Structure learning for gene regulatory networks.

Inference of biological network structures is often performed on high-dimensional data, yet is hindered by the limited sample size of high throughput "omics" data typically available. To overcome this challenge, often referred to as the "small n, large p problem," we exploit known organizing principles of biological networks that are sparse, modular, and likely share a large portion of their underlying architecture. We present SHINE-Structure Learning for Hierarchical Networks-a framework for defining data-driven structural constraints and incorporating a shared learning paradigm for efficiently learning multiple Markov networks from high-dimensional data at large p/n ratios not previously feasible. We evaluated SHINE on Pan-Cancer data comprising 23 tumor types, and found that learned tumor-specific networks exhibit expected graph properties of real biological networks, recapture previously validated interactions, and recapitulate findings in literature. Application of SHINE to the analysis of subtype-specific breast cancer networks identified key genes and biological processes for tumor maintenance and survival as well as potential therapeutic targets for modulating known breast cancer disease genes.

Inactivation of LATS1/2 drives luminal-basal plasticity to initiate basal-like mammary carcinomas.

Basal-like breast cancers, an aggressive breast cancer subtype that has poor treatment options, are thought to arise from luminal mammary epithelial cells that undergo basal plasticity through poorly understood mechanisms. Using genetic mouse models and ex vivo primary organoid cultures, we show that conditional co-deletion of the LATS1 and LATS2 kinases, key effectors of Hippo pathway signaling, in mature mammary luminal epithelial cells promotes the development of Krt14 and Sox9-expressing basal-like carcinomas that metastasize over time. Genetic co-deletion experiments revealed that phenotypes resulting from the loss of LATS1/2 activity are dependent on the transcriptional regulators YAP/TAZ. Gene expression analyses of LATS1/2-deleted mammary epithelial cells notably revealed a transcriptional program that associates with human basal-like breast cancers. Our study demonstrates in vivo roles for the LATS1/2 kinases in mammary epithelial homeostasis and luminal-basal fate control and implicates signaling networks induced upon the loss of LATS1/2 activity in the development of basal-like breast cancer.

Yap/Taz inhibit goblet cell fate to maintain lung epithelial homeostasis.

Proper lung function relies on the precise balance of specialized epithelial cells that coordinate to maintain homeostasis. Herein, we describe essential roles for the transcriptional regulators YAP/TAZ in maintaining lung epithelial homeostasis, reporting that conditional deletion of Yap and Wwtr1/Taz in the lung epithelium of adult mice results in severe defects, including alveolar disorganization and the development of airway mucin hypersecretion. Through in vivo lineage tracing and in vitro molecular experiments, we reveal that reduced YAP/TAZ activity promotes intrinsic goblet transdifferentiation of secretory airway epithelial cells. Global gene expression and chromatin immunoprecipitation sequencing (ChIP-seq) analyses suggest that YAP/TAZ act cooperatively with TEA domain (TEAD) transcription factors and the NuRD complex to suppress the goblet cell fate program, directly repressing the SPDEF gene. Collectively, our study identifies YAP/TAZ as critical factors in lung epithelial homeostasis and offers molecular insight into the mechanisms promoting goblet cell differentiation, which is a hallmark of many lung diseases.

Effect of longevity genetic variants on the molecular aging rate.

We conducted a genome-wide association study of 1320 centenarians from the New England Centenarian Study (median age = 104 years) and 2899 unrelated controls using >9 M genetic variants imputed to the HRC panel of ~65,000 haplotypes. The genetic variants with the most significant associations were correlated to 4131 proteins that were profiled in the serum of a subset of 224 study participants using a SOMAscan array. The genetic associations were replicated in a genome-wide association study of 480 centenarians and ~800 controls of Ashkenazi Jewish descent. The proteomic associations were replicated in a proteomic scan of approximately 1000 Ashkenazi Jewish participants from a third cohort. The analysis replicated a protein signature associated with APOE genotypes and confirmed strong overexpression of BIRC2 (p < 5E-16) and under-expression of APOB in carriers of the APOE2 allele (p < 0.05). The analysis also discovered and replicated associations between longevity variants and slower changes of protein biomarkers of aging, including a novel protein signature of rs2184061 (CDKN2A/CDKN2B in chromosome 9) that suggests a genetic regulation of GDF15. The analyses showed that longevity variants correlate with proteome signatures that could be manipulated to discover healthy-aging targets.

animalcules: interactive microbiome analytics and visualization in R.

