Competitive inhibition of natural antisense Sok-RNA interactions activates Hok-mediated cell killing in Escherichia coli

Short regulatory RNAs are widespread in bacteria, and many function through antisense recognition of mRNA. Among the best studied antisense transcripts are RNA antitoxins that repress toxin mRNA translation. The hok/sok locus of plasmid R1 from Escherichia coli is an established model for RNA antitoxin action. Base-pairing between hok mRNA and Sok-antisense-RNA increases plasmid maintenance through post-segregational-killing of plasmid-free progeny cells. To test the model and the idea that sequestration of Sok-RNA activity could provide a novel antimicrobial strategy, we designed anti Sok peptide nucleic acid (PNA) oligomers that, according to the model, would act as competitive inhibitors of hok mRNA::Sok-RNA interactions. In hok/sok-carrying cells, anti Sok PNAs were more bactericidal than rifampicin. Also, anti Sok PNAs induced ghost cell morphology and an accumulation of mature hok mRNA, consistent with cell killing through synthesis of Hok protein. The results support the sense/antisense model for hok mRNA repression by Sok-RNA and demonstrate that antisense agents can be used to out-compete RNA::RNA interactions in bacteria. Finally, BLAST analyses of ≈200 prokaryotic genomes revealed that many enteric bacteria have multiple hok/sok homologous and analogous RNA-regulated toxin–antitoxin loci. Therefore, it is possible to activate suicide in bacteria by targeting antitoxins

A protective role for nitric oxide and salicylic acid for arsenite phytotoxicity in rice (Oryza sativa L.)

The authors are thankful to Director, CSIR-National Botanical Research Institute (CSIR-NBRI), Lucknow for the facilities and for the financial support from the network projects (CSIR-INDEPTH), New Delhi, India. APS is thankful to CSIR New Delhi, India respectively, for the award of Research Associateship. RDT is gratefully thankful to Award of Emeritus Scientist (CSIR). GD is thankful to SERB-DST, New Delhi for award of NPDF. AK is thankful to UGC for award of DSKPDF. Award of Fast Track Scientist to SM from DST is gratefully acknowledged. We are also thankful to Mr. Dilip Chakraborty for technical assistance.Peer reviewedPostprin

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Unorthodox players in estrogen signaling: A G-protein coupled receptor and microRNAs

The steroid hormone 17β-estradiol (E2) regulates a number of developmental and physiological processes. The classical understanding of the estrogen signaling implies that the two nuclear receptors (ER), ERα and ERβ, mediate all the physiological responses of E2. ERs are transcription factors, which regulate the expression of their targets upon binding to E2. Estrogens also mediate a lot of rapid outcomes within a matter of seconds to minutes that cannot be explained by the classical understanding of estrogen signaling, which involves transcription of target genes mediated by the ERs. Whereas, many of the rapid effects of estrogen are attributed to the cytoplasmic pools of ERs, it has become clear in the recent years that there are other factors contributing to the rapid effects of estrogen and one of the emerging such factors is a G-protein coupled receptor, GPR30. MicroRNAs (miRNAs) are a new class of small non-coding RNAs, 20 to 25 nucleotides in length that regulate the expression of other genes. Upon binding the 3' untranslated region (UTR) of target mRNAs, miRNAs typically reduce their stability and/or translation. miRNAs are shown to be involved in regulating virtually every cellular process and their deregulation has been shown to contribute to many diseases. There are nearly 1,000 different miRNAs in humans, which regulate expression of thousands of other genes, reflecting that this is indeed a major layer of gene expression regulation. In my PhD, I had focused on these two different themes of estrogen signaling. In the first project, using a DNA microarray approach, we identified the target genes of the emerging new player in estrogen signaling, GPR30. In the second part, we have addressed some previously unexplained observations about the role of the 3ʼUTR of the ERα mRNA in regulating ERα levels and estrogen signaling

Multidirectional interplay between nuclear receptors and microRNAs

Nuclear receptors (NRs) form one of the largest superfamilies of transcription factors in metazoans. MicroRNAs (miRNAs) are small non-coding RNAs that bind the 3' untranslated region (3'UTR) of target mRNAs to reduce their stability and/or translation. miRNAs can directly regulate the protein output of target NR mRNAs, and, conversely, the expression of miRNAs can be modulated by NRs at the transcriptional level. At least one NR also regulates the posttranscriptional maturation of miRNAs by interacting with miRNA processing factors via NR co-regulators. Moreover, miRNAs regulate NR signaling by targeting the mRNAs of NR co-regulators and target genes. This complex set of interactions also leads to an extensive network of feedback and feedforward regulatory loops

