RNA binding protein enrichment analysis method based on single cell expression profile data

Abstract

The invention discloses an RNA (Ribonucleic Acid) binding protein enrichment analysis method based on single cell expression profile data, which comprises the following steps of: writing an interactive online Web application program by using Shiny, carrying a data visualization tool, and using an RBP (Reactive Binding Protein)-gene database as a database; the method is specially designed for single cell data, different single cell data can be integrated for enrichment analysis, and expression profiles and RBP dynamic changes in single cells can be effectively captured and analyzed; rBP enrichment analysis is customized for single cell data, a single group and multiple groups of single cell expression data are preprocessed, including normalization processing, and a statistical model is used to evaluate enrichment significance at each time point, so that the analysis accuracy and biological correlation are improved; a dynamic visualization tool is provided, the dynamic visualization tool comprises a customizable network diagram, an interactive bar diagram and detailed table display, and a user can adjust a plurality of parameters of a view according to needs and directly download images and data from an interface.

Description

Technical Field

[0001] This invention relates to the field of bioinformatics technology, specifically to a method for enriching RNA-binding proteins based on single-cell expression profile data. Background Technology

[0002] RNA binding proteins (RBPs), similar to transcription factors, are a class of proteins that specifically bind to RNA molecules and play important roles in multiple biological processes, including RNA processing, stability, and translation. Particularly at the single-cell level, RBPs influence cell fate decisions and developmental processes, revealing regulatory mechanisms in cellular heterogeneity and state transitions. Previously, due to technological limitations, RBP detection methods such as Northern blotting could only achieve low-throughput detection. However, in recent years, significant breakthroughs have been made in new high-throughput detection technologies. CLIP-seq (Crosslinking and Immunoprecipitation followed by sequencing) crosslinks RBPs to their RNA targets, followed by immunoprecipitation and sequencing analysis to identify RBP binding sites, enabling efficient and comprehensive RBP detection.

[0003] In the field of single-cell research, a large amount of RBP data has been accumulated and published in public datasets such as ENCODE. However, there are still few RBP enrichment analysis tools specifically for single-cell data, and there is still a gap in RBP research.

[0004] Enrichment analysis can quickly identify key RNA binding proteins (RBPs), which is crucial for understanding the RNA regulatory network at the cellular level. Therefore, we propose an RNA binding protein enrichment analysis method based on single-cell expression profile data. Summary of the Invention

[0005] The purpose of this invention is to provide a method for enriching RNA-binding proteins based on single-cell expression profile data, in order to solve the problems mentioned in the background art.

[0006] To address the aforementioned technical problems, this invention provides the following technical solution: an RNA-binding protein enrichment analysis method based on single-cell expression profile data, developed as an interactive online web application using Shiny, equipped with data visualization tools, and using the RBP-gene database, comprising the following steps:

[0007] Step S1: Provide a front-end page for users to import single-cell expression profile data and select species;

[0008] Step S2: Obtain the single-cell expression profile data and selected species imported by the user through the front-end page, combine them with the built-in RBP-gene database, perform statistical tests based on hypergeometric distribution, and conduct RBP enrichment analysis.

[0009] Step S3: Based on the enrichment analysis results, set the analysis parameters and visualize the dynamic interactive charts.

[0010] According to the above technical solution, the single-cell expression profile data is a single-group single-cell expression profile data;

[0011] According to the above technical solution, step S1 specifically includes: copying the gene name and pasting it into the search box, and selecting the corresponding species according to the species to which the gene belongs;

[0012] According to the above technical solution, for the enrichment analysis of single-cell expression profile data, in step S3, the dynamic visualization enrichment analysis results include bar charts displaying the enrichment results, tables displaying the enrichment results, and customized network diagrams displaying the enrichment results.

[0013] According to the above technical solution, the single-cell expression profile data consists of multiple sets of single-cell expression profile data;

[0014] According to the above technical solution, step S1 specifically includes: uploading expression profile data in .xls file format, with the first column being the gene name, the second column being the grouping information, the first row being the ID and group, and selecting the corresponding species according to the species to which the gene belongs;

[0015] According to the above technical solution, for the enrichment analysis of multiple groups of single-cell expression profile data, in step S3, the dynamic visualization enrichment analysis results include grouped dot plots showing the enrichment results, tables showing the enrichment results, integrated customized network diagrams showing the enrichment results, and grouped customized network diagrams showing the enrichment results.

