Identifying gene expression programs in single-cell RNA-seq data using linear correlation explanation

Single-cell RNA-sequencing (scRNA-seq) has emerged as a powerful technique for studying gene expression patterns at the single-cell level. Inferring gene regulatory networks (GRNs) from scRNA-seq data provides insight into cellular phenotypes from the genomic level. However, the high sparsity, noise and dropout events inherent in scRNA-seq data present challenges for GRN inference. In recent years, the dramatic increase in data on experimentally validated transcription factors binding to DNA has made it possible to infer GRNs by supervised methods. In this study, we address the problem of GRN inference by framing it as a graph link prediction task. In this paper, we propose a novel framework called GNNLink, which leverages known GRNs to deduce the potential regulatory interdependencies between genes. First, we preprocess the raw scRNA-seq data. Then, we introduce a graph convolutional network-based interaction graph encoder to effectively refine gene features by capturing interdependencies between nodes in the network. Finally, the inference of GRN is obtained by performing matrix completion operation on node features. The features obtained from model training can be applied to downstream tasks such as measuring similarity and inferring causality between gene pairs. To evaluate the performance of GNNLink, we compare it with six existing GRN reconstruction methods using seven scRNA-seq datasets. These datasets encompass diverse ground truth networks, including functional interaction networks, Loss of Function/Gain of Function data, non-specific ChIP-seq data and cell-type-specific ChIP-seq data. Our experimental results demonstrate that GNNLink achieves comparable or superior performance across these datasets, showcasing its robustness and accuracy. Furthermore, we observe consistent performance across datasets of varying scales. For reproducibility, we provide the data and source code of GNNLink on our GitHub repository: https://github.com/sdesignates/GNNLink.

In a single-cell RNA-seq (scRNA-seq) data set, a high proportion of missing values (or an excessive number of zeroes) are frequently observed. For the related follow-up tasks, such as clustering analysis and differential expression analysis, a data set without missing values is generally required. Many imputation approaches have been proposed for this purpose. Multiple imputation (MI) is a well-established approach to address possible biases in a follow-up analysis result based on one-time imputed data. There is a lack of investigation on this in the analysis of scRNA-seq data. In this study, we have investigated how to efficiently apply the MI approach to the clustering analysis and the differential expression analysis of scRNA-seq data. We proposed an MI procedure for clustering analysis and an MI procedure for differential expression analysis. To demonstrate the improvements achieved by MI in clustering analysis and differential expression analysis of scRNA-seq data, we analyzed three well-known scRNA-seq data sets. scIGANs, an scRNA-seq imputation method based on the generative adversarial networks (GANs), has been recently proposed for scRNA-seq data imputation. Multiple randomly imputed data sets can be conveniently generated by this method. We implemented our MI procedures based on scIGANs. We demonstrated that MI yielded improved performances on the clustering analysis and differential expression analysis results. Our applications to experimental scRNA-seq data illustrated the advantages of MI over one-time imputation of missing values in scRNA-seq data.

The development and widespread utilization of high-throughput sequencing technologies in biology has fueled the rapid growth of single-cell RNA sequencing (scRNA-seq) data over the past decade. The development of scRNA-seq technology has significantly expanded researchers’ understanding of cellular heterogeneity. Accurate cell type identification is the prerequisite for any research on heterogeneous cell populations. However, due to the high noise and high dimensionality of scRNA-seq data, improving the effectiveness of cell type identification remains a challenge. As an effective dimensionality reduction method, Principal Component Analysis (PCA) is an essential tool for visualizing high-dimensional scRNA-seq data and identifying cell subpopulations. However, traditional PCA has some defects when used in mining the nonlinear manifold structure of the data and usually suffers from over-density of principal components (PCs). Therefore, we present a novel method in this paper called joint -norm and random walk graph constrained PCA (RWPPCA). RWPPCA aims to retain the data’s local information in the process of mapping high-dimensional data to low-dimensional space, to more accurately obtain sparse principal components and to then identify cell types more precisely. Specifically, RWPPCA combines the random walk (RW) algorithm with graph regularization to more accurately determine the local geometric relationships between data points. Moreover, to mitigate the adverse effects of dense PCs, the -norm is introduced to make the PCs sparser, thus increasing their interpretability. Then, we evaluate the effectiveness of RWPPCA on simulated data and scRNA-seq data. The results show that RWPPCA performs well in cell type identification and outperforms other comparison methods.

