COVID-19 in Correctional and Detention Facilities - United States, February-April 2020.

An estimated 2.1 million U.S. adults are housed within approximately 5,000 correctional and detention facilities† on any given day (1). Many facilities face significant challenges in controlling the spread of highly infectious pathogens such as SARS-CoV-2, the virus that causes coronavirus disease 2019 (COVID-19). Such challenges include crowded dormitories, shared lavatories, limited medical and isolation resources, daily entry and exit of staff members and visitors, continual introduction of newly incarcerated or detained persons, and transport of incarcerated or detained persons in multiperson vehicles for court-related, medical, or security reasons (2,3). During April 22-28, 2020, aggregate data on COVID-19 cases were reported to CDC by 37 of 54 state and territorial health department jurisdictions. Thirty-two (86%) jurisdictions reported at least one laboratory-confirmed case from a total of 420 correctional and detention facilities. Among these facilities, COVID-19 was diagnosed in 4,893 incarcerated or detained persons and 2,778 facility staff members, resulting in 88 deaths in incarcerated or detained persons and 15 deaths among staff members. Prompt identification of COVID-19 cases and consistent application of prevention measures, such as symptom screening and quarantine, are critical to protecting incarcerated and detained persons and staff members.

Mitochondrial uncoupler BAM15 reverses diet-induced obesity and insulin resistance in mice.

Obesity is a health problem affecting more than 40% of US adults and 13% of the global population. Anti-obesity treatments including diet, exercise, surgery and pharmacotherapies have so far failed to reverse obesity incidence. Herein, we target obesity with a pharmacotherapeutic approach that decreases caloric efficiency by mitochondrial uncoupling. We show that a recently identified mitochondrial uncoupler BAM15 is orally bioavailable, increases nutrient oxidation, and decreases body fat mass without altering food intake, lean body mass, body temperature, or biochemical and haematological markers of toxicity. BAM15 decreases hepatic fat, decreases inflammatory lipids, and has strong antioxidant effects. Hyperinsulinemic-euglycemic clamp studies show that BAM15 improves insulin sensitivity in multiple tissue types. Collectively, these data demonstrate that pharmacologic mitochondrial uncoupling with BAM15 has powerful anti-obesity and insulin sensitizing effects without compromising lean mass or affecting food intake.

Proteome and Phosphoproteome Analysis of Brown Adipocytes Reveals That RICTOR Loss Dampens Global Insulin/AKT Signaling.

Stimulating brown adipose tissue (BAT) activity represents a promising therapy for overcoming metabolic diseases. mTORC2 is important for regulating BAT metabolism, but its downstream targets have not been fully characterized. In this study, we apply proteomics and phosphoproteomics to investigate the downstream effectors of mTORC2 in brown adipocytes. We compare wild-type controls to isogenic cells with an induced knockout of the mTORC2 subunit RICTOR (Rictor-iKO) by stimulating each with insulin for a 30-min time course. In Rictor-iKO cells, we identify decreases to the abundance of glycolytic and de novo lipogenesis enzymes, and increases to mitochondrial proteins as well as a set of proteins known to increase upon interferon stimulation. We also observe significant differences to basal phosphorylation because of chronic RICTOR loss including decreased phosphorylation of the lipid droplet protein perilipin-1 in Rictor-iKO cells, suggesting that RICTOR could be involved with regulating basal lipolysis or droplet dynamics. Finally, we observe mild dampening of acute insulin signaling response in Rictor-iKO cells, and a subset of AKT substrates exhibiting statistically significant dependence on RICTOR.

Thesaurus: quantifying phosphopeptide positional isomers.

Proteins can be phosphorylated at neighboring sites resulting in different functional states, and studying the regulation of these sites has been challenging. Here we present Thesaurus, a search engine that detects and quantifies phosphopeptide positional isomers from parallel reaction monitoring and data-independent acquisition mass spectrometry experiments. We apply Thesaurus to analyze phosphorylation events in the PI3K/AKT signaling pathway and show neighboring sites with distinct regulation.

Funding Hepatitis C Treatment in Correctional Facilities by Using a Nominal Pricing Mechanism.