BACKGROUND: Microbial communities that live in and on the human body play a vital role in health and disease. Recent advances in sequencing technologies have enabled the study of microbial communities at unprecedented resolution. However, these advances in data generation have presented novel challenges to researchers attempting to analyze and visualize these data. RESULTS: To address some of these challenges, we have developed animalcules, an easy-to-use interactive microbiome analysis toolkit for 16S rRNA sequencing data, shotgun DNA metagenomics data, and RNA-based metatranscriptomics profiling data. This toolkit combines novel and existing analytics, visualization methods, and machine learning models. For example, the toolkit features traditional microbiome analyses such as alpha/beta diversity and differential abundance analysis, combined with new methods for biomarker identification are. In addition, animalcules provides interactive and dynamic figures that enable users to understand their data and discover new insights. animalcules can be used as a standalone command-line R package or users can explore their data with the accompanying interactive R Shiny interface. CONCLUSIONS: We present animalcules, an R package for interactive microbiome analysis through either an interactive interface facilitated by R Shiny or various command-line functions. It is the first microbiome analysis toolkit that supports the analysis of all 16S rRNA, DNA-based shotgun metagenomics, and RNA-sequencing based metatranscriptomics datasets. animalcules can be freely downloaded from GitHub at https://github.com/compbiomed/animalcules or installed through Bioconductor at https://www.bioconductor.org/packages/release/bioc/html/animalcules.html . Video abstract.

Contextualized Protein-Protein Interactions.

Protein-protein interaction (PPI) databases are an important bioinformatics resource, yet existing literature-curated databases usually represent cell-type-agnostic interactions, which is at variance with our understanding that protein dynamics are context specific and highly dependent on their environment. Here, we provide a resource derived through data mining to infer disease- and tissue-relevant interactions by annotating existing PPI databases with cell-contextual information extracted from reporting studies. This resource is applicable to the reconstruction and analysis of disease-centric molecular interaction networks. We have made the data and method publicly available and plan to release scheduled updates in the future. We expect these resources to be of interest to a wide audience of researchers in the life sciences.

Protein signatures of centenarians and their offspring suggest centenarians age slower than other humans.

Using samples from the New England Centenarian Study (NECS), we sought to characterize the serum proteome of 77 centenarians, 82 centenarians' offspring, and 65 age-matched controls of the offspring (mean ages: 105, 80, and 79 years). We identified 1312 proteins that significantly differ between centenarians and their offspring and controls (FDR < 1%), and two different protein signatures that predict longer survival in centenarians and in younger people. By comparing the centenarian signature with 2 independent proteomic studies of aging, we replicated the association of 484 proteins of aging and we identified two serum protein signatures that are specific of extreme old age. The data suggest that centenarians acquire similar aging signatures as seen in younger cohorts that have short survival periods, suggesting that they do not escape normal aging markers, but rather acquire them much later than usual. For example, centenarian signatures are significantly enriched for senescence-associated secretory phenotypes, consistent with those seen with younger aged individuals, and from this finding, we provide a new list of serum proteins that can be used to measure cellular senescence. Protein co-expression network analysis suggests that a small number of biological drivers may regulate aging and extreme longevity, and that changes in gene regulation may be important to reach extreme old age. This centenarian study thus provides additional signatures that can be used to measure aging and provides specific circulating biomarkers of healthy aging and longevity, suggesting potential mechanisms that could help prolong health and support longevity.

nf-gwas-pipeline: A Nextflow Genome-Wide Association Study Pipeline.