[0016] According to the above technical solution, in step S3, for the dynamic interactive chart, users are provided with further viewing, modification, and download operations, and the modifications include chart color, font size, title, and arrangement scheme;

[0017] According to the above technical solution, it also includes RBP-Gene browser and database query, and the steps are as follows:

[0018] Step 1: Enter a target gene or RBP name and select the corresponding species, then select the gene or RBP to query;

[0019] Step 2: Based on the information entered in Step 1, and combined with the built-in RBP-gene database, perform a query and output the query results in a table.

[0020] According to the above technical solution, in step 2, the columns of the table are geneName, RBP, clusterNum, chromosome, narrowStart, NarrowEnd, strand, and ENTREZID in sequence. The table supports global search, column search, sorting, and downloading of xls and bed format tables.

[0021] Compared with existing technologies, the beneficial effects achieved by this invention are as follows: It proposes an RNA-binding protein enrichment analysis method based on single-cell expression profile data. In terms of data integration and processing, it is specifically designed for single-cell data, enabling the integration of different single-cell data for enrichment analysis, effectively capturing and analyzing the dynamic changes in expression profiles and RBP within single cells. Regarding the enrichment analysis method, it provides a customized RBP enrichment analysis for single-cell data, preprocessing single and multiple groups of single-cell expression data, including normalization, and using statistical models to evaluate the significance of enrichment at each time point, improving the accuracy and biological relevance of the analysis. In terms of visualization interaction and customization functions, it provides dynamic visualization tools, including customizable network diagrams, interactive bar charts, and detailed table displays. Users can adjust multiple parameters of the view as needed and directly download images and data from the interface. Attached Figure Description

[0022] The accompanying drawings are provided to further illustrate the invention and form part of the specification. They are used in conjunction with embodiments of the invention to explain the invention and do not constitute a limitation thereof. In the drawings:

[0023] Figure 1 This is a flowchart of the method of the present invention; Detailed Implementation

[0024] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0025] This invention provides a technical solution: an RNA-binding protein enrichment analysis method based on single-cell expression profile data, which is developed into an interactive online web application using Shiny, equipped with data visualization tools, and uses the RBP-gene database.

[0026] To provide human and mouse RBP databases, follow these steps:

[0027] Step 1: Select either the human RBP database or the mouse RBP database (available for download);

[0028] Step 2: Display the data table, with three columns: geneName, Protein Name, and Alpha Names.

[0029] Step 3, Data Retrieval (provides global search, single column search, and sorting).

[0030] The RBP-Gene browser and database query are provided, and the steps are as follows:

[0031] Step 1: Enter a target gene or RBP name and select the corresponding species, then select the gene or RBP to query;

[0032] Step 2: Based on the information entered in Step 1, and combined with the built-in RBP gene database, perform a query and output the query results in a table. The columns of the table are geneName, RBP, clusterNum, chromosome, narrowStart, NarrowEnd, strand, and ENTREZID. The table supports global search, column search, sorting, and downloading in xls and .bed format.

[0033] Example 1

[0034] The method for enriching RNA-binding proteins based on single-cell expression profile data includes the following steps:

[0035] Step S1: Provide a front-end page for users to import single-cell expression profile data and select species;

[0036] Specifically, copy the gene name, paste it into the search box, and select the corresponding species based on the species to which the gene belongs;

[0037] Step S2: Obtain the single-cell expression profile data and selected species imported by the user through the front-end page, combine it with the built-in RBP-gene database, perform statistical tests based on hypergeometric distribution, and conduct RBP enrichment analysis.

[0038] Step S3: Based on the enrichment analysis results, set the analysis parameters and visualize the results using dynamic interactive charts. The dynamic visualization of the enrichment analysis results includes bar charts, tables, and customized network diagrams. The dynamic interactive charts allow users to further view, modify, and download the results. Modifications include chart color, font size, title, and arrangement.

[0039] The horizontal axis of the bar chart is -log10(Pvalue), and the vertical axis is each RBP arranged from largest to smallest according to -log(Pvalue). The bar colors range from red to blue, and by default, the top 10 most significant ones (P<0.1) are displayed.

[0040] The columns of the table are RBP, pvalue, GeneRatio, BackRatio, and genes, respectively. It supports global search, single-column search, and sorting. The first 10 entries are displayed by default, and the number of entries displayed can be modified. The table can be downloaded.

[0041] The customized network diagram offers a variety of layout options, adjustable RBP count (0-50), adjustable font size (0-50), and adjustable color configuration.

