Hepatic p53 is regulated by transcription factor FOXO1 and acutely controls glycogen homeostasis

The tumor suppressor p53 is involved in the adaptation of hepatic metabolism to nutrient availability. Acute deletion of p53 in the mouse liver affects hepatic glucose and triglyceride metabolism. However, long-term adaptations upon the loss of hepatic p53 and its transcriptional regulators are unknown. Here we show that short-term, but not chronic, liver-specific deletion of p53 in mice reduces liver glycogen levels, and we implicate the transcription factor forkhead box O1 protein (FOXO1) in the regulation of p53 and its target genes. We demonstrate that acute p53 deletion prevents glycogen accumulation upon refeeding, whereas a chronic loss of p53 associates with a compensational activation of the glycogen synthesis pathway. Moreover, we identify fasting-activated FOXO1 as a repressor of p53 transcription in hepatocytes. We show that this repression is relieved by inactivation of FOXO1 by insulin, which likely mediates the upregulation of p53 expression upon refeeding. Strikingly, we find that high-fat diet-induced insulin resistance with persistent FOXO1 activation not only blunted the regulation of p53 but also the induction of p53 target genes like p21 during fasting, indicating overlapping effects of both FOXO1 and p53 on target gene expression in a context-dependent manner. Thus, we conclude that p53 acutely controls glycogen storage in the liver and is linked to insulin signaling via FOXO1, which has important implications for our understanding of the hepatic adaptation to nutrient availability

Rapid Molecular Assays for Specific Detection and Quantitation of Loa loa Microfilaremia

Loa loa is a filarial nematode that infects over 10 million people in Africa. Most infections cause no symptoms, but individuals with large numbers of blood-stage microfilariae are at risk for fatal reactions to ivermectin, an antiparasitic agent used to treat and prevent infections with Onchocerca volvulus, a related filarial parasite that may occur alongside L. loa. To address the urgent need for a point-of-care L. loa diagnostic assay, we screened a Loa microfilaria gene expression library and identified 18 Loa-specific DNA targets. From two targets, we developed a novel, rapid quantitative PCR assay for estimating L. loa microfilaria burden. The assay is highly sensitive (detects a single microfilaria in 20 µL of blood) and correlates well with microfilaria counts obtained with conventional microscopic techniques. The assay is species-specific for L. loa compared with related filarial parasites (including O. volvulus) and can be used in its current form in resource-rich areas as a diagnostic tool for L. loa infection. Although modifications will be required to make point-of-care use feasible, our assay provides a proof of concept for a potentially valuable tool to identify individuals at risk for adverse reactions to ivermectin and to facilitate the implementation of filarial control programs

A Randomized Trial Evaluating Prosaptide™ for HIV-Associated Sensory Neuropathies: Use of an Electronic Diary to Record Neuropathic Pain

Objectives: To examine the efficacy and safety of Prosaptide™ (PRO) for the treatment of painful HIV-associated sensory neuropathies (HIV-SN). Design: A randomized, double-blind, placebo-controlled, multicenter study in participants with sensory neuropathy. Pain modulating therapy was discontinued prior to baseline. Participants were stratified by sural sensory nerve action potential (SNAP) amplitude. Participants were trained to use an electronic diary (ED) to record pain. Setting: Peripheral neuropathies are common complications of HIV infection. The pathogenesis is unknown and currently treatments are restricted to symptomatic measures. We examined PRO against placebo (PBO) for treatment of painful HIV-SN and performed a post-hoc evaluation of an electronic diary (ED) to record HIV-associated neuropathic pain. Participants: Eligible participants included adults with neurologist-confirmed painful HIV-SN.Interventions 2, 4, 8, or 16 mg/d PRO or PBO administered via subcutaneous (SC) injection for six weeks. Neurotoxic antiretroviral drug usage was held constant.Outcome Measures Changes from baseline in the weekly average of evaluable daily random prompts measuring pain using the Gracely pain scale and adverse events. Results: 237 participants were randomized. The study was stopped after a planned futility analysis. There were no between-group differences in the frequency of adverse events or laboratory toxicities. The 6-week mean (sd) Gracely pain scale changes were −0.12 (0.23), −0.24 (0.35), −0.15 (0.32), −0.18 (0.34), and −0.18 (0.32) for the 2, 4, 8, 16 mg, and PBO arms respectively. A similar variability of pain changes recorded using the ED were noted compared to previous trials that used paper collection methods.Conclusions 6-week treatment with PRO was safe but not effective at reducing HIV-associated neuropathic pain. Use of an ED to record neuropathic pain is novel in HIV-SN, resulted in reasonable compliance in recording pain data, but did not decrease the variability of pain scores compared to historical paper collection methods. Trial Registration: Current Controlled Trials NCT0028637