Monoubiquitination of Histone H2B Blocks Eviction of Histone Variant H2A.Z from Inducible Enhancers

Covalent modifications of histones play a crucial role in the regulation of gene expression. Histone H2B monoubiquitination has mainly been described as a regulator of transcription elongation, but its role in transcription initiation is poorly documented. We investigated the role of this histone mark (H2Bub1) on different inducible enhancers, in particular those regulated by estrogen receptor a, by loss- and gain-of-function experiments with the specific E3-ubiquitin ligase complex of H2B: RNF20/RNF40. RNF20/RNF40 overexpression causes repression of the induced activity of these enhancers. Genome- wide profiles show that H2Bub1 levels are negatively correlated with the accessibility of enhancers to transcriptional activators. We found that the chromatin association of histone variant H2A.Z, which is evicted from enhancers for transcriptional activation, is stabilized by H2Bub1 by impairing access of the chromatin remodeler INO80. We propose that H2Bub1 acts as a gatekeeper of H2A.Z eviction and activation of inducible enhancers

Estrogenic GPR30 signalling induces proliferation and migration of breast cancer cells through CTGF

The steroid hormone oestrogen can signal through several receptors and pathways. Although the transcriptional responses mediated by the nuclear oestrogen receptors (ER) have been extensively characterized, the changes in gene expression elicited by signalling through the membrane-associated ER GPR30 have not been studied. We show here for ER-negative human breast cancer cells that the activation of GPR30 signalling by oestrogen or by hydroxytamoxifen (OHT), an ER antagonist but GPR30 agonist, induces a transcription factor network, which resembles that induced by serum in fibroblasts. The most strongly induced gene, CTGF, appears to be a target of these transcription factors. We found that the secreted factor connective tissue growth factor (CTGF) not only contributes to promote proliferation but also mediates the GPR30-induced stimulation of cell migration. These results provide a framework for understanding the physiological and pathological functions of GPR30. As the activation of GPR30 by OHT also induces CTGF in fibroblasts from breast tumour biopsies, these pathways may be involved in promoting aggressive behaviour of breast tumours in response to endogenous oestrogens or to OHT being used for endocrine therapy

Aggressiveness of non-EMT breast cancer cells relies on FBXO11 activity

Abstract Tumorigenesis is increasingly considered to rely on subclones of cells poised to undergo an epithelial to mesenchymal transition (EMT) program. We and others have provided evidence, however, that the tumorigenesis of human breast cancer is not always restricted to typical EMT cells but is also somewhat paradoxically conveyed by subclones of apparently differentiated, non-EMT cells. Here we characterize such non-EMT-like and EMT-like subclones. Through a loss-of-function screen we found that a member of the E3 ubiquitin ligase complexes, FBXO11, specifically fuels tumor formation of a non-EMT-like clone by restraining the p53/p21 pathway. Interestingly, in the related EMT-like clone, FBXO11 operates through the BCL2 pathway with little or no impact on tumorigenesis. These data command caution in attempts to assess tumorigenesis prospectively based on EMT profiling, and they emphasize the importance of next generation subtyping of tumors, that is at the level of clonal composition

Vps11 and Vps18 of Vps-C membrane traffic complexes are E3 ubiquitin ligases and fine-tune signalling

In response to extracellular signals, many signalling proteins associated with the plasma membrane are sorted into endosomes. This involves endosomal fusion, which depends on the complexes HOPS and CORVET. Whether and how their subunits themselves modulate signal transduction is unknown. We show that Vps11 and Vps18 (Vps11/18), two common subunits of the HOPS/CORVET complexes, are E3 ubiquitin ligases. Upon overexpression of Vps11/Vps18, we find perturbations of ubiquitination in signal transduction pathways. We specifically demonstrate that Vps11/18 regulate several signalling factors and pathways, including Wnt, estrogen receptor α (ERα), and NFκB. For ERα, we demonstrate that the Vps11/18-mediated ubiquitination of the scaffold protein PELP1 impairs the activation of ERα by c-Src. Thus, proteins involved in membrane traffic, in addition to performing their well-described role in endosomal fusion, fine-tune signalling in several different ways, including through ubiquitination