MOTIVATION: Single-cell RNA sequencing (scRNA-seq) enables transcriptome-wide gene expression measurements at single-cell resolution providing a comprehensive view of the compositions and dynamics of tissue and organism development. The evolution of scRNA-seq protocols has led to a dramatic increase of cells throughput, exacerbating many of the computational and statistical issues that previously arose for bulk sequencing. In particular, with scRNA-seq data all the analyses steps, including normalization, have become computationally intensive, both in terms of memory usage and computational time. In this perspective, new accurate methods able to scale efficiently are desirable. RESULTS: Here, we propose PsiNorm, a between-sample normalization method based on the power-law Pareto distribution parameter estimate. Here, we show that the Pareto distribution well resembles scRNA-seq data, especially those coming from platforms that use unique molecular identifiers. Motivated by this result, we implement PsiNorm, a simple and highly scalable normalization method. We benchmark PsiNorm against seven other methods in terms of cluster identification, concordance and computational resources required. We demonstrate that PsiNorm is among the top performing methods showing a good trade-off between accuracy and scalability. Moreover, PsiNorm does not need a reference, a characteristic that makes it useful in supervised classification settings, in which new out-of-sample data need to be normalized. AVAILABILITY AND IMPLEMENTATION: PsiNorm is implemented in the scone Bioconductor package and available at https://bioconductor.org/packages/scone/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Genetic variations of expression quantitative trait loci (eQTLs) play a critical role in influencing complex traits and diseases development. Two main factors that affect the statistical power of detecting eQTLs are: (i) relatively small size of samples available, and (ii) heavy burden of multiple testing due to a very large number of variants to be tested. The later issue is particularly severe when one tries to identify trans-eQTLs that are far away from the genes they influence. If one can exploit co-expressed genes jointly in eQTL-mapping, effective sample size can be increased. Furthermore, using the structure of the gene regulatory network (GRN) may help to identify trans-eQTLs without increasing multiple testing burden. RESULTS: In this article, we use the structure equation model (SEM) to model both GRN and effect of eQTLs on gene expression, and then develop a novel algorithm, named sparse SEM for eQTL mapping (SSEMQ), to conduct joint eQTL mapping and GRN inference. The SEM can exploit co-expressed genes jointly in eQTL mapping and also use GRN to determine trans-eQTLs. Computer simulations demonstrate that our SSEMQ significantly outperforms nine existing eQTL mapping methods. SSEMQ is further used to analyze two real datasets of human breast and whole blood tissues, yielding a number of cis- and trans-eQTLs. AVAILABILITY AND IMPLEMENTATION: R package ssemQr is available at https://github.com/Ivis4ml/ssemQr.git. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Combining the results of different experiments to exhibit complex patterns or to improve statistical power is a typical aim of data integration. The starting point of the statistical analysis often comes as a set of P-values resulting from previous analyses, that need to be combined flexibly to explore complex hypotheses, while guaranteeing a low proportion of false discoveries. RESULTS: We introduce the generic concept of composed hypothesis, which corresponds to an arbitrary complex combination of simple hypotheses. We rephrase the problem of testing a composed hypothesis as a classification task and show that finding items for which the composed null hypothesis is rejected boils down to fitting a mixture model and classifying the items according to their posterior probabilities. We show that inference can be efficiently performed and provide a thorough classification rule to control for type I error. The performance and the usefulness of the approach are illustrated in simulations and on two different applications. The method is scalable, does not require any parameter tuning, and provided valuable biological insight on the considered application cases. AVAILABILITY AND IMPLEMENTATION: The QCH methodology is available in the qch package hosted on CRAN. Additionally, R codes to reproduce the Einkorn example are available on the personal webpage of the first author: https://www6.inrae.fr/mia-paris/Equipes/Membres/Tristan-Mary-Huard. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Simultaneous recordings of gene network dynamics across large populations have revealed that cell characteristics vary considerably even in clonal lines. Inferring the variability of parameters that determine gene dynamics is key to understanding cellular behavior. However, this is complicated by the fact that the outcomes and effects of many reactions are not observable directly. Unobserved reactions can be replaced with time delays to reduce model dimensionality and simplify inference. However, the resulting models are non-Markovian, and require the development of new inference techniques. RESULTS: We propose a non-Markovian, hierarchical Bayesian inference framework for quantifying the variability of cellular processes within and across cells in a population. We illustrate our approach using a delayed birth-death process. In general, a distributed delay model, rather than a popular fixed delay model, is needed for inference, even if only mean reaction delays are of interest. Using in silico and experimental data we show that the proposed hierarchical framework is robust and leads to improved estimates compared to its non-hierarchical counterpart. We apply our method to data obtained using time-lapse microscopy and infer the parameters that describe the dynamics of protein production at the single cell and population level. The mean delays in protein production are larger than previously reported, have a coefficient of variation of around 0.2 across the population, and are not strongly correlated with protein production or growth rates. AVAILABILITY AND IMPLEMENTATION: Accompanying code in Python is available at https://github.com/mvcortez/Bayesian-Inference. CONTACT: kresimir.josic@gmail.com or jaekkim@kaist.ac.kr or cbskust@korea.ac.kr. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Accurate automatic annotation of protein function relies on both innovative models and robust datasets. Due to their importance in biological processes, the identification of DNA-binding proteins directly from protein sequence has been the focus of many studies. However, the datasets used to train and evaluate these methods have suffered from substantial flaws. We describe some of the weaknesses of the datasets used in previous DNA-binding protein literature and provide several new datasets addressing these problems. We suggest new evaluative benchmark tasks that more realistically assess real-world performance for protein annotation models. We propose a simple new model for the prediction of DNA-binding proteins and compare its performance on the improved datasets to two previously published models. In addition, we provide extensive tests showing how the best models predict across taxa. RESULTS: Our new gradient boosting model, which uses features derived from a published protein language model, outperforms the earlier models. Perhaps surprisingly, so does a baseline nearest neighbor model using BLAST percent identity. We evaluate the sensitivity of these models to perturbations of DNA-binding regions and control regions of protein sequences. The successful data-driven models learn to focus on DNA-binding regions. When predicting across taxa, the best models are highly accurate across species in the same kingdom and can provide some information when predicting across kingdoms. AVAILABILITY AND IMPLEMENTATION: The data and results for this article can be found at https://doi.org/10.5281/zenodo.5153906. The code for this article can be found at https://doi.org/10.5281/zenodo.5153683. The code, data and results can also be found at https://github.com/AZaitzeff/tools_for_dna_binding_proteins.

