scMAPA: Identification of cell-type-specific alternative polyadenylation in complex tissues.

BACKGROUND: Alternative polyadenylation (APA) causes shortening or lengthening of the 3'-untranslated region (3'-UTR) of genes (APA genes) in diverse cellular processes such as cell proliferation and differentiation. To identify cell-type-specific APA genes in scRNA-Seq data, current bioinformatic methods have several limitations. First, they assume certain read coverage shapes in the scRNA-Seq data, which can be violated in multiple APA genes. Second, their identification is limited between 2 cell types and not directly applicable to the data of multiple cell types. Third, they do not control undesired source of variance, which potentially introduces noise to the cell-type-specific identification of APA genes. FINDINGS: We developed a combination of a computational change-point algorithm and a statistical model, single-cell Multi-group identification of APA (scMAPA). To avoid the assumptions on the read coverage shape, scMAPA formulates a change-point problem after transforming the 3' biased scRNA-Seq data to represent the full-length 3'-UTR signal. To identify cell-type-specific APA genes while adjusting for undesired source of variation, scMAPA models APA isoforms in consideration of the cell types and the undesired source. In our novel simulation data and data from human peripheral blood mononuclear cells, scMAPA outperforms existing methods in sensitivity, robustness, and stability. In mouse brain data consisting of multiple cell types sampled from multiple regions, scMAPA identifies cell-type-specific APA genes, elucidating novel roles of APA for dividing immune cells and differentiated neuron cells and in multiple brain disorders. CONCLUSIONS: scMAPA elucidates the cell-type-specific function of APA events and sheds novel insights into the functional roles of APA events in complex tissues.

Tim-3 mediates T cell trogocytosis to limit antitumor immunity.

T cell immunoglobulin mucin domain-containing protein 3 (Tim-3) negatively regulates innate and adaptive immunity in cancer. To identify the mechanisms of Tim-3 in cancer immunity, we evaluated the effects of Tim-3 blockade in human and mouse melanoma. Here, we show that human programmed cell death 1-positive (PD-1+) Tim-3+CD8+ tumor-infiltrating lymphocytes (TILs) upregulate phosphatidylserine (PS), a receptor for Tim-3, and acquire cell surface myeloid markers from antigen-presenting cells (APCs) through transfer of membrane fragments called trogocytosis. Tim-3 blockade acted on Tim-3+ APCs in a PS-dependent fashion to disrupt the trogocytosis of activated tumor antigen-specific CD8+ T cells and PD-1+Tim-3+ CD8+ TILs isolated from patients with melanoma. Tim-3 and PD-1 blockades cooperated to disrupt trogocytosis of CD8+ TILs in 2 melanoma mouse models, decreasing tumor burden and prolonging survival. Deleting Tim-3 in dendritic cells but not in CD8+ T cells impeded the trogocytosis of CD8+ TILs in vivo. Trogocytosed CD8+ T cells presented tumor peptide-major histocompatibility complexes and became the target of fratricide T cell killing, which was reversed by Tim-3 blockade. Our findings have uncovered a mechanism Tim-3 uses to limit antitumor immunity.

Intestinal microbiota signatures of clinical response and immune-related adverse events in melanoma patients treated with anti-PD-1.

Ample evidence indicates that the gut microbiome is a tumor-extrinsic factor associated with antitumor response to anti-programmed cell death protein-1 (PD-1) therapy, but inconsistencies exist between published microbial signatures associated with clinical outcomes. To resolve this, we evaluated a new melanoma cohort, along with four published datasets. Time-to-event analysis showed that baseline microbiota composition was optimally associated with clinical outcome at approximately 1 year after initiation of treatment. Meta-analysis and other bioinformatic analyses of the combined data show that bacteria associated with favorable response are confined within the Actinobacteria phylum and the Lachnospiraceae/Ruminococcaceae families of Firmicutes. Conversely, Gram-negative bacteria were associated with an inflammatory host intestinal gene signature, increased blood neutrophil-to-lymphocyte ratio, and unfavorable outcome. Two microbial signatures, enriched for Lachnospiraceae spp. and Streptococcaceae spp., were associated with favorable and unfavorable clinical response, respectively, and with distinct immune-related adverse effects. Despite between-cohort heterogeneity, optimized all-minus-one supervised learning algorithms trained on batch-corrected microbiome data consistently predicted outcomes to programmed cell death protein-1 therapy in all cohorts. Gut microbial communities (microbiotypes) with nonuniform geographical distribution were associated with favorable and unfavorable outcomes, contributing to discrepancies between cohorts. Our findings shed new light on the complex interaction between the gut microbiome and response to cancer immunotherapy, providing a roadmap for future studies.

