Rules of engagement promote polarity in RNA trafficking

Many cell biological pathways exhibit overall polarity (net movement of molecules in one direction) even though individual molecular interactions in the pathway are freely reversible. The A2 RNA trafficking pathway exhibits polarity in moving specific RNA molecules from the nucleus to localization sites in the myelin compartment of oligodendrocytes or dendritic spines in neurons. The A2 pathway is mediated by a ubiquitously expressed trans-acting trafficking factor (hnRNP A2) that interacts with a specific 11 nucleotide cis-acting trafficking sequence termed the A2 response element (A2RE) found in several localized RNAs. Five different molecular partners for hnRNP A2 have been identified in the A2 pathway: hnRNP A2 itself, transportin, A2RE RNA, TOG (tumor overexpressed gene) and hnRNP E1, each playing a key role in one particular step of the A2 pathway. Sequential interactions of hnRNP A2 with different molecular partners at each step mediate directed movement of trafficking intermediates along the pathway. Specific "rules of engagement" (both and, either or, only if) govern sequential interactions of hnRNP A2 with each of its molecular partners. Rules of engagement are defined experimentally using three component binding assays to measure differential binding of hnRNP A2 to one partner in the presence of each of the other partners in the pathway. Here we describe rules of engagement for hnRNP A2 binding to each of its molecular partners and discuss how these rules of engagement promote polarity in the A2 RNA trafficking pathway. For molecules with multiple binding partners, specific rules of engagement govern different molecular interactions. Rules of engagement are ultimately determined by structural relationships between binding sites on individual molecules. In the A2 RNA trafficking pathway rules of engagement governing interactions of hnRNP A2 with different binding partners provide the basis for polarity of movement of intermediates along the pathway

Genetic diversity and phylogeography of broomcorn millet (Panicum miliaceum L.) across Eurasia

Broomcorn millet (Panicum miliaceum L.) is one of the world's oldest cultivated cereals, with several lines of recent evidence indicating that it was grown in northern China from at least 10 000 cal bp. Additionally, a cluster of archaeobotanical records of P. miliaceum dated to at least 7000 cal bp exists in eastern Europe. These two centres of early records could either represent independent domestications or cross-continental movement of this cereal that would predate that of any other crop by some 2 millennia. Here, we analysed genetic diversity among 98 landrace accessions from across Eurasia using 16 microsatellite loci, to explore phylogeographic structure in the Old World range of this historically important crop. The major genetic split in the data divided the accessions into an eastern and a western grouping with an approximate boundary in northwestern China. A substantial number of accessions belonging to the ‘western’ genetic group were also found in northeastern China. Further resolution subdivided the western and eastern genepools into 2 and 4 clusters respectively, each showing clear geographic patterning. The genetic data are consistent with both the single and multiple domestication centre hypotheses and add specific detail to what these hypotheses would entail regarding the spread of broomcorn millet. Discrepancies exist between the predictions from the genetic data and the current archaeobotanical record, highlighting priorities for investigation into early farming in Central Asia

DNA immunization as a technology platform for monoclonal antibody induction

To combat the threat of many emerging infectious diseases, DNA immunization offers a unique and powerful approach to the production of high-quality monoclonal antibodies (mAbs) against various pathogens. Compared with traditional protein-based immunization approaches, DNA immunization is efficient for testing novel immunogen designs, does not require the production or purification of proteins from a pathogen or the use of recombinant protein technology and is effective at generating mAbs against conformation-sensitive targets. Although significant progress in the use of DNA immunization to generate mAbs has been made over the last two decades, the literature does not contain an updated summary of this experience. The current review provides a comprehensive analysis of the literature, including our own work, describing the use of DNA immunization to produce highly functional mAbs, in particular, those against emerging infectious diseases. Critical factors such as immunogen design, delivery approach, immunization schedule, use of immune modulators and the role of final boost immunization are discussed in detail

Keyboard Contamination in Intensive Care Unit: Is Cleaning Enough? Prospective Research of In Situ Effectiveness of a Tea Tree Oil (KTEO) Film