MOTIVATION: Aberrant DNA methylation is strongly associated with heterogeneity in tumors. This study investigated the prognostic value of CpG island methylator phenotype in hepatocellular carcinoma (HCC). RESULTS: A total of 319 HCC samples with 21 121 CpG sites were included in this study and 215 disease-free survival (DFS) and overall survival (OS)-related CpG sites were identified. These CpG sites were divided into seven clusters by using consensus clustering method. Cluster 4, which constructed the prognostic prediction model as the seed cluster to evaluate survival risk for DFS and OS of HCC patients, had the lowest methylation level with the worse prognosis. The low-risk group patients had a significantly prolonged DFS and OS than the patients in the high-risk group (P = 0.008 and P < 0.001, respectively). A receiver operating characteristic curve results for predicting DFS and OS were 0.691 and 0.695, respectively. These results suggested that the CpG site methylation appears to be an informative prognostic biomarker in HCC. The CpG site methylation-related prognostic model may be an innovative insight to evaluate clinical outcomes for HCC patients. AVAILABILITY AND IMPLEMENTATION: The code of the analysis is available at https://www.bioconductor.org. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Classical Molecular Dynamics (MD) is a standard computational approach to model time-dependent processes at the atomic level. The inherent sparsity of increasingly huge generated trajectories demands clustering algorithms to reduce other post-simulation analysis complexity. The Quality Threshold (QT) variant is an appealing one from the vast number of available clustering methods. It guarantees that all members of a particular cluster will maintain a collective similarity established by a user-defined threshold. Unfortunately, its high computational cost for processing big data limits its application in the molecular simulation field. RESULTS: In this work, we propose a methodological parallel between QT clustering and another well-known algorithm in the field of Graph Theory, the Maximum Clique Problem. Molecular trajectories are represented as graphs whose nodes designate conformations, while unweighted edges indicate mutual similarity between nodes. The use of a binary-encoded RMSD matrix coupled to the exploitation of bitwise operations to extract clusters significantly contributes to reaching a very affordable algorithm compared to the few implementations of QT for MD available in the literature. Our alternative provides results in good agreement with the exact one while strictly preserving the collective similarity of clusters. AVAILABILITY AND IMPLEMENTATION: The source code and documentation of BitQT are free and publicly available on GitHub (https://github.com/LQCT/BitQT.git) and ReadTheDocs (https://bitqt.readthedocs.io/en/latest/), respectively. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

The increasing number of single cell and bulk RNAseq datasets describing complex gene expression profiles in different organisms, organs or cell types calls for an intuitive tool allowing rapid comparative analysis. Here, we present Swift Profiling Of Transcriptomes (SPOT) as a web tool that allows not only differential expression analysis but also fast ranking of genes fitting transcription profiles of interest. Based on a heuristic approach the spot algorithm ranks the genes according to their proximity to the user-defined gene expression profile of interest. The best hits are visualized as a table, bar chart or dot plot and can be exported as an Excel file. While the tool is generally applicable, we tested it on RNAseq data from malaria parasites that undergo multiple stage transformations during their complex life cycle as well as on data from multiple human organs during development and cell lines infected by SARS-CoV-2. SPOT should enable non-bioinformaticians to easily analyse their own and any available dataset. AVAILABILITY AND IMPLEMENTATION: SPOT is freely available for (academic) use at: https://frischknechtlab.shinyapps.io/SPOT/ and https://github.com/EliasFarr/SPOT. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

SUMMARY: Comparing results from multiple MD simulations performed under different conditions is essential during the initial stages of analysis. We propose a tool called MD Contact Comparison (MDContactCom) that compares residue-residue contact fluctuations of two MD trajectories, quantifies the differences, identifies sites that exhibit large differences and visualizes those sites on the protein structure. Using this method, it is possible to identify sites affected by varying simulation conditions and reveal the path of propagation of the effect even when differences between the 3D structure of the molecule and the fluctuation RMSF of each residue is unclear. MDContactCom can monitor differences in complex protein dynamics between two MD trajectories and identify candidate sites to be analyzed in more detail. As such, MDContactCom is a versatile software package for analyzing most MD simulations. AVAILABILITY AND IMPLEMENTATION: MDContactCom is freely available for download on GitLab. The software is implemented in Python3. https://gitlab.com/chiemotono/mdcontactcom. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Reference sequences are essential in creating a baseline of knowledge for many common bioinformatics methods, especially those using genomic sequencing. RESULTS: We have created refget, a Global Alliance for Genomics and Health API specification to access reference sequences and sub-sequences using an identifier derived from the sequence itself. We present four reference implementations across in-house and cloud infrastructure, a compliance suite and a web report used to ensure specification conformity across implementations. AVAILABILITY AND IMPLEMENTATION: The refget specification can be found at: https://w3id.org/ga4gh/refget. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