Single-cell RNA sequencing (scRNA-seq) has recently brought new insight into cell differentiation processes and functional variation in cell subtypes from homogeneous cell populations. A lack of prior knowledge makes unsupervised machine learning methods, such as clustering, suitable for analyzing scRNA-seq. However, there are several limitations to overcome, including high dimensionality, clustering result instability, and parameter adjustment complexity. In this study, we propose a method by combining structure entropy and k nearest neighbor to identify cell subpopulations in scRNA-seq data. In contrast to existing clustering methods for identifying cell subtypes, minimized structure entropy results in natural communities without specifying the number of clusters. To investigate the performance of our model, we applied it to eight scRNA-seq datasets and compared our method with three existing methods (nonnegative matrix factorization, single-cell interpretation via multikernel learning, and structural entropy minimization principle). The experimental results showed that our approach achieves, on average, better performance in these datasets compared to the benchmark methods.

Single-cell RNA sequencing technologies have been pivotal in advancing the development of algorithms for clustering heterogeneous cell populations. Existing methods for utilizing scRNA-seq data to identify cell types tend to neglect the beneficial impact of dropout events and perform clustering focusing solely on quantitative perspective. Here, we introduce a novel method named scQA, notable for its ability to concurrently identify cell types and cell type-specific key genes from both qualitative and quantitative perspectives. In contrast to other methods, scQA not only identifies cell types but also extracts key genes associated with these cell types, enabling bidirectional clustering for scRNA-seq data. Through an iterative process, our approach aims to minimize the number of landmarks to approximately a dozen while maximizing the inclusion of quasi-trend-preserved genes with dropouts both qualitatively and quantitatively. It then clusters cells by employing an ingenious label propagation strategy, obviating the requirement for a predetermined number of cell types. Validated on 20 publicly available scRNA-seq datasets, scQA consistently outperforms other salient tools. Furthermore, we confirm the effectiveness and potential biological significance of the identified key genes through both external and internal validation. In conclusion, scQA emerges as a valuable tool for investigating cell heterogeneity due to its distinctive fusion of qualitative and quantitative facets, along with bidirectional clustering capabilities. Furthermore, it can be seamlessly integrated into border scRNA-seq analyses. The source codes are publicly available at https://github.com/LD-Lyndee/scQA.

TransAnno-Net: A Deep Learning Framework for Accurate Cell Type Annotation of Mouse Lung Tissue Using Self-supervised Pretraining.

MLSpatial: A machine-learning method to reconstruct the spatial distribution of cells from scRNA-seq by extracting spatial features

En1 and Lmx1b do not recapitulate embryonic dorsal-ventral limb patterning functions during mouse digit tip regeneration.

The incidence and mortality rates of lung cancer are exceptionally high. Many patients are diagnosed with early stage lung cancer but experience rapid recurrence post-surgery. Many research studies have shown that the unfavorable prognosis of patients may be associated with micro-metastasis in the lymph nodes. Our research aimed to develop a nomogram to predict the prognosis of lung adenocarcinoma (LUAD). Single-cell RNA sequencing (scRNA-seq) data were analyzed to identify 11 cell clusters. Patterns of incoming and outgoing signals were identified across the entire cell population. A weighted gene co-expression network analysis (WGCNA) was conducted to uncover critical genes in LUAD. The intersecting marker genes were used to construct the prognostic model. scRNA-seq data were analyzed to identify 19 cell clusters. We identified 3,464 marker genes from the scRNA-seq dataset, 1,994 differentially expressed genes from the bulk RNA sequencing (RNA-seq) dataset, and 1,863 genes associated with a key module identified by the WGCNA. After performing the intersection, univariate Cox, and least absolute shrinkage and selection operator analyses, a prognostic model was established based on the expression levels of 13 signature genes. Subsequent functional experiments confirmed the role of selected regulated genes. Through the integration of scRNA-seq data and bulk RNA-seq data, we developed an innovative model to predict the prognosis of patients. The risk score was found to be a significant independent predictor and clinical-pathological features of LUAD.

Most sequencing-based spatial transcriptomics (ST) technologies do not achieve single-cell resolution where each captured location (spot) may contain a mixture of cells from heterogeneous cell types, and several cell-type decomposition methods have been proposed to estimate cell type proportions of each spot by integrating with single-cell RNA sequencing (scRNA-seq) data. However, these existing methods did not fully consider the effect of distribution difference between scRNA-seq and ST data for decomposition, leading to biased cell-type-specific genes derived from scRNA-seq for ST data. To address this issue, we develop an instance-based transfer learning framework to adjust scRNA-seq data by ST data to correctly match cell-type-specific gene expression. We evaluate the effect of raw and adjusted scRNA-seq data on cell-type decomposition by eight leading decomposition methods using both simulated and real datasets. Experimental results show that data adjustment can effectively reduce distribution difference and improve decomposition, thus enabling for a more precise depiction on spatial organization of cell types. We highlight the importance of data adjustment in integrative analysis of scRNA-seq with ST data and provide guidance for improved cell-type decomposition.