A tool for conducting Genome-Wide Association Study (GWAS) in a systematic, automated and reproducible manner is overdue. We developed an automated GWAS pipeline by combining multiple analysis tools - including bcftools, vcftools, the R packages SNPRelate/GENESIS/GMMAT and ANNOVAR - through Nextflow, which is a portable, flexible, and reproducible reactive workflow framework for developing pipelines. The GWAS pipeline integrates the steps of data quality control and assessment and genetic association analyses, including analysis of cross-sectional and longitudinal studies with either single variants or gene-based tests, into a unified analysis workflow. The pipeline is implemented in Nextflow, dependencies are distributed through Docker, and the code is publicly available on Github.

Naturally occurring hotspot cancer mutations in Gα13 promote oncogenic signaling.

Heterotrimeric G-proteins are signaling switches broadly divided into four families based on the sequence and functional similarity of their Gα subunits: Gs, Gi/o, Gq/11, and G12/13 Artificial mutations that activate Gα subunits of each of these families have long been known to induce oncogenic transformation in experimental systems. With the advent of next-generation sequencing, activating hotspot mutations in Gs, Gi/o, or Gq/11 proteins have also been identified in patient tumor samples. In contrast, patient tumor-associated G12/13 mutations characterized to date lead to inactivation rather than activation. By using bioinformatic pathway analysis and signaling assays, here we identified cancer-associated hotspot mutations in Arg-200 of Gα13 (encoded by GNA13) as potent activators of oncogenic signaling. First, we found that components of a G12/13-dependent signaling cascade that culminates in activation of the Hippo pathway effectors YAP and TAZ is frequently altered in bladder cancer. Up-regulation of this signaling cascade correlates with increased YAP/TAZ activation transcriptional signatures in this cancer type. Among the G12/13 pathway alterations were mutations in Arg-200 of Gα13, which we validated to promote YAP/TAZ-dependent (TEAD) and MRTF-A/B-dependent (SRE.L) transcriptional activity. We further showed that this mechanism relies on the same RhoGEF-RhoGTPase cascade components that are up-regulated in bladder cancers. Moreover, Gα13 Arg-200 mutants induced oncogenic transformation in vitro as determined by focus formation assays. In summary, our findings on Gα13 mutants establish that naturally occurring hotspot mutations in Gα subunits of any of the four families of heterotrimeric G-proteins are putative cancer drivers.

Gene therapy for Alzheimer's disease targeting CD33 reduces amyloid beta accumulation and neuroinflammation.

Neuroinflammation is a key contributor to the pathology of Alzheimer's disease (AD). CD33 (Siglec-3) is a transmembrane sialic acid-binding receptor on the surface of microglial cells. CD33 is upregulated on microglial cells from post-mortem AD patient brains, and high levels of CD33 inhibit uptake and clearance of amyloid beta (Aß) in microglial cell cultures. Furthermore, knockout of CD33 reduces amyloid plaque burden in mouse models of AD. Here, we tested whether a gene therapy strategy to reduce CD33 on microglia in AD could decrease Aß plaque load. Intracerebroventricular injection of an adeno-associated virus (AAV) vector-based system encoding an artificial microRNA targeting CD33 (miRCD33) into APP/PS1 mice reduced CD33 mRNA and TBS-soluble Aß40 and Aß42 levels in brain extracts. Treatment of APP/PS1 mice with miRCD33 vector at an early age (2 months) was more effective at reducing Aß plaque burden than intervening at later times (8 months). Furthermore, early intervention downregulated several microglial receptor transcripts (e.g. CD11c, CD47 and CD36) and pro-inflammatory activation genes (e.g. Tlr4 and Il1b). Marked reductions in the chemokine Ccl2 and the pro-inflammatory cytokine Tnfα were observed at the protein level in the brain of APP/PS1 mice treated with miRCD33 vector. Overall, our data indicate that CD33 is a viable target for AAV-based knockdown strategies to reduce AD pathology. One Sentence Summary: A gene therapy approach for Alzheimer's disease using adeno-associated virus vector-based knockdown of CD33 reduced amyloid beta accumulation and neuroinflammation.