SUMMARY: PERMANOVA (permutational multivariate analysis of variance based on distances) has been widely used for testing the association between the microbiome and a covariate of interest. Statistical significance is established by permutation, which is computationally intensive for large sample sizes. As large-scale microbiome studies, such as American Gut Project (AGP), become increasingly popular, a computationally efficient version of PERMANOVA is much needed. To achieve this end, we derive the asymptotic distribution of the PERMANOVA pseudo-F statistic and provide analytical P-value calculation based on chi-square approximation. We show that the asymptotic P-value is close to the PERMANOVA P-value even under a moderate sample size. Moreover, it is more accurate and an order-of-magnitude faster than the permutation-free method MDMR. We demonstrated the use of our procedure D-MANOVA on the AGP dataset. AVAILABILITY AND IMPLEMENTATION: D-MANOVA is implemented by the dmanova function in the CRAN package GUniFrac. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Protein-protein interactions (PPI) play crucial roles in many biological processes, and identifying PPI sites is an important step for mechanistic understanding of diseases and design of novel drugs. Since experimental approaches for PPI site identification are expensive and time-consuming, many computational methods have been developed as screening tools. However, these methods are mostly based on neighbored features in sequence, and thus limited to capture spatial information. RESULTS: We propose a deep graph-based framework deep Graph convolutional network for Protein-Protein-Interacting Site prediction (GraphPPIS) for PPI site prediction, where the PPI site prediction problem was converted into a graph node classification task and solved by deep learning using the initial residual and identity mapping techniques. We showed that a deeper architecture (up to eight layers) allows significant performance improvement over other sequence-based and structure-based methods by more than 12.5% and 10.5% on AUPRC and MCC, respectively. Further analyses indicated that the predicted interacting sites by GraphPPIS are more spatially clustered and closer to the native ones even when false-positive predictions are made. The results highlight the importance of capturing spatially neighboring residues for interacting site prediction. AVAILABILITY AND IMPLEMENTATION: The datasets, the pre-computed features, and the source codes along with the pre-trained models of GraphPPIS are available at https://github.com/biomed-AI/GraphPPIS. The GraphPPIS web server is freely available at https://biomed.nscc-gz.cn/apps/GraphPPIS. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: The development of an open-source platform to predict protein 1D features and 3D structure is an important task. In this paper, we report an open-source toolkit for protein 3D structure modeling, named OPUS-X. It contains three modules: OPUS-TASS2, which predicts protein torsion angles, secondary structure and solvent accessibility; OPUS-Contact, which measures the distance and orientation information between different residue pairs; and OPUS-Fold2, which uses the constraints derived from the first two modules to guide folding. RESULTS: OPUS-TASS2 is an upgraded version of our previous method OPUS-TASS. OPUS-TASS2 integrates protein global structure information and significantly outperforms OPUS-TASS. OPUS-Contact combines multiple raw co-evolutionary features with protein 1D features predicted by OPUS-TASS2, and delivers better results than the open-source state-of-the-art method trRosetta. OPUS-Fold2 is a complementary version of our previous method OPUS-Fold. OPUS-Fold2 is a gradient-based protein folding framework based on the differentiable energy terms in opposed to OPUS-Fold that is a sampling-based method used to deal with the non-differentiable terms. OPUS-Fold2 exhibits comparable performance to the Rosetta folding protocol in trRosetta when using identical inputs. OPUS-Fold2 is written in Python and TensorFlow2.4, which is user-friendly to any source-code-level modification. AVAILABILITYAND IMPLEMENTATION: The code and pre-trained models of OPUS-X can be downloaded from https://github.com/OPUS-MaLab/opus_x. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: With the great progress of deep learning-based inter-residue contact/distance prediction, the discrete space formed by fragment assembly cannot satisfy the distance constraint well. Thus, the optimal solution of the continuous space may not be achieved. Designing an effective closed-loop continuous dihedral angle optimization strategy that complements the discrete fragment assembly is crucial to improve the performance of the distance-assisted fragment assembly method. RESULTS: In this article, we proposed a de novo protein structure prediction method called IPTDFold based on closed-loop iterative partition sampling, topology adjustment and residue-level distance deviation optimization. First, local dihedral angle crossover and mutation operators are designed to explore the conformational space extensively and achieve information exchange between the conformations in the population. Then, the dihedral angle rotation model of loop region with partial inter-residue distance constraints is constructed, and the rotation angle satisfying the constraints is obtained by differential evolution algorithm, so as to adjust the spatial position relationship between the secondary structures. Finally, the residue distance deviation is evaluated according to the difference between the conformation and the predicted distance, and the dihedral angle of the residue is optimized with biased probability. The final model is generated by iterating the above three steps. IPTDFold is tested on 462 benchmark proteins, 24 FM targets of CASP13 and 20 FM targets of CASP14. Results show that IPTDFold is significantly superior to the distance-assisted fragment assembly method Rosetta_D (Rosetta with distance). In particular, the prediction accuracy of IPTDFold does not decrease as the length of the protein increases. When using the same FastRelax protocol, the prediction accuracy of IPTDFold is significantly superior to that of trRosetta without orientation constraints, and is equivalent to that of the full version of trRosetta. AVAILABILITYAND IMPLEMENTATION: The source code and executable are freely available at https://github.com/iobio-zjut/IPTDFold. