AMP-Activated Kinase Restricts Rift Valley Fever Virus Infection by Inhibiting Fatty Acid Synthesis

The cell intrinsic innate immune responses provide a first line of defense against viral infection, and often function by targeting cellular pathways usurped by the virus during infection. In particular, many viruses manipulate cellular lipids to form complex structures required for viral replication, many of which are dependent on de novo fatty acid synthesis. We found that the energy regulator AMPK, which potently inhibits fatty acid synthesis, restricts infection of the Bunyavirus, Rift Valley Fever Virus (RVFV), an important re-emerging arthropod-borne human pathogen for which there are no effective vaccines or therapeutics. We show restriction of RVFV both by AMPK and its upstream activator LKB1, indicating an antiviral role for this signaling pathway. Furthermore, we found that AMPK is activated during RVFV infection, leading to the phosphorylation and inhibition of acetyl-CoA carboxylase, the first rate-limiting enzyme in fatty acid synthesis. Activating AMPK pharmacologically both restricted infection and reduced lipid levels. This restriction could be bypassed by treatment with the fatty acid palmitate, demonstrating that AMPK restricts RVFV infection through its inhibition of fatty acid biosynthesis. Lastly, we found that this pathway plays a broad role in antiviral defense since additional viruses from disparate families were also restricted by AMPK and LKB1. Therefore, AMPK is an important component of the cell intrinsic immune response that restricts infection through a novel mechanism involving the inhibition of fatty acid metabolism

Gene Expression of the Tumour Suppressor LKB1 Is Mediated by Sp1, NF-Y and FOXO Transcription Factors

The serine/threonine kinase LKB1 is a tumour suppressor that regulates multiple biological pathways, including cell cycle control, cell polarity and energy metabolism by direct phosphorylation of 14 different AMP-activated protein kinase (AMPK) family members. Although many downstream targets have been described, the regulation of LKB1 gene expression is still poorly understood. In this study, we performed a functional analysis of the human LKB1 upstream regulatory region. We used 200 base pair deletion constructs of the 5′-flanking region fused to a luciferase reporter to identify the core promoter. It encompasses nucleotides −345 to +52 relative to the transcription start site and coincides with a DNase I hypersensitive site. Based on extensive deletion and substitution mutant analysis of the LKB1 promoter, we identified four cis-acting elements which are critical for transcriptional activation. Using electrophoretic mobility shift assays as well as chromatin immunoprecipitations, we demonstrate that the transcription factors Sp1, NF-Y and two forkhead box O (FOXO) family members FOXO3 and FOXO4 bind to these elements. Overexpression of these factors significantly increased the LKB1 promoter activity. Conversely, small interfering RNAs directed against NF-Y alpha and the two FOXO proteins greatly reduced endogenous LKB1 expression and phosphorylation of LKB1's main substrate AMPK in three different cell lines. Taken together, these results demonstrate that Sp1, NF-Y and FOXO transcription factors are involved in the regulation of LKB1 transcription

Post-translational regulation contributes to the loss of LKB1 expression through SIRT1 deacetylase in osteosarcomas

background: The most prevalent form of bone cancer is osteosarcoma (OS), which is associated with poor prognosis in case of metastases formation. Mice harbouring liver kinase B1 (LKB1+/−) develop osteoblastoma-like tumours. Therefore, we asked whether loss of LKB1 gene has a role in the pathogenesis of human OS. methods: Osteosarcomas (n=259) were screened for LKB1 and sirtuin 1 (SIRT1) protein expression using immunohistochemistry and western blot. Those cases were also screened for LKB1 genetic alterations by next-generation sequencing, Sanger sequencing, restriction fragment length polymorphism and fluorescence in situ hybridisation approaches. We studied LKB1 protein degradation through SIRT1 expression. MicroRNA expression investigations were also conducted to identify the microRNAs involved in the SIRT1/LKB1 pathway. results: Forty-one per cent (106 out of 259) OS had lost LKB1 protein expression with no evident genetic anomalies. We obtained evidence that SIRT1 impairs LKB1 protein stability, and that SIRT1 depletion leads to accumulation of LKB1 in OS cell lines resulting in growth arrest. Further investigations revealed the role of miR-204 in the regulation of SIRT1 expression, which impairs LKB1 stability. conclusions: We demonstrated the involvement of sequential regulation of miR-204/SIRT1/LKB1 in OS cases and showed a mechanism for the loss of expression of LKB1 tumour suppressor in this malignancy

Triad pattern algorithm for predicting strong promoter candidates in bacterial genomes