The cost of treating all incarcerated people who have hepatitis C with direct-acting antiviral agents (DAAs) greatly stresses correctional facility budgets. Complex federal laws bar pharmaceutical companies from simply discounting expensive medications to prices that facilities can afford. This article discusses means by which correctional facilities may qualify under federal law as "safety-net providers" to allow sale of DAAs at a price <10% of the average manufacturer price (AMP). No new laws would need to be enacted to implement this strategy. Using fiscal year 2018 pricing data from the Georgia Department of Corrections, we derived an estimate for the AMP and then used this estimate to calculate a nominal price. The United States would save ∼$3 billion if manufacturers sold DAAs at a nominal price to correctional facilities. Use of this strategy would help solve the conundrum of how state and county governments can pay for hepatitis C treatment and would ultimately save money for society.

Chromatogram libraries improve peptide detection and quantification by data independent acquisition mass spectrometry.

Data independent acquisition (DIA) mass spectrometry is a powerful technique that is improving the reproducibility and throughput of proteomics studies. Here, we introduce an experimental workflow that uses this technique to construct chromatogram libraries that capture fragment ion chromatographic peak shape and retention time for every detectable peptide in a proteomics experiment. These coordinates calibrate protein databases or spectrum libraries to a specific mass spectrometer and chromatography setup, facilitating DIA-only pipelines and the reuse of global resource libraries. We also present EncyclopeDIA, a software tool for generating and searching chromatogram libraries, and demonstrate the performance of our workflow by quantifying proteins in human and yeast cells. We find that by exploiting calibrated retention time and fragmentation specificity in chromatogram libraries, EncyclopeDIA can detect 20-25% more peptides from DIA experiments than with data dependent acquisition-based spectrum libraries alone.

Five Questions Concerning Managing Hepatitis C in the Justice System: Finding Practical Solutions for Hepatitis C Virus Elimination.

An estimated 30% of Americans with hepatitis C virus (HCV) pass through a jail or prison annually. One in 7 incarcerated persons is viremic. Screening and treatment is cost-effective and beneficial to society as a whole. Yet at current (2018) levels of funding for HCV management, prisons are not aggressively seeking cases; few incarcerated persons with HCV actually receive treatment. This article explores barriers to screening for and treating hepatitis C in state prisons, and ways that states may overcome these barriers, such as nominal pricing. While high prices for direct-acting antivirals discourage treatment, potential strategies exist to lower prices.

The phosphatidic acid-binding, polybasic domain is responsible for the differences in the phosphoregulation of lipins 1 and 3.

Lipins 1, 2, and 3 are Mg2+-dependent phosphatidic acid phosphatases and catalyze the penultimate step of triacylglycerol synthesis. We have previously investigated the biochemistry of lipins 1 and 2 and shown that di-anionic phosphatidic acid (PA) augments their activity and lipid binding and that lipin 1 activity is negatively regulated by phosphorylation. In the present study, we show that phosphorylation does not affect the catalytic activity of lipin 3 or its ability to associate with PA in vitro The lipin proteins each contain a conserved polybasic domain (PBD) composed of nine lysine and arginine residues located between the conserved N- and C-terminal domains. In lipin 1, the PBD is the site of PA binding and sensing of the PA electrostatic charge. The specific arrangement and number of the lysines and arginines of the PBD vary among the lipins. We show that the different PBDs of lipins 1 and 3 are responsible for the presence of phosphoregulation on the former but not the latter enzyme. To do so, we generated lipin 1 that contained the PBD of lipin 3 and vice versa. The lipin 1 enzyme with the lipin 3 PBD lost its ability to be regulated by phosphorylation but remained downstream of phosphorylation by mammalian target of rapamycin. Conversely, the presence of the lipin 1 PBD in lipin 3 subjected the enzyme to negative intramolecular control by phosphorylation. These results indicate a mechanism for the observed differences in lipin phosphoregulation in vitro.

An atlas of human kinase regulation.

