The rockefeller university

Coronavirus disease (COVID-19), caused by the virus SARS-CoV-2, is already responsible for more than 4.3 million confirmed cases and 295,000 deaths worldwide as of May 15, 2020. Ongoing efforts to control the pandemic include the development of peptide-based vaccines and diagnostic tests. In these approaches, HLA allelic diversity plays a crucial role. Despite its importance, current knowledge of HLA allele frequencies in South America is very limited. In this study, we have performed a literature review of datasets reporting HLA frequencies of South American populations, available in scientific literature and/or in the Allele Frequency Net Database. This allowed us to enrich the current scenario with more than 12.8 million data points. As a result, we are presenting updated HLA allelic frequencies based on country, including 91 alleles that were previously thought to have frequencies either under 5% or of an unknown value. Using alleles with an updated frequency of at least >= 5% in any South American country, we predicted epitopes in SARS-CoV-2 proteins using NetMHCpan (I and II) and MHC flurry. Then, the best predicted epitopes (class-I and -II) were selected based on their binding to South American alleles (Coverage Score). Class II predicted epitopes were also filtered based on their three-dimensional exposure. We obtained 14 class-I and four class-II candidate epitopes with experimental evidence (reported in the Immune Epitope Database and Analysis Resource), having good coverage scores for South America. Additionally, we are presenting 13 HLA-I and 30 HLA-II novel candidate epitopes without experimental evidence, including 16 class-II candidates in highly exposed conserved areas of the NTD and RBD regions of the Spike protein. These novel candidates have even better coverage scores for South America than those with experimental evidence. Finally, we show that recent similar studies presenting candidate epitopes also predicted some of our candidates but discarded them in the selection process, resulting in candidates with suboptimal coverage for South America. In conclusion, the candidate epitopes presented provide valuable information for the development of epitope-based strategies against SARS-CoV-2, such as peptide vaccines and diagnostic tests. Additionally, the updated HLA allelic frequencies provide a better representation of South America and may impact different immunogenetic studies.

Mendelian susceptibility to mycobacterial diseases (MSMD; Online Mendelian Inheritance in Man, OMIM #209950) is an inborn error of immunity (IEI) characterized by extreme susceptibility to invasive infections sustained by poorly virulent mycobacteria, including Mycobacterium bovis, bacillus Calmette-Guérin (BCG) vaccines, and environmental mycobacteria [1,2,3,4]. M. tuberculosis may also be involved in rare cases [5]. Many genes involved in interferon (IFN)-γ production (IL12B, IL12RB1, IL12RB2, IL23R, TYK2, ISG15, RORC), in response to IFN-γ (IFN-γR1, IFN-γR2, STAT1, JAK1, CYBB), both (IRF8, SPPL2A, NEMO) or IFN-γ itself are responsible for MSDM [4,5,6,7,8]. The clinical features depend on the genotype and the residual activity of IFN-γ.

Metabolic reprogramming is a key feature of many cancers, but how and when it contributes to tumorigenesis remains unclear. Here we demonstrate that metabolic reprogramming induced by mitochondrial fusion can be rate-limiting for immortalization of tumor-initiating cells (TICs) and trigger their irreversible dedication to tumorigenesis. Using single-cell transcriptomics, we find that Drosophila brain tumors contain a rapidly dividing stem cell population defined by upregulation of oxidative phosphorylation (OxPhos). We combine targeted metabolomics and in vivo genetic screening to demonstrate that OxPhos is required for tumor cell immortalization but dispensable in neural stem cells (NSCs) giving rise to tumors. Employing an in vivo NADH/NAD+ sensor, we show that NSCs precisely increase OxPhos during immortalization. Blocking OxPhos or mitochondrial fusion stalls TICs in quiescence and prevents tumorigenesis through impaired NAD+ regeneration. Our work establishes a unique connection between cellular metabolism and immortalization of tumor-initiating cells.

Muscle cell plasma membrane is frequently damaged by mechanical activity, and its repair requires the membrane protein dysferlin. We previously identified that, similar to dysferlin deficit, lack of annexin A2 (AnxA2) also impairs repair of skeletal myofibers. Here, we have studied the mechanism of AnxA2-mediated muscle cell membrane repair in cultured muscle cells. We find that injury-triggered increase in cytosolic calcium causes AnxA2 to bind dysferlin and accumulate on dysferlin-containing vesicles as well as with dysferlin at the site of membrane injury. AnxA2 accumulates on the injured plasma membrane in cholesterol-rich lipid microdomains and requires Src kinase activity and the presence of cholesterol. Lack of AnxA2 and its failure to translocate to the plasma membrane, both prevent calcium-triggered dysferlin translocation to the plasma membrane and compromise repair of the injured plasma membrane. Our studies identify that Anx2 senses calcium increase and injury-triggered change in plasma membrane cholesterol to facilitate dysferlin delivery and repair of the injured plasma membrane.