After the SARS-CoV-2 pandemic, disinfection practices and microbial load reduction have become even more important and rigorous. To determine the contamination of keyboard surface and the relative risk to transfer healthcare-associated pathogens to susceptible patients, as it frequently happens in Intensive Care Unit (ICU), a standard keyboard (SK), a cleanable keyless keyboard (KK) with smooth surface and a standard keyboard coated with a 3M Tegaderm film added with active essential oil (tea tree oil) (KTEO) were tested. S. aureus, including MRSA strains, were detected in ICU, with values ranging from 15% to 57%. Gram negative strains belonging to the Enterobacteriaceae family were also found with values ranging from 14% to 71%. Similar Gram positive and Gram negative strains were found on all surfaces, but with low percentage, and only environmental bacteria were detected using the settling plates method. The Microbial Challenge Test performed on KTEO showed high rates of decrease for all the pathogens with statistical significance both at 24 and 48h (p=0.003* and p=0.040*, respectively). Our results suggest that the use of KTEO may be a feasible strategy for reducing the transmission of pathogens in health care setting and may be complementary to surface cleaning protocols

Therapeutic targeting of Krüppel-like factor 4 abrogates microglial activation

<p>Abstract</p> <p>Background</p> <p>Neuroinflammation occurs as a result of microglial activation in response to invading micro-organisms or other inflammatory stimuli within the central nervous system. According to our earlier findings, Krüppel-like factor 4 (Klf4), a zinc finger transcription factor, is involved in microglial activation and subsequent release of proinflammatory cytokines, tumor necrosis factor alpha, macrophage chemoattractant protein-1 and interleukin-6 as well as proinflammatory enzymes, inducible nitric oxide synthase and cyclooxygenase-2 in lipopolysaccharide-treated microglial cells. Our current study focuses on finding the molecular mechanism of the anti-inflammatory activities of honokiol in lipopolysaccharide-treated microglia with emphasis on the regulation of Klf4.</p> <p>Methods</p> <p>For <it>in vitro </it>studies, mouse microglial BV-2 cell lines as well as primary microglia were treated with 500 ng/mL lipopolysaccharide as well as 1 μM and 10 μM of honokiol. We cloned full-length Klf4 cDNA in pcDNA3.1 expression vector and transfected BV-2 cells with this construct using lipofectamine for overexpression studies. For <it>in vivo </it>studies, brain tissues were isolated from BALB/c mice treated with 5 mg/kg body weight of lipopolysaccharide either with or without 2.5 or 5 mg/kg body weight of honokiol. Expression of Klf4, cyclooxygenase-2, inducible nitric oxide synthase and phospho-nuclear factor-kappa B was measured using immunoblotting. We also measured the levels of cytokines, reactive oxygen species and nitric oxide in different conditions.</p> <p>Results</p> <p>Our findings suggest that honokiol can substantially downregulate the production of proinflammatory cytokines and inflammatory enzymes in lipopolysaccharide-stimulated microglia. In addition, honokiol downregulates lipopolysaccharide-induced upregulation of both Klf4 and phospho-nuclear factor-kappa B in these cells. We also found that overexpression of Klf4 in BV-2 cells suppresses the anti-inflammatory action of honokiol.</p> <p>Conclusions</p> <p>Honokiol potentially reduces inflammation in activated microglia in a Klf4-dependent manner.</p

Assessing prediction of diabetes in older adults using different adiposity measures: a 7 year prospective study in 6,923 older men and women

The aim of this study was to examine whether waist circumference (WC) or WHR improve diabetes prediction beyond body mass index in older men and women, and to define optimal cut-off points. In this prospective study, non-diabetic men (n = 3,519) and women (n = 3,404) aged 60-79 years were followed up for 7 years. There were 169 and 128 incident cases of type 2 diabetes in men and women, respectively. BMI, WC and WHR all showed strong associations with incident type 2 diabetes independent of potential confounders. In men, the adjusted relative risks (top vs lowest quartile) were 4.71 (95% CI 2.45-9.03) for BMI, 3.53 (95% CI 1.92-6.48) for WC and 2.76 (95% CI 1.58-4.82) for WHR. For women, the corresponding relative risks were 4.10 (95% CI 2.16-7.79), 12.18 (95% CI 4.83-30.74) and 5.61 (95% CI 2.84-11.09) for BMI, WC and WHR, respectively. Receiver-operating characteristic curve analysis revealed similar associations for BMI and WC in predicting diabetes in men (AUC = 0.726 and 0.713, respectively); WHR was the weakest predictor (AUC = 0.656). In women, WC was a significantly stronger predictor (AUC = 0.780) than either BMI (AUC = 0.733) or WHR (AUC = 0.728; p &lt; 0.01 for both). Inclusion of both WC and BMI did not improve prediction beyond BMI alone in men or WC alone in women. Optimal sensitivity and specificity for the prediction of type 2 diabetes was observed at a WC of 100 cm in men and 92 cm in women. In older men, BMI and WC yielded similar prediction of risk of type 2 diabetes, whereas WC was clearly a superior predictor in older wome

