BioinformaticsDashboardv0.0..Rmd

---
title: "Bioinformatics Dashboard v0.0 🧬 🦠 🧫"
output: 
  flexdashboard::flex_dashboard:
    orientation: columns
    vertical_layout: fill
  runtime: shiny 
---

```{r setup, include=FALSE}
if (!requireNamespace("pacman", quietly = TRUE)) {
  install.packages("pacman")
}

pacman::p_load(flexdashboard, shiny, ggplot2, plotly, clusterProfiler, readxl, tidyverse, DESeq2, biomaRt, tidyr, shinyjs, rentrez, dplyr, ggtext, cowplot, UpSetR)

library(flexdashboard)
library(shiny)
library(ggplot2)
library(plotly)
library(clusterProfiler)
library(readxl)
library(tidyverse)
library(DESeq2)
library(biomaRt)
library(tidyr)
library(shinyjs)
library(rentrez)
library(dplyr) 
library(ggtext)
library(cowplot)
library(UpSetR)
```

Column {.tabset}
-----------------------------------------------------------------------

### RNAseq analysis
Analyzes RNAseq data using DESeq2 and GSEA. Visualizes using volcano plot, and other plots to show the GSEA analysis results. Please make sure the uploaded data is in xlsx format, has the first column with the gene names, and there is an even number of data columns. The control condition should be first, the mutant condition second. 
```{r}

ui <- fluidPage(
  titlePanel("Interactive Volcano Plot with Gene and GO Term Search"),
  useShinyjs(),
  
  passwordInput("password", "Enter Password:", value = "", placeholder = "Password"),
  actionButton("submit_password", "Submit"),
  
  uiOutput("main_ui"),
  
  #if you define a side_ui --> absolute panel, it would have to be defined here
)

# Server logic
server <- function(input, output, session) {
  # Password handling
  correct_password <- "my_secret_password"
  
  observeEvent(input$submit_password, {
    if (input$password == correct_password) {
      showModal(modalDialog(
        title = "Access Granted",
        "Welcome! You can now search for genes and view the volcano plot.",
        easyClose = TRUE,
        footer = NULL
      ))
      
      # Hide the password input and button after validation
      shinyjs::hide("password")
      shinyjs::hide("submit_password")
      
      # Main UI appears after password is correct
      output$main_ui <- renderUI({
        sidebarLayout(
          sidebarPanel(
            fluidRow(
              fileInput("file", "Choose XLSX File", multiple = FALSE, accept = c(".xlsx", "text/xlsx")),
              actionButton("analyze_button", "Analyze"),
              tags$hr(),
              textInput("gene_search", "Search for a gene or keyword (separate multiple genes with ';'):", ""),
              actionButton("search_gene", "Search Gene"),
              tags$hr(),
              selectInput("GO_search", "Select a GO term:", choices = NULL),
              actionButton("search_GO_term", "Search GO Term"),
              tags$hr(),
              selectInput("description_search", "Search for the name of a pathway:", choices = NULL),
              actionButton("search_description", "Search Description"),
              tags$hr(),
              sliderInput("pvalue", "P-value: ",
                          min = 0, max = 1,
                          value = 0.01, step = 0.00001),
              tags$hr(),
              sliderInput("log2fc", "Log2FoldChange: ", 
                          min = 0.0001, max = 100, 
                          value = 2.5, step = 0.05),
              tags$hr(), 
              actionButton("visualize_gse", "Visualize the GSEGO Results:")
            )
          ),
          mainPanel(
            plotlyOutput("volcanoPlot"),
            plotOutput("dotPlotTitle", width = "100%", height = "100px"),
            plotOutput("dotPlot", width = "100%", height = "1000px"), 
            plotOutput("conceptNetworkTitle", width = "100%", height = "100px"),
            plotOutput("conceptNetwork", width = "100%", height = "600px"),
            plotOutput("heatMapTitle", width = "100%", height = "100px"),
            plotOutput("heatMap", width = "100%", height = "400px"), 
            plotOutput("upsetPlotTitle", width = "100%", height = "100px"),
            plotOutput("upsetPlot", width = "100%", height = "1000px"),
            plotOutput("pubmedPathwayPlotTitle", width = "100%", height = "100px"),