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Human proteins that are secreted into different body fluids from various cells and tissues can be promising disease indicators. Modern proteomics research empowered by both qualitative and quantitative profiling techniques has made great progress in protein discovery in various human fluids. However, due to the large number of proteins and diverse modifications present in the fluids, as well as the existing technical limits of major proteomics platforms (e.g. mass spectrometry), large discrepancies are often generated from different experimental studies. As a result, a comprehensive proteomics landscape across major human fluids are not well determined. RESULTS: To bridge this gap, we have developed a deep learning framework, named DeepSec, to identify secreted proteins in 12 types of human body fluids. DeepSec adopts an end-to-end sequence-based approach, where a Convolutional Neural Network is built to learn the abstract sequence features followed by a Bidirectional Gated Recurrent Unit with fully connected layer for protein classification. DeepSec has demonstrated promising performances with average area under the ROC curves of 0.85-0.94 on testing datasets in each type of fluids, which outperforms existing state-of-the-art methods available mostly on blood proteins. As an illustration of how to apply DeepSec in biomarker discovery research, we conducted a case study on kidney cancer by using genomics data from the cancer genome atlas and have identified 104 possible marker proteins. AVAILABILITY: DeepSec is available at https://bmbl.bmi.osumc.edu/deepsec/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: To facilitate the process of tailor-making a deep neural network for exploring the dynamics of genomic DNA, we have developed a hands-on package called ezGeno. ezGeno automates the search process of various parameters and network structures and can be applied to any kind of 1D genomic data. Combinations of multiple abovementioned 1D features are also applicable. RESULTS: For the task of predicting TF binding using genomic sequences as the input, ezGeno can consistently return the best performing set of parameters and network structure, as well as highlight the important segments within the original sequences. For the task of predicting tissue-specific enhancer activity using both sequence and DNase feature data as the input, ezGeno also regularly outperforms the hand-designed models. Furthermore, we demonstrate that ezGeno is superior in efficiency and accuracy compared to the one-layer DeepBind model and AutoKeras, an open-source AutoML package. AVAILABILITY AND IMPLEMENTATION: The ezGeno package can be freely accessed at https://github.com/ailabstw/ezGeno. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: The well-known fact that protein structures are more conserved than their sequences forms the basis of several areas of computational structural biology. Methods based on the structure analysis provide more complete information on residue conservation in evolutionary processes. This is crucial for the determination of evolutionary relationships between proteins and for the identification of recurrent structural patterns present in biomolecules involved in similar functions. However, algorithmic structural alignment is much more difficult than multiple sequence alignment. This study is devoted to the development and applications of DAMA-a novel effective environment capable to compute and analyze multiple structure alignments. RESULTS: DAMA is based on local structural similarities, using local 3D structure descriptors and thus accounts for nearest-neighbor molecular environments of aligned residues. It is constrained neither by protein topology nor by its global structure. DAMA is an extension of our previous study (DEDAL) which demonstrated the applicability of local descriptors to pairwise alignment problems. Since the multiple alignment problem is NP-complete, an effective heuristic approach has been developed without imposing any artificial constraints. The alignment algorithm searches for the largest, consistent ensemble of similar descriptors. The new method is capable to capture most of the biologically significant similarities present in canonical test sets and is discriminatory enough to prevent the emergence of larger, but meaningless, solutions. Tests performed on the test sets, including protein kinases, demonstrate DAMA's capability of identifying equivalent residues, which should be very useful in discovering the biological nature of proteins similarity. Performance profiles show the advantage of DAMA over other methods, in particular when using a strict similarity measure QC, which is the ratio of correctly aligned columns, and when applying the methods to more difficult cases. AVAILABILITY AND IMPLEMENTATION: DAMA is available online at http://dworkowa.imdik.pan.pl/EP/DAMA. Linux binaries of the software are available upon request. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Motions of transmembrane receptors on cancer cell surfaces can reveal biophysical features of the cancer cells, thus providing a method for characterizing cancer cell phenotypes. While conventional analysis of receptor motions in the cell membrane mostly relies on the mean-squared displacement plots, much information is lost when producing these plots from the trajectories. Here we employ deep learning to classify breast cancer cell types based on the trajectories of epidermal growth factor receptor (EGFR). Our model is an artificial neural network trained on the EGFR motions acquired from six breast cancer cell lines of varying invasiveness and receptor status: MCF7 (hormone receptor positive), BT474 (HER2-positive), SKBR3 (HER2-positive), MDA-MB-468 (triple negative, TN), MDA-MB-231 (TN) and BT549 (TN). RESULTS: The model successfully classified the trajectories within individual cell lines with 83% accuracy and predicted receptor status with 85% accuracy. To further validate the method, epithelial-mesenchymal transition (EMT) was induced in benign MCF10A cells, noninvasive MCF7 cancer cells and highly invasive MDA-MB-231 cancer cells, and EGFR trajectories from these cells were tested. As expected, after EMT induction, both MCF10A and MCF7 cells showed higher rates of classification as TN cells, but not the MDA-MB-231 cells. Whereas deep learning-based cancer cell classifications are primarily based on the optical transmission images of cell morphology and the fluorescence images of cell organelles or cytoskeletal structures, here we demonstrated an alternative way to classify cancer cells using a dynamic, biophysical feature that is readily accessible. AVAILABILITY AND IMPLEMENTATION: A python implementation of deep learning-based classification can be found at https://github.com/soonwoohong/Deep-learning-for-EGFR-trajectory-classification. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Co-evolution analysis can be used to accurately predict residue-residue contacts from multiple sequence alignments. The introduction of machine-learning techniques has enabled substantial improvements in precision and a shift from predicting binary contacts to predict distances between pairs of residues. These developments have significantly improved the accuracy of de novo prediction of static protein structures. With AlphaFold2 lifting the accuracy of some predicted protein models close to experimental levels, structure prediction research will move on to other challenges. One of those areas is the prediction of more than one conformation of a protein. Here, we examine the potential of residue-residue distance predictions to be informative of protein flexibility rather than simply static structure. RESULTS: We used DMPfold to predict distance distributions for every residue pair in a set of proteins that showed both rigid and flexible behaviour. Residue pairs that were in contact in at least one reference structure were classified as rigid, flexible or neither. The predicted distance distribution of each residue pair was analysed for local maxima of probability indicating the most likely distance or distances between a pair of residues. We found that rigid residue pairs tended to have only a single local maximum in their predicted distance distributions while flexible residue pairs more often had multiple local maxima. These results suggest that the shape of predicted distance distributions contains information on the rigidity or flexibility of a protein and its constituent residues. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Scalable clustering algorithms are needed to analyze millions of cells in single cell RNA-seq (scRNA-seq) data. RESULTS: Here, we present an open source python package called FlowGrid that can integrate into the Scanpy workflow to perform clustering on very large scRNA-seq datasets. FlowGrid implements a fast density-based clustering algorithm originally designed for flow cytometry data analysis. We introduce a new automated parameter tuning procedure, and show that FlowGrid can achieve comparable clustering accuracy as state-of-the-art clustering algorithms but at a substantially reduced run time for very large single cell RNA-seq datasets. For example, FlowGrid can complete a one-hour clustering task for one million cells in about five min. AVAILABILITY AND IMPLEMENTATION: https://github.com/holab-hku/FlowGrid. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Multi-omics data in molecular biology has accumulated rapidly over the years. Such data contains valuable information for research in medicine and drug discovery. Unfortunately, data-driven research in medicine and drug discovery is challenging for a majority of small research labs due to the large volume of data and the complexity of analysis pipeline. RESULTS: We present BioVLAB-Cancer-Pharmacogenomics, a bioinformatics system that facilitates analysis of multi-omics data from breast cancer to analyze and investigate intratumor heterogeneity and pharmacogenomics on Amazon Web Services. Our system takes multi-omics data as input to perform tumor heterogeneity analysis in terms of TCGA data and deconvolve-and-match the tumor gene expression to cell line data in CCLE using DNA methylation profiles. We believe that our system can help small research labs perform analysis of tumor multi-omics without worrying about computational infrastructure and maintenance of databases and tools. AVAILABILITY AND IMPLEMENTATION: http://biohealth.snu.ac.kr/software/biovlab_cancer_pharmacogenomics. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: High-throughput experiments are generating ever increasing amounts of various -omics data, so shedding new light on the link between human disorders, their genetic causes and the related impact on protein behavior and structure. While numerous bioinformatics tools now exist that predict which variants in the human exome cause diseases, few tools predict the reasons why they might do so. Yet, understanding the impact of variants at the molecular level is a prerequisite for the rational development of targeted drugs or personalized therapies. RESULTS: We present the updated MutaFrame webserver, which aims to meet this need. It offers two deleteriousness prediction softwares, DEOGEN2 and SNPMuSiC, and is designed for bioinformaticians and medical researchers who want to gain insights into the origins of monogenic diseases. It contains information at two levels for each human protein: its amino acid sequence and its three-dimensional structure; we used the experimental structures whenever available, and modeled structures otherwise. MutaFrame also includes higher-level information, such as protein essentiality and protein-protein interactions. It has a user-friendly interface for the interpretation of results and a convenient visualization system for protein structures, in which the variant positions introduced by the user and other structural information are shown. In this way, MutaFrame aids our understanding of the pathogenic processes caused by single-site mutations and their molecular and contextual interpretation. AVAILABILITY AND IMPLEMENTATION: Mutaframe webserver at http://mutaframe.com/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Intrinsically disordered protein regions interact with proteins, nucleic acids and lipids. Regions that bind lipids are implicated in a wide spectrum of cellular functions and several human diseases. Motivated by the growing amount of experimental data for these interactions and lack of tools that can predict them from the protein sequence, we develop DisoLipPred, the first predictor of the disordered lipid-binding residues (DLBRs). RESULTS: DisoLipPred relies on a deep bidirectional recurrent network that implements three innovative features: transfer learning, bypass module that sidesteps predictions for putative structured residues, and expanded inputs that cover physiochemical properties associated with the protein-lipid interactions. Ablation analysis shows that these features drive predictive quality of DisoLipPred. Tests on an independent test dataset and the yeast proteome reveal that DisoLipPred generates accurate results and that none of the related existing tools can be used to indirectly identify DLBR. We also show that DisoLipPred's predictions complement the results generated by predictors of the transmembrane regions. Altogether, we conclude that DisoLipPred provides high-quality predictions of DLBRs that complement the currently available methods. AVAILABILITY AND IMPLEMENTATION: DisoLipPred's webserver is available at