FERONIA controls pectin- and nitric oxide-mediated male-female interaction.

Species that propagate by sexual reproduction actively guard against the fertilization of an egg by multiple sperm (polyspermy). Flowering plants rely on pollen tubes to transport their immotile sperm to fertilize the female gametophytes inside ovules. In Arabidopsis, pollen tubes are guided by cysteine-rich chemoattractants to target the female gametophyte1,2. The FERONIA receptor kinase has a dual role in ensuring sperm delivery and blocking polyspermy3. It has previously been reported that FERONIA generates a female gametophyte environment that is required for sperm release4. Here we show that FERONIA controls several functionally linked conditions to prevent the penetration of female gametophytes by multiple pollen tubes in Arabidopsis. We demonstrate that FERONIA is crucial for maintaining de-esterified pectin at the filiform apparatus, a region of the cell wall at the entrance to the female gametophyte. Pollen tube arrival at the ovule triggers the accumulation of nitric oxide at the filiform apparatus in a process that is dependent on FERONIA and mediated by de-esterified pectin. Nitric oxide nitrosates both precursor and mature forms of the chemoattractant LURE11, respectively blocking its secretion and interaction with its receptor, to suppress pollen tube attraction. Our results elucidate a mechanism controlled by FERONIA in which the arrival of the first pollen tube alters ovular conditions to disengage pollen tube attraction and prevent the approach and penetration of the female gametophyte by late-arriving pollen tubes, thus averting polyspermy.

Yap suppresses T-cell function and infiltration in the tumor microenvironment.

A major challenge for cancer immunotherapy is sustaining T-cell activation and recruitment in immunosuppressive solid tumors. Here, we report that the levels of the Hippo pathway effector Yes-associated protein (Yap) are sharply induced upon the activation of cluster of differentiation 4 (CD4)-positive and cluster of differentiation 8 (CD8)-positive T cells and that Yap functions as an immunosuppressive factor and inhibitor of effector differentiation. Loss of Yap in T cells results in enhanced T-cell activation, differentiation, and function, which translates in vivo to an improved ability for T cells to infiltrate and repress tumors. Gene expression analyses of tumor-infiltrating T cells following Yap deletion implicates Yap as a mediator of global T-cell responses in the tumor microenvironment and as a negative regulator of T-cell tumor infiltration and patient survival in diverse human cancers. Collectively, our results indicate that Yap plays critical roles in T-cell biology and suggest that Yap inhibition improves T-cell responses in cancer.

How the AHR Became Important in Cancer: The Role of Chronically Active AHR in Cancer Aggression.

For decades, the aryl hydrocarbon receptor (AHR) was studied for its role in environmental chemical toxicity i.e., as a quirk of nature and a mediator of unintended consequences of human pollution. During that period, it was not certain that the AHR had a "normal" physiological function. However, the ongoing accumulation of data from an ever-expanding variety of studies on cancer, cancer immunity, autoimmunity, organ development, and other areas bears witness to a staggering array of AHR-controlled normal and pathological activities. The objective of this review is to discuss how the AHR has gone from a likely contributor to genotoxic environmental carcinogen-induced cancer to a master regulator of malignant cell progression and cancer aggression. Particular focus is placed on the association between AHR activity and poor cancer outcomes, feedback loops that control chronic AHR activity in cancer, and the role of chronically active AHR in driving cancer cell invasion, migration, cancer stem cell characteristics, and survival.

hypeR: an R package for geneset enrichment workflows.