Structural and Functional Diversity of Acidic Scorpion Potassium Channel Toxins

Background: Although the basic scorpion K + channel toxins (KTxs) are well-known pharmacological tools and potential drug candidates, characterization the acidic KTxs still has the great significance for their potential selectivity towards different K + channel subtypes. Unfortunately, research on the acidic KTxs has been ignored for several years and progressed slowly. Principal Findings: Here, we describe the identification of nine new acidic KTxs by cDNA cloning and bioinformatic analyses. Seven of these toxins belong to three new a-KTx subfamilies (a-KTx28, a-KTx29, and a-KTx30), and two are new members of the known k-KTx2 subfamily. ImKTx104 containing three disulfide bridges, the first member of the a-KTx28 subfamily, has a low sequence homology with other known KTxs, and its NMR structure suggests ImKTx104 adopts a modified cystine-stabilized a-helix-loop-b-sheet (CS-a/b) fold motif that has no apparent a-helixs and b-sheets, but still stabilized by three disulfide bridges. These newly described acidic KTxs exhibit differential pharmacological effects on potassium channels. Acidic scorpion toxin ImKTx104 was the first peptide inhibitor found to affect KCNQ1 channel, which is insensitive to the basic KTxs and is strongly associated with human cardiac abnormalities. ImKTx104 selectively inhibited KCNQ1 channel with a Kd of 11.69 mM, but was less effective against the basic KTxs-sensitive potassium channels. In addition to the ImKTx104 toxin, HeTx204 peptide, containing a cystine-stabilized a-helix-loop-helix (CS-a/a) fold scaffold motif

A modified sequence capture approach allowing standard and methylation analyses of the same enriched genomic DNA sample

Background: Bread wheat has a large complex genome that makes whole genome resequencing costly. Therefore, genome complexity reduction techniques such as sequence capture make re-sequencing cost effective. With a high-quality draft wheat genome now available it is possible to design capture probe sets and to use them to accurately genotype and anchor SNPs to the genome. Furthermore, in addition to genetic variation, epigenetic variation provides a source of natural variation contributing to changes in gene expression and phenotype that can be profiled at the base pair level using sequence capture coupled with bisulphite treatment. Here, we present a new 12 Mbp wheat capture probe set, that allows both the profiling of genotype and methylation from the same DNA sample. Furthermore, we present a method, based on Agilent SureSelect Methyl-Seq, that will use a single capture assay as a starting point to allow both DNA sequencing and methyl-seq. Results: Our method uses a single capture assay that is sequentially split and used for both DNA sequencing and methyl-seq. The resultant genotype and epi-type data is highly comparable in terms of coverage and SNP/methylation site identification to that generated from separate captures for DNA sequencing and methyl-seq. Furthermore, by defining SNP frequencies in a diverse landrace from the Watkins collection we highlight the importance of having genotype data to prevent false positive methylation calls. Finally, we present the design of a new 12 Mbp wheat capture and demonstrate its successful application to re-sequence wheat. Conclusions: We present a cost-effective method for performing both DNA sequencing and methyl-seq from a single capture reaction thus reducing reagent costs, sample preparation time and DNA requirements for these complementary analyses

Catechol-O-Methyltransferase Val158Met Polymorphism Associates with Individual Differences in Sleep Physiologic Responses to Chronic Sleep Loss