The coordinated regulation of protein kinases is a rapid mechanism that integrates diverse cues and swiftly determines appropriate cellular responses. However, our understanding of cellular decision-making has been limited by the small number of simultaneously monitored phospho-regulatory events. Here, we have estimated changes in activity in 215 human kinases in 399 conditions derived from a large compilation of phosphopeptide quantifications. This atlas identifies commonly regulated kinases as those that are central in the signaling network and defines the logic relationships between kinase pairs. Co-regulation along the conditions predicts kinase-complex and kinase-substrate associations. Additionally, the kinase regulation profile acts as a molecular fingerprint to identify related and opposing signaling states. Using this atlas, we identified essential mediators of stem cell differentiation, modulators of Salmonella infection, and new targets of AKT1. This provides a global view of human phosphorylation-based signaling and the necessary context to better understand kinase-driven decision-making.

Plug-and-play analysis of the human phosphoproteome by targeted high-resolution mass spectrometry.

Systematic approaches to studying cellular signaling require phosphoproteomic techniques that reproducibly measure the same phosphopeptides across multiple replicates, conditions, and time points. Here we present a method to mine information from large-scale, heterogeneous phosphoproteomics data sets to rapidly generate robust targeted mass spectrometry (MS) assays. We demonstrate the performance of our method by interrogating the IGF-1/AKT signaling pathway, showing that even rarely observed phosphorylation events can be consistently detected and precisely quantified.

The proteomic landscape of triple-negative breast cancer.

Triple-negative breast cancer is a heterogeneous disease characterized by poor clinical outcomes and a shortage of targeted treatment options. To discover molecular features of triple-negative breast cancer, we performed quantitative proteomics analysis of twenty human-derived breast cell lines and four primary breast tumors to a depth of more than 12,000 distinct proteins. We used this data to identify breast cancer subtypes at the protein level and demonstrate the precise quantification of biomarkers, signaling proteins, and biological pathways by mass spectrometry. We integrated proteomics data with exome sequence resources to identify genomic aberrations that affect protein expression. We performed a high-throughput drug screen to identify protein markers of drug sensitivity and understand the mechanisms of drug resistance. The genome and proteome provide complementary information that, when combined, yield a powerful engine for therapeutic discovery. This resource is available to the cancer research community to catalyze further analysis and investigation.

Genetic inhibition of hepatic acetyl-CoA carboxylase activity increases liver fat and alters global protein acetylation.

Lipid deposition in the liver is associated with metabolic disorders including fatty liver disease, type II diabetes, and hepatocellular cancer. The enzymes acetyl-CoA carboxylase 1 (ACC1) and ACC2 are powerful regulators of hepatic fat storage; therefore, their inhibition is expected to prevent the development of fatty liver. In this study we generated liver-specific ACC1 and ACC2 double knockout (LDKO) mice to determine how the loss of ACC activity affects liver fat metabolism and whole-body physiology. Characterization of LDKO mice revealed unexpected phenotypes of increased hepatic triglyceride and decreased fat oxidation. We also observed that chronic ACC inhibition led to hyper-acetylation of proteins in the extra-mitochondrial space. In sum, these data reveal the existence of a compensatory pathway that protects hepatic fat stores when ACC enzymes are inhibited. Furthermore, we identified an important role for ACC enzymes in the regulation of protein acetylation in the extra-mitochondrial space.

Lipin 2 binds phosphatidic acid by the electrostatic hydrogen bond switch mechanism independent of phosphorylation.

Lipin 2 is a phosphatidic acid phosphatase (PAP) responsible for the penultimate step of triglyceride synthesis and dephosphorylation of phosphatidic acid (PA) to generate diacylglycerol. The lipin family of PA phosphatases is composed of lipins 1-3, which are members of the conserved haloacid dehalogenase superfamily. Although genetic alteration of LPIN2 in humans is known to cause Majeed syndrome, little is known about the biochemical regulation of its PAP activity. Here, in an attempt to gain a better general understanding of the biochemical nature of lipin 2, we have performed kinetic and phosphorylation analyses. We provide evidence that lipin 2, like lipin 1, binds PA via the electrostatic hydrogen bond switch mechanism but has a lower rate of catalysis. Like lipin 1, lipin 2 is highly phosphorylated, and we identified 15 phosphosites. However, unlike lipin 1, the phosphorylation of lipin 2 is not induced by insulin signaling nor is it sensitive to inhibition of the mammalian target of rapamycin. Importantly, phosphorylation of lipin 2 does not negatively regulate either membrane binding or PAP activity. This suggests that lipin 2 functions as a constitutively active PA phosphatase in stark contrast to the high degree of phosphorylation-mediated regulation of lipin 1. This knowledge of lipin 2 regulation is important for a deeper understanding of how the lipin family functions with respect to lipid synthesis and, more generally, as an example of how the membrane environment around PA can influence its effector proteins.

