycobacterium tuberculosis. The bacterium was subjected to multiple stress conditions to identify key players of cellular stress response and elucidate a TA-coexpression network. This study provides direct experimental evidence for transcriptional activation of each of the seventy-nine TA loci following mycobacterial exposure to growth-limiting environments clearly establishing TA loci as stress-responsive modules in M. tuberculosis. TA locus activation was found to be stress-specific with multiple loci activated in a duration-based response to a particular stress. Conditions resulting in arrest of cellular translation led to greater up-regulation of TA genes suggesting that TA loci have a primary role in arresting translation in the cell. Our study identifed higBA2 and vapBC46 as key loci that were activated in all the conditions tested. Besides, relBE1, higBA3, vapBC35, vapBC22 and higBA1 were also upregulated in multpile stresses. Certain TA modules exhibited co-activation across multiple conditions suggestive of a common regulatory mechanism.

Gene regulatory networking reveals the molecular cue to lysophosphatidic acid-induced metabolic adaptations in ovarian cancer cells.

Extravasation and metastatic progression are two main reasons for the high mortality rate associated with cancer. The metastatic potential of cancer cells depends on a plethora of metabolic challenges prevailing within the tumor microenvironment. To achieve higher rates of proliferation, cancer cells reprogram their metabolism, increasing glycolysis and biosynthetic activities. Just why this metabolic reprogramming predisposes cells towards increased oncogenesis remains elusive. The accumulation of myriad oncolipids in the tumor microenvironment has been shown to promote the invasiveness of cancer cells, with lysophosphatidic acid (LPA) being one such critical factor enriched in ovarian cancer patients. Cellular bioenergetic studies confirm that oxidative phosphorylation is suppressed and glycolysis is increased with long exposure to LPA in ovarian cancer cells compared with non-transformed epithelial cells. We sought to uncover the regulatory complexity underlying this oncolipid-induced metabolic perturbation. Gene regulatory networking using RNA-Seq analysis identified the oncogene ETS-1 as a critical mediator of LPA-induced metabolic alterations for the maintenance of invasive phenotype. Moreover, LPA receptor-2 specific PtdIns3K-AKT signaling induces ETS-1 and its target matrix metalloproteases. Abrogation of ETS-1 restores cellular bioenergetics towards increased oxidative phosphorylation and reduced glycolysis, and this effect was reversed by the presence of LPA. Furthermore, the bioenergetic status of LPA-treated ovarian cancer cells mimics hypoxia through induction of hypoxia-inducible factor-1α, which was found to transactivate ets-1. Studies in primary tumors generated in syngeneic mice corroborated the in vitro findings. Thus, our study highlights the phenotypic changes induced by the pro-metastatic factor ETS-1 in ovarian cancer cells. The relationship between enhanced invasiveness and metabolic plasticity further illustrates the critical role of metabolic adaptation of cancer cells as a driver of tumor progression. These findings reveal oncolipid-induced metabolic predisposition as a new mechanism of tumorigenesis and propose metabolic inhibitors as a potential approach for future management of aggressive ovarian cancer.

Streptococcus uberis ST439 and ST475 induce differential inflammatory responses in a mouse intramammary infection model.

Streptococcus uberis causing mastitis is a growing challenge to the dairy industry. Molecular, epidemiological and population structure studies have revealed clonal diversity among the infecting strains. In this study, mouse intramammary infection model was used to uncover the host immune response to two epidemiologically important live strains of S. uberis (SU1and SU2) obtained from subclinical case of mastitis possessing specific and unique multi locus sequence types (ST), pulsed field gel electrophoresis (PFGE) pulsotypes and virulence profiles. Temporal (2h, 4h, 8h, 12h, 24h and 48h) expression of key inflammatory mediators (IL2, IL4, IL6, IL12, TNFα, IFNγ, GMCSF, TLR2, TLR4, TLR9, TLR11, TLR12, CD14, IL1ß, RANTES, Lactoferrin, and CXCl1) by reverse transcription and probe-based quantitative real-time PCR showed relative mRNA levels higher (p<0.05) in response to SU2 compared with SU1 with 24h PI serving as a critical point for the deviating behavior (SU1 versus SU2). Further employing the predicted biological processes under the influence of this pool of tested genes, the delineation of gene regulatory networks suggested SU1-favoring its persistence in the host environment; in contrast, SU2-which elevated gene expression indicating towards pathogen clearance or immune surveillance. This study suggested how these unique strains could manipulate the host immune response to influence the severity of mastitis; our results expand the available information on host pathogen interaction and provide a firm foundation needing further investigations to gain control over this pathogen.