Val158Met polymorphism was a novel marker in healthy adults of differential vulnerability to chronic partial sleep deprivation (PSD), a condition distinct from total sleep loss and one experienced by millions on a daily and persistent basis. allelic frequencies were higher in whites than African Americans.-related treatment responses and risk factors for symptom exacerbation

Resilience to Disturbance Despite Limited Dispersal and Self-Recruitment in Tropical Barrel Sponges: Implications for Conservation and Management

While estimates of connectivity are important for effective management, few such estimates are available for reef invertebrates other than for corals. Barrel sponges are one of the largest and most conspicuous members of the coral reef fauna across the Indo-Pacific and given their large size, longevity and ability to process large volumes of water, they have a major role in reef functioning. Here we used a panel of microsatellite markers to characterise the genetic structure of two barrel sponge species, Xestospongia testudinaria and a currently undescribed Xestospongia species. We sampled across seven populations in the Wakatobi Marine National Park, SE Sulawesi (Indonesia) spanning a spatial scale of approximately 2 to 70 km, and present the first estimates of demographic connectivity for coral reef sponges. Genetic analyses showed high levels of genetic differentiation between all populations for both species, but contrasting patterns of genetic structuring for the two species. Autocorrelation analyses showed the likely dispersal distances of both species to be in the order of 60 and 140 m for Xestopongia sp. and Xestospongia testudinaria, respectively, which was supported by assignment tests that showed high levels of self-recruitment (>80%). We also found consistently high inbreeding coefficients across all populations for both species. Our study highlights the potential susceptibility of barrel sponges to environmental perturbations because they are generally long-lived, slow growing, have small population sizes and are likely to be reliant on self-recruitment. Surprisingly, despite these features we actually found the highest abundance of both barrel sponge species (although they were generally smaller) at a site that has been severely impacted by humans over the last fifty years. This suggests that barrel sponges exhibit environmental adaptation to declining environmental quality and has important implications for the management and conservation of these important reef species. © 2014 Bell et al

Emerging role of the calcium-activated, small conductance, SK3 K ⁺ channel in distal tubule function: Regulation by TRPV4

The Ca2+-activated, maxi-K (BK) K+ channel, with low Ca2+-binding affinity, is expressed in the distal tubule of the nephron and contributes to flow-dependent K+ secretion. In the present study we demonstrate that the Ca2+-activated, SK3 (KCa2.3) K + channel, with high Ca2+-binding affinity, is also expressed in the mouse kidney (RT-PCR, immunoblots). Immunohistochemical evaluations using tubule specific markers demonstrate significant expression of SK3 in the distal tubule and the entire collecting duct system, including the connecting tubule (CNT) and cortical collecting duct (CCD). In CNT and CCD, main sites for K+ secretion, the highest levels of expression were along the apical (luminal) cell membranes, including for both principal cells (PCs) and intercalated cells (ICs), posturing the channel for Ca2+- dependent K+ secretion. Fluorescent assessment of cell membrane potential in native, split-opened CCD, demonstrated that selective activation of the Ca2+-permeable TRPV4 channel, thereby inducing Ca2+ influx and elevating intracellular Ca2+ levels, activated both the SK3 channel and the BK channel leading to hyperpolarization of the cell membrane. The hyperpolarization response was decreased to a similar extent by either inhibition of SK3 channel with the selective SK antagonist, apamin, or by inhibition of the BK channel with the selective antagonist, iberiotoxin (IbTX). Addition of both inhibitors produced a further depolarization, indicating cooperative effects of the two channels on Vm. It is concluded that SK3 is functionally expressed in the distal nephron and collecting ducts where induction of TRPV4-mediated Ca2+ influx, leading to elevated intracellular Ca2+ levels, activates this high Ca2+- affinity K+ channel. Further, with sites of expression localized to the apical cell membrane, especially in the CNT and CCD, SK3 is poised to be a key pathway for Ca2+-dependent regulation of membrane potential and K+ secretion. © 2014 Berrout et al

The Complete Genome Sequence of ‘Candidatus Liberibacter solanacearum’, the Bacterium Associated with Potato Zebra Chip Disease