Fecal microbiota transplant overcomes resistance to anti-PD-1 therapy in melanoma patients.

Anti-programmed cell death protein 1 (PD-1) therapy provides long-term clinical benefits to patients with advanced melanoma. The composition of the gut microbiota correlates with anti-PD-1 efficacy in preclinical models and cancer patients. To investigate whether resistance to anti-PD-1 can be overcome by changing the gut microbiota, this clinical trial evaluated the safety and efficacy of responder-derived fecal microbiota transplantation (FMT) together with anti-PD-1 in patients with PD-1-refractory melanoma. This combination was well tolerated, provided clinical benefit in 6 of 15 patients, and induced rapid and durable microbiota perturbation. Responders exhibited increased abundance of taxa that were previously shown to be associated with response to anti-PD-1, increased CD8+ T cell activation, and decreased frequency of interleukin-8-expressing myeloid cells. Responders had distinct proteomic and metabolomic signatures, and transkingdom network analyses confirmed that the gut microbiome regulated these changes. Collectively, our findings show that FMT and anti-PD-1 changed the gut microbiome and reprogrammed the tumor microenvironment to overcome resistance to anti-PD-1 in a subset of PD-1 advanced melanoma.

Role of animal movement and indirect contact among farms in transmission of porcine epidemic diarrhea virus.

Epidemiological models of the spread of pathogens in livestock populations primarily focus on direct contact between farms based on animal movement data, and in some cases, local spatial spread based on proximity between premises. The roles of other types of indirect contact among farms is rarely accounted for. In addition, data on animal movements is seldom available in the United States. However, the spread of porcine epidemic diarrhea virus (PEDv) in U.S. swine represents one of the best documented emergences of a highly infectious pathogen in the U.S. livestock industry, providing an opportunity to parameterize models of pathogen spread via direct and indirect transmission mechanisms in swine. Using observed data on pig movements during the initial phase of the PEDv epidemic, we developed a network-based and spatially explicit epidemiological model that simulates the spread of PEDv via both indirect and direct movement-related contact in order to answer unresolved questions concerning factors facilitating between-farm transmission. By modifying the likelihood of each transmission mechanism and fitting this model to observed epidemiological dynamics, our results suggest that between-farm transmission was primarily driven by direct mechanisms related to animal movement and indirect mechanisms related to local spatial spread based on geographic proximity. However, other forms of indirect transmission among farms, including contact via contaminated vehicles and feed, were responsible for high consequence transmission events resulting in the introduction of the virus into new geographic areas. This research is among the first reports of farm-level animal movements in the U.S. swine industry and, to our knowledge, represents the first epidemiological model of commercial U.S. swine using actual data on farm-level animal movement.

A Probabilistic Approach to Receptive Field Mapping in the Frontal Eye Fields.

Studies of the neuronal mechanisms of perisaccadic vision often lack the resolution needed to determine important changes in receptive field (RF) structure. Such limited analytical power can lead to inaccurate descriptions of visuomotor processing. To address this issue, we developed a precise, probabilistic technique that uses a generalized linear model (GLM) for mapping the visual RFs of frontal eye field (FEF) neurons during stable fixation (Mayo et al., 2015). We previously found that full-field RF maps could be obtained using 1-8 dot stimuli presented at frame rates of 10-150 ms. FEF responses were generally robust to changes in the number of stimuli presented or the rate of presentation, which allowed us to visualize RFs over a range of spatial and temporal resolutions. Here, we compare the quality of RFs obtained over different stimulus and GLM parameters to facilitate future work on the detailed mapping of FEF RFs. We first evaluate the interactions between the number of stimuli presented per trial, the total number of trials, and the quality of RF mapping. Next, we vary the spatial resolution of our approach to illustrate the tradeoff between visualizing RF sub-structure and sampling at high resolutions. We then evaluate local smoothing as a possible correction for situations where under-sampling occurs. Finally, we provide a preliminary demonstration of the usefulness of a probabilistic approach for visualizing full-field perisaccadic RF shifts. Our results present a powerful, and perhaps necessary, framework for studying perisaccadic vision that is applicable to FEF and possibly other visuomotor regions of the brain.