SUMMARY: Geneset enrichment is a popular method for annotating high-throughput sequencing data. Existing tools fall short in providing the flexibility to tackle the varied challenges researchers face in such analyses, particularly when analyzing many signatures across multiple experiments. We present a comprehensive R package for geneset enrichment workflows that offers multiple enrichment, visualization, and sharing methods in addition to novel features such as hierarchical geneset analysis and built-in markdown reporting. hypeR is a one-stop solution to performing geneset enrichment for a wide audience and range of use cases. AVAILABILITY AND IMPLEMENTATION: The most recent version of the package is available at https://github.com/montilab/hypeR.

Pipeliner: A Nextflow-Based Framework for the Definition of Sequencing Data Processing Pipelines.

The advent of high-throughput sequencing technologies has led to the need for flexible and user-friendly data preprocessing platforms. The Pipeliner framework provides an out-of-the-box solution for processing various types of sequencing data. It combines the Nextflow scripting language and Anaconda package manager to generate modular computational workflows. We have used Pipeliner to create several pipelines for sequencing data processing including bulk RNA-sequencing (RNA-seq), single-cell RNA-seq, as well as digital gene expression data. This report highlights the design methodology behind Pipeliner that enables the development of highly flexible and reproducible pipelines that are easy to extend and maintain on multiple computing environments. We also provide a quick start user guide demonstrating how to setup and execute available pipelines with toy datasets.

TREM2 Acts Downstream of CD33 in Modulating Microglial Pathology in Alzheimer's Disease.

The microglial receptors CD33 and TREM2 have been associated with risk for Alzheimer's disease (AD). Here, we investigated crosstalk between CD33 and TREM2. We showed that knockout of CD33 attenuated amyloid beta (Aß) pathology and improved cognition in 5xFAD mice, both of which were abrogated by additional TREM2 knockout. Knocking out TREM2 in 5xFAD mice exacerbated Aß pathology and neurodegeneration but reduced Iba1+ cell numbers, all of which could not be rescued by additional CD33 knockout. RNA-seq profiling of microglia revealed that genes related to phagocytosis and signaling (IL-6, IL-8, acute phase response) are upregulated in 5xFAD;CD33-/- and downregulated in 5xFAD;TREM2-/- mice. Differential gene expression in 5xFAD;CD33-/- microglia depended on the presence of TREM2, suggesting TREM2 acts downstream of CD33. Crosstalk between CD33 and TREM2 includes regulation of the IL-1ß/IL-1RN axis and a gene set in the "receptor activity chemokine" cluster. Our results should facilitate AD therapeutics targeting these receptors.

Nanoengineering of aggregation-free and thermally-stable gold nanoparticles in mesoporous frameworks.

Loading catalytically active, aggregation-free and thermally stable metal nanoparticles (NPs) on a high surface area support represents a major interest in heterogeneous catalysis. Current synthetic approaches to these hybrid catalysts, however, still lack controllability in the thermal stability of metal NPs, particularly at high temperatures in the absence of organic ligands. We herein report a facile "co-assembly" methodology to prepare aggregation-free, ligand-free and thermally stable mesoporous hybrid nanocatalysts of metal-oxides and metal-carbons. Immobilization of catalytically active gold NPs (AuNPs) within high surface area mesoporous frameworks was achieved via the polymer-directed co-assembly of chemically and structurally equivalent Pluronic P-123 and poly(ethylene oxide)-modified metallic gold NPs (AuNP-PEO) as co-structure-directing-agents. The in situ immobilization of AuNPs partially embedded into periodically ordered mesoporous frameworks imposed a three-dimensional "nanoconfinement" effect and essentially enhanced the long-term thermal stability of AuNPs up to 800 °C. The mesoporous hybrids retained a high surface accessibility of AuNPs and they had a fantastic high-temperature catalytic durability (>130 h at 375 °C) confirmed by two model catalytic reactions, including aerobic oxidation of benzyl alcohol and CO oxidation, respectively. Our results may offer a new realm of possibilities for the rational applications of thermally stable nanocatalysts in renewable energy technology and high-temperature catalysis.