Zebra Chip (ZC) is an emerging plant disease that causes aboveground decline of potato shoots and generally results in unusable tubers. This disease has led to multi-million dollar losses for growers in the central and western United States over the past decade and impacts the livelihood of potato farmers in Mexico and New Zealand. ZC is associated with ‘Candidatus Liberibacter solanacearum’, a fastidious alpha-proteobacterium that is transmitted by a phloem-feeding psyllid vector, Bactericera cockerelli Sulc. Research on this disease has been hampered by a lack of robust culture methods and paucity of genome sequence information for ‘Ca. L. solanacearum’. Here we present the sequence of the 1.26 Mbp metagenome of ‘Ca. L. solanacearum’, based on DNA isolated from potato psyllids. The coding inventory of the ‘Ca. L. solanacearum’ genome was analyzed and compared to related Rhizobiaceae to better understand ‘Ca. L. solanacearum’ physiology and identify potential targets to develop improved treatment strategies. This analysis revealed a number of unique transporters and pathways, all potentially contributing to ZC pathogenesis. Some of these factors may have been acquired through horizontal gene transfer. Taxonomically, ‘Ca. L. solanacearum’ is related to ‘Ca. L. asiaticus’, a suspected causative agent of citrus huanglongbing, yet many genome rearrangements and several gene gains/losses are evident when comparing these two Liberibacter. species. Relative to ‘Ca. L. asiaticus’, ‘Ca. L. solanacearum’ probably has reduced capacity for nucleic acid modification, increased amino acid and vitamin biosynthesis functionalities, and gained a high-affinity iron transport system characteristic of several pathogenic microbes

Assessment of Regional Variability in COVID-19 Outcomes Among Patients With Cancer in the United States.

Importance: The COVID-19 pandemic has had a distinct spatiotemporal pattern in the United States. Patients with cancer are at higher risk of severe complications from COVID-19, but it is not well known whether COVID-19 outcomes in this patient population were associated with geography. Objective: To quantify spatiotemporal variation in COVID-19 outcomes among patients with cancer. Design, Setting, and Participants: This registry-based retrospective cohort study included patients with a historical diagnosis of invasive malignant neoplasm and laboratory-confirmed SARS-CoV-2 infection between March and November 2020. Data were collected from cancer care delivery centers in the United States. Exposures: Patient residence was categorized into 9 US census divisions. Cancer center characteristics included academic or community classification, rural-urban continuum code (RUCC), and social vulnerability index. Main Outcomes and Measures: The primary outcome was 30-day all-cause mortality. The secondary composite outcome consisted of receipt of mechanical ventilation, intensive care unit admission, and all-cause death. Multilevel mixed-effects models estimated associations of center-level and census division-level exposures with outcomes after adjustment for patient-level risk factors and quantified variation in adjusted outcomes across centers, census divisions, and calendar time. Results: Data for 4749 patients (median [IQR] age, 66 [56-76] years; 2439 [51.4%] female individuals, 1079 [22.7%] non-Hispanic Black individuals, and 690 [14.5%] Hispanic individuals) were reported from 83 centers in the Northeast (1564 patients [32.9%]), Midwest (1638 [34.5%]), South (894 [18.8%]), and West (653 [13.8%]). After adjustment for patient characteristics, including month of COVID-19 diagnosis, estimated 30-day mortality rates ranged from 5.2% to 26.6% across centers. Patients from centers located in metropolitan areas with population less than 250 000 (RUCC 3) had lower odds of 30-day mortality compared with patients from centers in metropolitan areas with population at least 1 million (RUCC 1) (adjusted odds ratio [aOR], 0.31; 95% CI, 0.11-0.84). The type of center was not significantly associated with primary or secondary outcomes. There were no statistically significant differences in outcome rates across the 9 census divisions, but adjusted mortality rates significantly improved over time (eg, September to November vs March to May: aOR, 0.32; 95% CI, 0.17-0.58). Conclusions and Relevance: In this registry-based cohort study, significant differences in COVID-19 outcomes across US census divisions were not observed. However, substantial heterogeneity in COVID-19 outcomes across cancer care delivery centers was found. Attention to implementing standardized guidelines for the care of patients with cancer and COVID-19 could improve outcomes for these vulnerable patients