BackgroundSingle-cell RNA sequencing (scRNA-seq) provides an effective tool to investigate the transcriptomic characteristics at the single-cell resolution. Due to the low amounts of transcripts in single cells and the technical biases in experiments, the raw scRNA-seq data usually includes large noise and makes the downstream analyses complicated. Although many methods have been proposed to impute the noisy scRNA-seq data in recent years, few of them take into account the prior associations across genes in imputation and integrate multiple types of imputation data to identify cell types.ResultsWe present a new framework, NetImpute, towards the identification of cell types from scRNA-seq data by integrating multiple types of biological networks. We employ a statistic method to detect the noise data items in scRNA-seq data and develop a new imputation model to estimate the real values of data noise by integrating the PPI network and gene pathways. Meanwhile, based on the data imputed by multiple types of biological networks, we propose an integrated approach to identify cell types from scRNA-seq data. Comprehensive experiments demonstrate that the proposed network-based imputation model can estimate the real values of noise data items accurately and integrating the imputation data based on multiple types of biological networks can improve the identification of cell types from scRNA-seq data.ConclusionsIncorporating the prior gene associations in biological networks can potentially help to improve the imputation of noisy scRNA-seq data and integrating multiple types of network-based imputation data can enhance the identification of cell types. The proposed NetImpute provides an open framework for incorporating multiple types of biological network data to identify cell types from scRNA-seq data.

Cell type prediction is one of the most challenging goals in single-cell RNA sequencing (scRNA-seq) data. Existing methods use unsupervised learning to identify signature genes in each cluster, followed by a literature survey to look up those genes for assigning cell types. However, finding potential marker genes in each cluster is cumbersome, which impedes the systematic analysis of single-cell RNA sequencing data. To address this challenge, we proposed a framework based on regularized multi-task learning (RMTL) that enables us to simultaneously learn the subpopulation associated with a particular cell type. Learning the structure of subpopulations is treated as a separate task in the multi-task learner. Regularization is used to modulate the multi-task model (e.g., W1, W2, … Wt) jointly, according to the specific prior. For validating our model, we trained it with reference data constructed from a single-cell RNA sequencing experiment and applied it to a query dataset. We also predicted completely independent data (the query dataset) from the reference data which are used for training. We have checked the efficacy of the proposed method by comparing it with other state-of-the-art techniques well known for cell type detection. Results revealed that the proposed method performed accurately in detecting the cell type in scRNA-seq data and thus can be utilized as a useful tool in the scRNA-seq pipeline.

Single-cell RNA sequencing (scRNA-seq) maps gene expression heterogeneity within a tissue. However, identifying biological signals in this data is challenging due to confounding technical factors, sparsity, and high dimensionality. Data factorization methods address this by separating and identifying signals in the data, such as gene expression programs, but the resulting factors must be manually interpreted. We developed Single-Cell Interpretable REsidual Decomposition (sciRED) to improve the interpretation of scRNA-seq factor analysis. sciRED removes known confounding effects, uses rotations to improve factor interpretability, maps factors to known covariates, identifies unexplained factors that may capture hidden biological phenomena and determines the genes and biological processes represented by the resulting factors. We apply sciRED to multiple scRNA-seq datasets and identify sex-specific variation in a kidney map, discern strong and weak immune stimulation signals in a PBMC dataset, reduce ambient RNA contamination in a rat liver atlas to help identify strain variation, and reveal rare cell type signatures and anatomical zonation gene programs in a healthy human liver map. These demonstrate that sciRED is useful in characterizing diverse biological signals within scRNA-seq data.

BackgroundCell clustering is a prerequisite for identifying differentially expressed genes (DEGs) in single-cell RNA sequencing (scRNA-seq) data. Obtaining a perfect clustering result is of central importance for subsequent analyses, but not easy. Additionally, the increase in cell throughput due to the advancement of scRNA-seq protocols exacerbates many computational issues, especially regarding method runtime. To address these difficulties, a new, accurate, and fast method for detecting DEGs in scRNA-seq data is needed.ResultsHere, we propose single-cell minimum enclosing ball (scMEB), a novel and fast method for detecting single-cell DEGs without prior cell clustering results. The proposed method utilizes a small part of known non-DEGs (stably expressed genes) to build a minimum enclosing ball and defines the DEGs based on the distance of a mapped gene to the center of the hypersphere in a feature space.ConclusionsWe compared scMEB to two different approaches that could be used to identify DEGs without cell clustering. The investigation of 11 real datasets revealed that scMEB outperformed rival methods in terms of cell clustering, predicting genes with biological functions, and identifying marker genes. Moreover, scMEB was much faster than the other methods, making it particularly effective for finding DEGs in high-throughput scRNA-seq data. We have developed a package scMEB for the proposed method, which could be available at https://github.com/FocusPaka/scMEB.

Hematopoietic Cell Cluster Identifier (HCCI): A Knowledgebase Computational Tool to Facilitate the Characterization of Hematopoietic Cell Populations in Single Cell RNA-Sequencing