http://biomine.cs.vcu.edu/servers/DisoLipPred/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: The solvent accessible surface is an essential structural property measure related to the protein structure and protein function. Relative solvent accessible area (RSA) is a standard measure to describe the degree of residue exposure in the protein surface or inside of protein. However, this computation will fail when the residues information is missing. RESULTS: In this article, we proposed a novel method for estimation RSA using the Cα atom distance matrix with the deep learning method (EAGERER). The new method, EAGERER, achieves Pearson correlation coefficients of 0.921-0.928 on two independent test datasets. We empirically demonstrate that EAGERER can yield better Pearson correlation coefficients than existing RSA estimators, such as coordination number, half sphere exposure and SphereCon. To the best of our knowledge, EAGERER represents the first method to estimate the solvent accessible area using limited information with a deep learning model. It could be useful to the protein structure and protein function prediction. AVAILABILITYAND IMPLEMENTATION: The method is free available at https://github.com/cliffgao/EAGERER. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: The evolutionary processes of mutation and recombination, upon which selection operates, are fundamental to understand the observed molecular diversity. Unlike nucleotide sequences, the estimation of the recombination rate in protein sequences has been little explored, neither implemented in evolutionary frameworks, despite protein sequencing methods are largely used. RESULTS: In order to accommodate this need, here I present a computational framework, called ProteinEvolverABC, to jointly estimate recombination and substitution rates from alignments of protein sequences. The framework implements the approximate Bayesian computation approach, with and without regression adjustments and includes a variety of substitution models of protein evolution, demographics and longitudinal sampling. It also implements several nuisance parameters such as heterogeneous amino acid frequencies and rate of change among sites and, proportion of invariable sites. The framework produces accurate coestimation of recombination and substitution rates under diverse evolutionary scenarios. As illustrative examples of usage, I applied it to several viral protein families, including coronaviruses, showing heterogeneous substitution and recombination rates. AVAILABILITY AND IMPLEMENTATION: ProteinEvolverABC is freely available from https://github.com/miguelarenas/proteinevolverabc, includes a graphical user interface for helping the specification of the input settings, extensive documentation and ready-to-use examples. Conveniently, the simulations can run in parallel on multicore machines. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Accurate disease phenotype prediction plays an important role in the treatment of heterogeneous diseases like cancer in the era of precision medicine. With the advent of high throughput technologies, more comprehensive multi-omics data is now available that can effectively link the genotype to phenotype. However, the interactive relation of multi-omics datasets makes it particularly challenging to incorporate different biological layers to discover the coherent biological signatures and predict phenotypic outcomes. In this study, we introduce omicsGAN, a generative adversarial network model to integrate two omics data and their interaction network. The model captures information from the interaction network as well as the two omics datasets and fuse them to generate synthetic data with better predictive signals. RESULTS: Large-scale experiments on The Cancer Genome Atlas breast cancer, lung cancer and ovarian cancer datasets validate that (i) the model can effectively integrate two omics data (e.g. mRNA and microRNA expression data) and their interaction network (e.g. microRNA-mRNA interaction network). The synthetic omics data generated by the proposed model has a better performance on cancer outcome classification and patients survival prediction compared to original omics datasets. (ii) The integrity of the interaction network plays a vital role in the generation of synthetic data with higher predictive quality. Using a random interaction network does not allow the framework to learn meaningful information from the omics datasets; therefore, results in synthetic data with weaker predictive signals. AVAILABILITY AND IMPLEMENTATION: Source code is available at: https://github.com/CompbioLabUCF/omicsGAN. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: The COVID-19 pandemic has ignited a broad scientific interest in viral research in general and coronavirus research in particular. The identification and characterization of viral species in natural reservoirs typically involves de novo assembly. However, existing genome, metagenome and transcriptome assemblers often are not able to assemble many viruses (including coronaviruses) into a single contig. Coverage variation between datasets and within dataset, presence of close strains, splice variants and contamination set a high bar for assemblers to process viral datasets with diverse properties. RESULTS: We developed coronaSPAdes, a novel assembler for RNA viral species recovery in general and coronaviruses in particular. coronaSPAdes leverages the knowledge about viral genome structures to improve assembly extending ideas initially implemented in biosyntheticSPAdes. We have shown that coronaSPAdes outperforms existing SPAdes modes and other popular short-read metagenome and viral assemblers in the recovery of full-length RNA viral genomes. AVAILABILITY AND IMPLEMENTATION: coronaSPAdes version used in this article is a part of SPAdes 3.15 release and is freely available at http://cab.spbu.ru/software/spades. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Information about bacteria biotopes (BB) is important for fundamental research and applications in microbiology. BB task at BioNLP-OST 2019 focuses on the extraction of locations and phenotypes of microorganisms from PubMed abstracts and full-text excerpts. The subtask BB-rel+ner aims to recognize relevant entities and extract interrelationships about BBs. The corresponding corpus owns some distinctive features (e.g. nested entities) which are challenging to deal with. Therefore, previous methods achieved low performance on entity and relation extraction and limited the mutual effect between named entity recognition and relation extraction. There is still much room for improvement. RESULTS: We propose a span-based model to extract entities and relations jointly from biomedical text regarding the BBs. For alleviating the problem of annotated data deficiency in domain-specific task, we employ a BERT (Bidirectional Encoder Representations from Transformers) model pre-trained on the domain-specific corpus to encode sentences. Our model considers all spans in a sentence as potential entity mentions and computes relation scores between the most confident entity spans based on representations of spans and contexts between spans. Experiments on the BB-rel+ner 2019 corpus demonstrate that our model achieves significantly better performance than the state-of-the-art method, with a reduction of 21.6% slot error rate (SER) for extracting relations. Our model is also effective in recognizing nested entities. Furthermore, the model can be applied to the CHEMPROT corpus for joint extraction of chemical-protein entities and relations, achieving state-of-the-art performance. AVAILABILITY AND IMPLEMENTATION: Our source code is available at https://github.com/zmmzGitHub/SpanMB_BERT. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Single-cell multi-omics sequencing data can provide a comprehensive molecular view of cells. However, effective approaches for the integrative analysis of such data are challenging. Existing manifold alignment methods demonstrated the state-of-the-art performance on single-cell multi-omics data integration, but they are often limited by requiring that single-cell datasets be derived from the same underlying cellular structure. RESULTS: In this study, we present Pamona, a partial Gromov-Wasserstein distance-based manifold alignment framework that integrates heterogeneous single-cell multi-omics datasets with the aim of delineating and representing the shared and dataset-specific cellular structures across modalities. We formulate this task as a partial manifold alignment problem and develop a partial Gromov-Wasserstein optimal transport framework to solve it. Pamona identifies both shared and dataset-specific cells based on the computed probabilistic couplings of cells across datasets, and it aligns cellular modalities in a common low-dimensional space, while simultaneously preserving both shared and dataset-specific structures. Our framework can easily incorporate prior information, such as cell type annotations or cell-cell correspondence, to further improve alignment quality. We evaluated Pamona on a comprehensive set of publicly available benchmark datasets. We demonstrated that Pamona can accurately identify shared and dataset-specific cells, as well as faithfully recover and align cellular structures of heterogeneous single-cell modalities in a common space, outperforming the comparable existing methods. AVAILABILITYAND IMPLEMENTATION: Pamona software is available at https://github.com/caokai1073/Pamona. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: An unsolved fundamental problem in biology is to predict phenotypes from a new genotype under environmental perturbations. The emergence of multiple omics data provides new opportunities but imposes great challenges in the predictive modeling of genotype-phenotype associations. Firstly, the high-dimensionality of genomics data and the lack of coherent labeled data often make the existing supervised learning techniques less successful. Secondly, it is challenging to integrate heterogeneous omics data from different resources. Finally, few works have explicitly modeled the information transmission from DNA to phenotype, which involves multiple intermediate molecular types. Higher-level features (e.g. gene expression) usually have stronger discriminative and interpretable power than lower-level features (e.g. somatic mutation). RESULTS: We propose a novel Cross-LEvel Information Transmission (CLEIT) network framework to address the above issues. CLEIT aims to represent the asymmetrical multi-level organization of the biological system by integrating multiple incoherent omics data and to improve the prediction power of low-level features. CLEIT first learns the latent representation of the high-level domain then uses it as ground-truth embedding to improve the representation learning of the low-level domain in the form of contrastive loss. Besides, CLEIT can leverage the unlabeled heterogeneous omics data to improve the generalizability of the predictive model. We demonstrate the effectiveness and significant performance boost of CLEIT in predicting anti-cancer drug sensitivity from somatic mutations via the assistance of gene expressions when compared with state-of-the-art methods. CLEIT provides a general framework to model information transmissions and integrate multi-modal data in a multi-level system. AVAILABILITYAND IMPLEMENTATION: The source code is freely available at https://github.com/XieResearchGroup/CLEIT. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: The simultaneous availability of ATAC-seq and RNA-seq experiments allows to obtain a more in-depth knowledge on the regulatory mechanisms occurring in gene regulatory networks. In this article, we highlight and analyze two novel aspects that leverage on the possibility of pairing RNA-seq and ATAC-seq data. Namely we investigate the causality of the relationships between transcription factors, chromatin and target genes and the internal consistency between the two omics, here measured in terms of structural balance in the sample correlations along elementary length-3 cycles. RESULTS: We propose a framework that uses the a priori knowledge on the data to infer elementary causal regulatory motifs (namely chains and forks) in the network. It is based on the notions of conditional independence and partial correlation, and can be applied to both longitudinal and non-longitudinal data. Our analysis highlights a strong connection between the causal regulatory motifs that are selected by the data and the structural balance of the underlying sample correlation graphs: strikingly, >97% of the selected regulatory motifs belong to a balanced subgraph. This result shows that internal consistency, as measured by structural balance, is close to a necessary condition for 3-node regulatory motifs to satisfy causality rules. AVAILABILITY AND IMPLEMENTATION: The analysis was carried out in MATLAB and the code can be found at https://github.com/albertozenere/Multi-omics-elementary-regulatory-motifs. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: DNA and RNA modifications can now be identified using nanopore sequencing. However, we currently lack a flexible software to efficiently encode, store, analyze and visualize DNA and RNA modification data. RESULTS: Here, we present ModPhred, a versatile toolkit that facilitates DNA and RNA modification analysis from nanopore sequencing reads in a user-friendly manner. ModPhred integrates probabilistic DNA and RNA modification information within the FASTQ and BAM file formats, can be used to encode multiple types of modifications simultaneously, and its output can be easily coupled to genomic track viewers, facilitating the visualization and analysis of DNA and RNA modification information in individual reads in a simple and computationally efficient manner. AVAILABILITY AND IMPLEMENTATION: ModPhred is available at https://github.com/novoalab/modPhred, is implemented in Python3, and is released under an MIT license. Docker images with all dependencies preinstalled are also provided. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Smoldyn is a particle-based biochemical simulator that is frequently used for systems biology and biophysics research. Previously, users could only define models using text-based input or a C/C++ application programming interface (API), which were convenient, but limited extensibility. RESULTS: We added a Python API to Smoldyn to improve integration with other software tools, such as Jupyter notebooks, other Python code libraries and other simulators. It includes low-level functions that closely mimic the existing C/C++ API and higher-level functions that are more convenient to use. These latter functions follow modern object-oriented Python conventions. AVAILABILITY AND IMPLEMENTATION: Smoldyn is open source and free, available at http://www.smoldyn.org and can be installed with the Python package manager pip. It runs on Mac, Windows and Linux. Documentation is available at http://www.smoldyn.org/SmoldynManual.pdf and https://smoldyn.readthedocs.io/en/latest/python/api.html.

MOTIVATION: A few algorithms have been developed for splitting the genome in nearly independent blocks of linkage disequilibrium. Due to the complexity of this problem, these algorithms rely on heuristics, which makes them suboptimal. RESULTS: Here, we develop an optimal solution for this problem using dynamic programming. AVAILABILITY: This is now implemented as function snp_ldsplit as part of R package bigsnpr. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Profiling the taxonomic composition of microbial communities commonly involves the classification of ribosomal RNA gene fragments. As a trade-off to maintain high classification accuracy, existing tools are typically limited to the genus level. Here, we present mTAGs, a taxonomic profiling tool that implements the alignment of metagenomic sequencing reads to degenerate consensus reference sequences of small subunit ribosomal RNA genes. It uses DNA fragments, that is, paired-end sequencing reads, as count units and provides relative abundance profiles at multiple taxonomic ranks, including operational taxonomic units based on a 97% sequence identity cutoff. At the genus rank, mTAGs outperformed other tools across several metrics, such as the F1 score by >11% across data from different environments, and achieved competitive (F1 score) or better results (Bray-Curtis dissimilarity) at the sub-genus level. AVAILABILITY AND IMPLEMENTATION: The software tool mTAGs is implemented in Python. The source code and binaries are freely available (https://github.com/SushiLab/mTAGs). The data underlying this article are available in Zenodo, at https://doi.org/10.5281/zenodo.4352762. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

SUMMARY: pKa values of ionizable residues and isoelectric points of proteins provide valuable local and global insights about their structure and function. These properties can be estimated with reasonably good accuracy using Poisson-Boltzmann and Monte Carlo calculations at a considerable computational cost (from some minutes to several hours). pKPDB is a database of over 12 M theoretical pKa values calculated over 120k protein structures deposited in the Protein Data Bank. By providing precomputed pKa and pI values, users can retrieve results instantaneously for their protein(s) of interest while also saving countless hours and resources that would be spent on repeated calculations. Furthermore, there is an ever-growing imbalance between experimental pKa and pI values and the number of resolved structures. This database will complement the experimental and computational data already available and can also provide crucial information regarding buried residues that are under-represented in experimental measurements. AVAILABILITY AND IMPLEMENTATION: Gzipped csv files containing p Ka and isoelectric point values can be downloaded from https://pypka.org/pKPDB. To query a single PDB code please use the PypKa free server at https://pypka.org. The pKPDB source code can be found at https://github.com/mms-fcul/pKPDB. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

MOTIVATION: Previously we presented swarm, an open-source amplicon clustering programme that produces fine-scale molecular operational taxonomic units (OTUs) that are free of arbitrary global clustering thresholds. Here, we present swarm v3 to address issues of contemporary datasets that are growing towards tera-byte sizes. RESULTS: When compared with previous swarm versions, swarm v3 has modernized C++ source code, reduced memory footprint by up to 50%, optimized CPU-usage and multithreading (more than 7 times faster with default parameters), and it has been extensively tested for its robustness and logic. AVAILABILITY AND IMPLEMENTATION: Source code and binaries are available at https://github.com/torognes/swarm. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

SUMMARY: MomentClosure.jl is a Julia package providing automated derivation of the time-evolution equations of the moments of molecule numbers for virtually any chemical reaction network using a wide range of moment closure approximations. It extends the capabilities of modelling stochastic biochemical systems in Julia and can be particularly useful when exact analytic solutions of the chemical master equation are unavailable and when Monte Carlo simulations are computationally expensive. AVAILABILITY AND IMPLEMENTATION: MomentClosure.jl is freely accessible under the MIT licence. Source code and documentation are available at https://github.com/augustinas1/MomentClosure.jl.