[0042] Example 2

[0043] An RNA-binding protein enrichment analysis method based on multiple sets of single-cell expression profile data includes the following steps:

[0044] Step S1: Provide a front-end page for users to import multiple sets of single-cell expression profile data and select species;

[0045] Specifically, the expression profile data is uploaded in .xls file format, with the first column being the gene name, the second column being the grouping information, the first row being the ID and group, and the corresponding species being selected according to the species to which the gene belongs;

[0046] Step S2: Obtain multiple sets of single-cell expression profile data and selected species imported by the user through the front-end page, combine them with the built-in RBP-gene database, perform statistical tests based on hypergeometric distribution, and conduct RBP enrichment analysis.

[0047] Step S3: Based on the enrichment analysis results, set the analysis parameters and visualize the results using dynamic interactive charts. The dynamic visualization of the enrichment analysis results includes grouped dot plots, tables, integrated customized network diagrams, and grouped customized network diagrams. The dynamic interactive charts allow users to further view, modify, and download the results. Modifications include chart color, font size, title, and arrangement.

[0048] The horizontal axis of the scatter plot represents different groups, and the vertical axis represents each RBP arranged from largest to smallest according to -log(Pvalue), with the dots colored from red to blue. The size of the dot indicates the number of enriched genes (P<0.1).

[0049] The columns of the table are RBP, pvalue, GeneRatio, BackRatio, and genes, respectively. It supports global search, single-column search, and sorting. The first 10 entries are displayed by default, and the number of entries displayed can be modified. The table can be downloaded.

[0050] The integrated, customized network diagram combines and displays enriched RBPs and genes, offering multiple arrangement options, adjustable RBP count (0-50), adjustable font size (0-50), and adjustable color scheme.

[0051] The customized network diagram grouping displays enriched RBPs and genes, shows grouping information, offers multiple arrangement options, and allows for adjustment of the number of RBPs (0-50), font size (0-50), and color scheme.

[0052] Finally, it should be noted that the above descriptions are merely preferred embodiments of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for enriching RNA-binding proteins based on single-cell expression profile data, characterized in that: The application was developed using Shiny as an interactive online web application, equipped with data visualization tools, and uses the RBP-gene database. The steps included are as follows: Step S1: Provide a front-end page for users to import single-cell expression profile data and select species; Step S2: Obtain the single-cell expression profile data and selected species imported by the user through the front-end page, combine them with the built-in RBP-gene database, perform statistical tests based on hypergeometric distribution, and conduct RBP enrichment analysis. Step S3: Based on the enrichment analysis results, set the analysis parameters and visualize the dynamic interactive charts.

2. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 1, characterized in that: The single-cell expression profile data refers to single-group single-cell expression profile data.

3. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 2, characterized in that: Step S1 specifically includes: copying the gene name and pasting it into the search box, and selecting the corresponding species according to the species to which the gene belongs.

4. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 2, characterized in that: For enrichment analysis of single-cell expression profile data, in step S3, the dynamic visualization enrichment analysis results include bar charts, tables, and customized network diagrams.

5. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 1, characterized in that: The single-cell expression profile data consists of multiple sets of single-cell expression profile data.

6. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 5, characterized in that: Step S1 specifically includes: uploading expression profile data in .xls file format, with the first column being the gene name, the second column being the grouping information, the first row being the ID and group, and selecting the corresponding species according to the species to which the gene belongs.

7. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 5, characterized in that: For enrichment analysis of multiple single-cell expression profile data, in step S3, the dynamic visualization enrichment analysis results include enrichment results displayed by grouped dot plots, enrichment results displayed in tables, enrichment results displayed by integrated customized network graphs, and enrichment results displayed by grouped customized network graphs.

8. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 1, characterized in that: In step S3, for the dynamic interactive chart, users are provided with further viewing, modification, and download operations. The modifications include chart color, font size, title, and arrangement scheme.

9. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 1, characterized in that: It also includes the RBP-Gene browser and database query, the steps of which are as follows: Step 1: Enter a target gene or RBP name and select the corresponding species, then select the gene or RBP to query; Step 2: Based on the information entered in Step 1, and combined with the built-in RBP-gene database, perform a query and output the query results in a table.

10. The method for enriching RNA-binding proteins based on single-cell expression profile data according to claim 9, characterized in that: In step 2, the columns of the table are geneName, RBP, clusterNum, chromosome, narrowStart, NarrowEnd, strand, and ENTREZID. The table supports global search, column search, sorting, and downloading in xls and bed formats.

Images