Abstract Background Bacterial promoters, which increase the efficiency of gene expression, differ from other promoters by several characteristics. This difference, not yet widely exploited in bioinformatics, looks promising for the development of relevant computational tools to search for strong promoters in bacterial genomes. Results We describe a new triad pattern algorithm that predicts strong promoter candidates in annotated bacterial genomes by matching specific patterns for the group I σ70 factors of Escherichia coli RNA polymerase. It detects promoter-specific motifs by consecutively matching three patterns, consisting of an UP-element, required for interaction with the α subunit, and then optimally-separated patterns of -35 and -10 boxes, required for interaction with the σ70 subunit of RNA polymerase. Analysis of 43 bacterial genomes revealed that the frequency of candidate sequences depends on the A+T content of the DNA under examination. The accuracy of in silico prediction was experimentally validated for the genome of a hyperthermophilic bacterium, Thermotoga maritima, by applying a cell-free expression assay using the predicted strong promoters. In this organism, the strong promoters govern genes for translation, energy metabolism, transport, cell movement, and other as-yet unidentified functions. Conclusion The triad pattern algorithm developed for predicting strong bacterial promoters is well suited for analyzing bacterial genomes with an A+T content of less than 62%. This computational tool opens new prospects for investigating global gene expression, and individual strong promoters in bacteria of medical and/or economic significance.</p

The management of patients with primary chronic anal fissure: a position paper

Anal fissure is one of the most common and painful proctologic diseases. Its treatment has long been discussed and several different therapeutic options have been proposed. In the last decades, the understanding of its pathophysiology has led to a progressive reduction of invasive and potentially invalidating treatments in favor of conservative treatment based on anal sphincter muscle relaxation. Despite some systematic reviews and an American position statement, there is ongoing debate about the best treatment for anal fissure. This review is aimed at identifying the best treatment option drawing on evidence-based medicine and on the expert advice of 6 colorectal surgeons with extensive experience in this field in order to produce an Italian position statement for anal fissures. While there is little chance of a cure with conservative behavioral therapy, medical treatment with calcium channel blockers, diltiazem and nifepidine or glyceryl trinitrate, had a considerable success rate ranging from 50 to 90%. Use of 0.4% glyceryl trinitrate in standardized fashion seems to have the best results despite a higher percentage of headache, while the use of botulinum toxin had inconsistent results. Nonresponding patients should undergo lateral internal sphincterotomy. The risk of incontinence after this procedure seems to have been overemphasized in the past. Only a carefully selected group of patients, without anal hypertonia, could benefit from anoplasty

Older People’s Adherence to Community-Based Group Exercise Programmes: A Multiple-Case Study

Physical inactivity is a global phenomenon, with estimates of one in four adults not being active enough to achieve health benefits, thus heightening the risk of developing non-communicable diseases. In order to realise the health and wellbeing gains associated with physical activity the behaviour must be sustained. Community-based group exercise programmes (CBGEP) utilising social support have been shown to be one means of not only increasing activity levels for older people, but sustaining physical activity. A mixed-methods systematic review revealed a gap in the literature around older people’s long-term adherence to real-life CBGEP within a UK context. This study therefore sought to address this gap by understanding older people’s ongoing adherence to CBGEP with a view to gaining further insight about which factors contribute to enabling people to sustain their physical activity levels. A multiple case study research design was employed to understand older people’s (≥ 60 years, n=27) adherence (≥ 69%, for ≥ 1 year) to three current CBGEP in the South- West of England. Qualitative data (participant observation, focus groups, documents, and interviews) were collected and analysed using inductive thematic analysis followed by the analytic technique of explanation building. In order to gain deeper insights into adherence, the humanisation framework was utilised in an a priori manner to further understand adherence from a humanising perspective. Quantitative data were analysed using descriptive statistics and used to set the context of the study. This study found that older people’s adherence to CBGEP was mediated through six factors: factors relating to the individual, the instructor, programme design, social features, participant perceived benefits, and a humanised exercise environment. These all served to explain older people’s adherence to CBGEP. The humanising qualities of these CBGEP must be considered if we wish to support older people in sustaining a physically active lifestyle as they age. These findings are of interest to practitioners and policy makers in how CBGEP serve to aid older people in maintaining a physically active lifestyle with a view to preventing non-communicable diseases and in maintaining social connectivity

Lawson criterion for ignition exceeded in an inertial fusion experiment

For more than half a century, researchers around the world have been engaged in attempts to achieve fusion ignition as a proof of principle of various fusion concepts. Following the Lawson criterion, an ignited plasma is one where the fusion heating power is high enough to overcome all the physical processes that cool the fusion plasma, creating a positive thermodynamic feedback loop with rapidly increasing temperature. In inertially confined fusion, ignition is a state where the fusion plasma can begin "burn propagation" into surrounding cold fuel, enabling the possibility of high energy gain. While "scientific breakeven" (i.e., unity target gain) has not yet been achieved (here target gain is 0.72, 1.37 MJ of fusion for 1.92 MJ of laser energy), this Letter reports the first controlled fusion experiment, using laser indirect drive, on the National Ignition Facility to produce capsule gain (here 5.8) and reach ignition by nine different formulations of the Lawson criterion