A practical recipe to survey phosphoproteomes.

The field of cellular signaling is fueled by the discovery of novel protein phosphorylation events. Phosphoproteomics focuses on the large-scale identification and characterization of serine, threonine, and tyrosine phosphorylation of proteins. Phosphopeptide enrichment followed by mass spectrometry has emerged as the most powerful technique for unbiased, discovery-driven analysis by offering high sensitivity, resolution, and speed. Methods for mass spectrometry-based phosphoproteomics analysis have improved substantially over the last decade, making the discipline more approachable to the broader scientific community. Herein we describe the status of the field of phosphoproteomics and provide a robust workflow covering the major aspects of large-scale phosphorylation analysis from phosphopeptide enrichment via IMAC to data analysis.

Not all fat is equal: differential gene expression and potential therapeutic targets in subcutaneous adipose, visceral adipose, and endometrium of obese women with and without endometrial cancer.

OBJECTIVE: To identify obesity-related cancer genes in endometrial and adipose tissue depots of body mass index-matched morbidly obese women with and without endometrial cancer. METHODS: Eight women undergoing hysterectomy (4 women with and 4 women without endometrial cancer) were matched by age (52.6 years) and body mass index (44.5 kg/m). Endometrium, visceral adipose tissue, and subcutaneous adipose tissue were collected and subjected to microarray analysis using Affymetrix Human Genome U133 Plus 2.0 Arrays. Gene set enrichment analysis used to extract biological information from the gene expression data and t test metric ranked and compared genes in the expression data set. Protein expression was measured in the endometrial samples, and serum was collected for hormone/metabolite assays. RESULTS: No significant differences were detected in hormone/metabolite levels between groups. Gene set enrichment analysis comparisons demonstrated that endometrial, visceral adipose and subcutaneous adipose tissues displayed 40, 47, and 38 alternatively regulated gene set pathways when comparing patients with and without cancer. Nineteen gene sets were alternately regulated in both visceral and subcutaneous adipose tissues; however, eighteen of these were regulated in the opposite direction. Five pathways were significantly and alternately regulated in all 3 tissue types and included glycolysis/gluconeogenesis, ribosome, peroxisome proliferator activated receptor signaling, pathogenic Escherichia coli infection, and natural killer-mediated cytotoxicity. In the malignant endometrium, liver kinase B1 underexpression was observed in all patients with cancer. Liver kinase B1 underexpression decreased adenosine monophosphate-activated protein kinase activity toward acetyl-CoA carboxylase and implied enhanced lipid biosynthesis in obesity-induced endometrial cancer. CONCLUSIONS: Subcutaneous and visceral adipose tissue depots have opposite patterns of gene expression in obese patients with and without endometrial cancer. The altered de novo lipogenesis and individual gene targets identified provide new potential targets for cancer treatment and prevention for at-risk obese women.

Quantitative proteomic analysis of the adipocyte plasma membrane.

The adipocyte is a key regulator of mammalian metabolism. To advance our understanding of this important cell, we have used quantitative proteomics to define the protein composition of the adipocyte plasma membrane (PM) in the presence and absence of insulin. Using this approach, we have identified a high confidence list of 486 PM proteins, 52 of which potentially represent novel cell surface proteins, including a member of the adiponectin receptor family and an unusually high number of hydrolases with no known function. Several novel insulin-responsive proteins including the sodium/hydrogen exchanger, NHE6 and the collagens III and VI were also identified, and we provide evidence of PM-ER association suggestive of a unique functional association between these two organelles in the adipocyte. Together these studies provide a wealth of potential therapeutic targets for the manipulation of adipocyte function and a valuable resource for metabolic research and PM biology.