Characterization of MHC-bound peptides by mass spectrometry (MS) is an essential technique for immunologic studies. Many efforts have been made to quantify the number of MHC-presented ligands by MS and to define the limits of detection of a specific MHC ligand. However, these experiments are often complex and comparisons across different laboratories are challenging. Therefore, we compared and orthogonally validated quantitation of peptide:MHC complexes by radio immunoassay and flow cytometry using TCR mimic antibodies in three model systems to establish a method to control the experimental input of peptide MHC:complexes for MS analysis. Following isolation of MHC-bound peptides we identified and quantified an MHC ligand of interest with high correlation to the initial input. We found that the diversity of the presented ligandome, as well as the peptide sequence itself affected the detection of the target peptide. Furthermore, results were applicable from these model systems to unmodified cell lines with a tight correlation between HLA-A*02 complex input and the number of identified HLA-A*02 ligands. Overall, this framework provides an easily accessible experimental setup that offers the opportunity to control the peptide:MHC input and in this way compare immunopeptidome experiments not only within but also between laboratories, independent of their experimental approach. Significance: Although immunopeptidomics is an essential tool for the characterization of MHCbound peptides on the cell surface, there are no easily applicable established protocols available that allow comparison of immunopeptidome experiments across laboratories. Here, we demonstrate that controlling the peptide:MHC input for immunopurification and LC-MS/MS experiments by flow cytometry in pre-defined model systems allows the generation of qualitative and quantitative data that can easily be compared between investigators, independently of their methods for MHC ligand isolation for MS.

We present an approach for preparing cryo-electron microscopy (cryo-EM) grids to study short-lived molecular states. Using piezoelectric dispensing, two independent streams of similar to 50-pl droplets of sample are deposited within 10 ms of each other onto the surface of a nanowire EM grid, and the mixing reaction stops when the grid is vitrified in liquid ethane similar to 100 ms later. We demonstrate this approach for four biological systems where short-lived states are of high interest.

The Van der Pol equation is a paradigmatic model of relaxation oscillations. This remarkable nonlinear phenomenon of self-sustained oscillatory motion underlies important rhythmic processes in nature and electrical engineering. Relaxation oscillations in a real system are usually coupled to environmental noise, which further enriches their dynamics, but makes theoretical analysis of such systems and determination of the equation parameter values a difficult task. In a companion paper we have proposed an analytic approach to a similar problem for another classical nonlinear model-the bistable Duffing oscillator. Here we extend our techniques to the case of the Van der Pol equation driven by white noise. We analyze the statistics of solutions and propose a method to estimate parameter values from the oscillator's time series. We use experimental data of active oscillations in a biophysical system to demonstrate how our method applies to real observations and can be generalized for more complex models.

Data on the efficacy and safety of interferon (IFN)-alpha for the treatment of essential thrombocythemia (ET) and polycythemia vera (PV) are inconsistent. We conducted a systematic review and meta-analysis and searched MEDLINE and EMBASE via Ovid, Scopus, COCHRANE registry of clinical trials, and Web of Science from inception through 03/2019 for studies of pegylated IFN (peg-IFN) and non-pegylated IFN (non-peg-IFN) in PV and ET patients. Random-effects models were used to pool response rates for the primary outcome of overall response rate (ORR) defined as a composite of complete response, partial response, complete hematologic response (CHR) and partial hematologic response. Peg-IFN and non-peg-IFN were compared by meta-regression analyses. In total, 44 studies with 1359 patients (730 ET, 629 PV) were included. ORR were 80.6% (95% confidence interval: 76.6-84.1%, CHR: 59.0% [51.5%-66.1%]) and 76.7% (67.4-84.0%; CHR: 48.5% [37.8-59.4%]) for ET and PV patients, respectively. In meta-regression analyses results did not differ significantly for non-peg-IFN vs. peg-IFN. Annualized rates of thromboembolic complications and treatment discontinuation due to adverse events were low at 1.2% and 8.8% for ET and 0.5% and 6.5% for PV patients, respectively. Both peg-IFN and non-peg-IFN can be effective and safe long-term treatments for ET and PV.

Symmetry breaking is essential for polarization of cells and generation of left-right body asymmetry. Here the authors investigate the arrangement of hair cells in zebrafish and show that mirror-symmetric patterns arise from a combination of biochemical and mechanical symmetry-breaking events. Actively regulated symmetry breaking, which is ubiquitous in biological cells, underlies phenomena such as directed cellular movement and morphological polarization. Here, we investigate how an organ-level polarity pattern emerges through symmetry breaking at the cellular level during the formation of a mechanosensory organ. Combining theory, genetic perturbations and in vivo imaging, we study the development and regeneration of the fluid-motion sensors in the zebrafish's lateral line. We find that two interacting symmetry-breaking events-one mediated by biochemical signalling and the other by cellular mechanics-give rise to precise rotations of cell pairs, which produce a mirror-symmetric polarity pattern in the receptor organ.

The characterization of specific metaboliteprotein interactions is important in chemical biology and drug discovery. For example, nuclear receptors (NRs) are a family of ligand-activated transcription factors that regulate diverse physiological processes in animals and are key targets for therapeutic development. However, the identification and characterization of physiological ligands for many NRs remains challenging, because of limitations in domain-specific analysis of ligand binding in cells. To address these limitations, we developed a domain-specific covalent chemical reporter for peroxisome proliferator-activated receptors (PPARs) and demonstrated its utility to screen and characterize the potency of candidate NR ligands in live cells. These studies demonstrate targeted and domain-specific chemical reporters provide excellent tools to evaluate endogenous and exogenous (diet, microbiota, therapeutics) ligands of PPARs in mammalian cells, as well as additional protein targets for further investigation.