Investigation of first ray mobility during gait by kinematic fluoroscopic imaging-a novel method

<p>Abstract</p> <p>Background</p> <p>It is often suggested that sagittal instability at the first tarso-metatarsal joint level is a primary factor for hallux valgus and that sagittal instability increases with the progression of the deformity. The assessment of the degree of vertical instability is usually made by clinical evaluation while any measurements mostly refer to a static assessment of medial ray mobility (i.e. the plantar/dorsal flexion in the sagittal plane). Testing methods currently available cannot attribute the degree of mobility to the corresponding anatomical joints making up the medial column of the foot. The aim of this study was to develop a technique which allows for a quantification of the in-vivo sagittal mobility of the joints of the medial foot column during the roll-over process under full weight bearing.</p> <p>Methods</p> <p>Mobility of first ray bones was investigated by dynamic distortion-free fluoroscopy (25 frames/s) of 14 healthy volunteers and 8 patients with manifested clinical instability of the first ray. A CAD-based evaluation method allowed the determination of mobility and relative displacements and rotations of the first ray bones within the sagittal plane during the stance phase of gait.</p> <p>Results</p> <p>Total flexion of the first ray was found to be 13.63 (SD 6.14) mm with the healthy volunteers and 13.06 (SD 8.01) mm with the patients (resolution: 0.245 mm/pixel). The dorsiflexion angle was 5.27 (SD 2.34) degrees in the healthy volunteers and increased to 5.56 (SD 3.37) degrees in the patients. Maximum rotations were found at the naviculo-cuneiform joints and least at the first tarso-metatarsal joint level in both groups.</p> <p>Conclusions</p> <p>Dynamic fluoroscopic assessment has been shown to be a valuable tool for characterisation of the kinematics of the joints of the medial foot column during gait.</p> <p>A significant difference in first ray flexion and angular rotation between the patients and healthy volunteers however could not be found.</p

Genes targeted by the estrogen and progesterone receptors in the human endometrial cell lines HEC1A and RL95-2

<p>Abstract</p> <p>Background</p> <p>When the steroid hormones estrogen and progesterone bind to nuclear receptors, they have transcriptional impact on target genes in the human endometrium. These transcriptional changes have a critical function in preparing the endometrium for embryo implantation.</p> <p>Methods</p> <p>382 genes were selected, differentially expressed in the receptive endometrium, to study their responsiveness of estrogen and progesterone. The endometrial cell lines HEC1A and RL95-2 were used as experimental models for the non-receptive and receptive endometrium, respectively. Putative targets for activated steroid hormone receptors were investigated by chromatin immunoprecipitation (ChIP) using receptor-specific antibodies. Promoter occupancy of the selected genes by steroid receptors was detected in ChIP-purified DNA by quantitative PCR (qPCR). Expression analysis by reverse transcriptase (RT)-PCR was used to further investigate hormone dependent mRNA expression regulation of a subset of genes.</p> <p>Results</p> <p>ChIP-qPCR analysis demonstrated that each steroid hormone receptor had distinct group of target genes in the endometrial cell lines. After estradiol treatment, expression of estrogen receptor target genes predominated in HEC1A cells (n = 137) compared to RL95-2 cells (n = 35). In contrast, expression of progesterone receptor target genes was higher in RL95-2 cells (n = 83) than in HEC1A cells (n = 7) after progesterone treatment. RT-PCR analysis of 20 genes demonstrated transcriptional changes after estradiol or progesterone treatment of the cell lines.</p> <p>Conclusions</p> <p>Combined results from ChIP-qPCR and RT-PCR analysis showed different patterns of steroid hormone receptor occupancy at target genes, corresponding to activation or suppression of gene expression after hormone treatment of HEC1A and RL95-2 cell lines.</p