Global gene expression profiling data analysis reveals key gene families and biological processes inhibited by Mithramycin in sarcoma cell lines.

The role of Mithramycin as an anticancer drug has been well studied. Sarcoma is a type of cancer arising from cells of mesenchymal origin. Though incidence of sarcoma is not of significant percentage, it becomes vital to understand the role of Mithramycin in controlling tumor progression of sarcoma. In this article, we have analyzed the global gene expression profile changes induced by Mithramycin in two different sarcoma lines from whole genome gene expression profiling microarray data. We have found that the primary mode of action of Mithramycin is by global repression of key cellular processes and gene families like phosphoproteins, kinases, alternative splicing, regulation of transcription, DNA binding, regulation of histone acetylation, negative regulation of gene expression, chromosome organization or chromatin assembly and cytoskeleton.

Ameliorated de novo transcriptome assembly using Illumina paired end sequence data with Trinity Assembler.

Advent of Next Generation Sequencing has led to possibilities of de novo transcriptome assembly of organisms without availability of complete genome sequence. Among various sequencing platforms available, Illumina is the most widely used platform based on data quality, quantity and cost. Various de novo transcriptome assemblers are also available today for construction of de novo transcriptome. In this study, we aimed at obtaining an ameliorated de novo transcriptome assembly with sequence reads obtained from Illumina platform and assembled using Trinity Assembler. We found that, primary transcriptome assembly obtained as a result of Trinity can be ameliorated on the basis of transcript length, coverage, and depth and protein homology. Our approach to ameliorate is reproducible and could enhance the sensitivity and specificity of the assembled transcriptome which could be critical for validation of the assembled transcripts and for planning various downstream biological assays.

Global insights into high temperature and drought stress regulated genes by RNA-Seq in economically important oilseed crop Brassica juncea.

BACKGROUND: Brassica juncea var. Varuna is an economically important oilseed crop of family Brassicaceae which is vulnerable to abiotic stresses at specific stages in its life cycle. Till date no attempts have been made to elucidate genome-wide changes in its transcriptome against high temperature or drought stress. To gain global insights into genes, transcription factors and kinases regulated by these stresses and to explore information on coding transcripts that are associated with traits of agronomic importance, we utilized a combinatorial approach of next generation sequencing and de-novo assembly to discover B. juncea transcriptome associated with high temperature and drought stresses. RESULTS: We constructed and sequenced three transcriptome libraries namely Brassica control (BC), Brassica high temperature stress (BHS) and Brassica drought stress (BDS). More than 180 million purity filtered reads were generated which were processed through quality parameters and high quality reads were assembled de-novo using SOAPdenovo assembler. A total of 77750 unique transcripts were identified out of which 69,245 (89%) were annotated with high confidence. We established a subset of 19110 transcripts, which were differentially regulated by either high temperature and/or drought stress. Furthermore, 886 and 2834 transcripts that code for transcription factors and kinases, respectively, were also identified. Many of these were responsive to high temperature, drought or both stresses. Maximum number of up-regulated transcription factors in high temperature and drought stress belonged to heat shock factors (HSFs) and dehydration responsive element-binding (DREB) families, respectively. We also identified 239 metabolic pathways, which were perturbed during high temperature and drought treatments. Analysis of gene ontologies associated with differentially regulated genes forecasted their involvement in diverse biological processes. CONCLUSIONS: Our study provides first comprehensive discovery of B. juncea transcriptome under high temperature and drought stress conditions. Transcriptome resource generated in this study will enhance our understanding on the molecular mechanisms involved in defining the response of B. juncea against two important abiotic stresses. Furthermore this information would benefit designing of efficient crop improvement strategies for tolerance against conditions of high temperature regimes and water scarcity.