            plotOutput("pubmedPathwayPlot", width = "100%", height = "1500px")
          )
        )
      })
      
      # Reactive values to store results and search criteria
      searchValues <- reactiveValues(
        gene_search = "",
        GO_search = "All",
        description_search = "",
        df_inverted = NULL  # Store df_inverted here
      )
      
      # Process uploaded file and perform DESeq2 analysis
      observeEvent(input$analyze_button, {
        req(input$file)  # Ensure a file is uploaded
        
        # Read in gene counts data
        genecounts <- tryCatch({
          read_excel(input$file$datapath, sheet = 1, col_names = TRUE)
        }, error = function(e) {
          showModal(modalDialog(title = "Error", "Could not read the Excel file.", easyClose = TRUE))
          return(NULL)
        })
        
        if (is.null(genecounts)) return(NULL)  # Stop further processing if reading failed
        
        genecounts <- as.data.frame(genecounts)
        rownames(genecounts) <- genecounts[, 1]
        genecounts$Gene_Name <- NULL
        genecounts <- genecounts[, -1]
        
        num_samples <- ncol(genecounts)
        
        # Check if the number of samples is even
        if (num_samples %% 2 != 0) {
          showModal(modalDialog(
            title = "Error",
            "The number of samples must be even for proper grouping.",
            easyClose = TRUE
          ))
          return(NULL)
        }
        
        # Create the condition data frame
        condition <- data.frame(genotype = rep(c('C', 'R'), each = num_samples / 2), row.names = colnames(genecounts))
        
        # Create DESeq2 dataset
        dds <- DESeqDataSetFromMatrix(countData = genecounts, colData = condition, design = ~genotype)
        de <- DESeq(dds)
        res_reactive <- reactiveVal()
        res_reactive(results(de))
        res <<- results(de)
        
        # Create additional columns for plotting
        res$pvalue_log10 <- -log10(res$pvalue)
        pvalue_threshold <- 0.05
        fold_change_threshold <- 2
        
        res$significance <- ifelse(res$pvalue < pvalue_threshold, "Significant", "Not Significant")
        res$new_column <- rownames(res)
        res$diffexpressed <- ifelse(res$log2FoldChange > 0, "UP", ifelse(res$log2FoldChange < 0, "DOWN", "NO_CHANGE"))
        
        # Generate gene list for GSEA
        organism = "org.Hs.eg.db"
        original_gene_list <- res$log2FoldChange
        names(original_gene_list) <- res$new_column
        gene_list <<- na.omit(original_gene_list)
        gene_list = sort(gene_list, decreasing = TRUE)
        
        # Perform GO enrichment analysis
        gse <<- gseGO(geneList = gene_list, 
                      ont = "ALL", 
                      keyType = "SYMBOL", 
                      minGSSize = 3, 
                      maxGSSize = 800, 
                      pvalueCutoff = 0.05, 
                      verbose = TRUE, 
                      OrgDb = organism, 
                      pAdjustMethod = "none")
        
        # Store inverted results for GO terms in reactive values
        searchValues$df_inverted <- gse@result %>% separate_rows(core_enrichment, sep = "/")
        
        # Update GO term and description choices in UI
        updateSelectInput(session, "GO_search", choices = unique(searchValues$df_inverted$ID))
        updateSelectInput(session, "description_search", choices = unique(searchValues$df_inverted$Description))
        
        # Reactive filtering of results based on user input
        filteredRes <- reactive({
          data <- as.data.frame(res)
          
          # Apply gene search filter
          if (searchValues$gene_search != "") {
            genes <- strsplit(searchValues$gene_search, ";")[[1]]
            genes <- trimws(genes)
            data <- data %>%
              filter(rowSums(sapply(genes, function(gene) grepl(gene, new_column, ignore.case = TRUE))) > 0)
          }
          
          # Apply GO term filter
          if (searchValues$GO_search != "All") {
            selected_genes <- searchValues$df_inverted %>%
              filter(ID == searchValues$GO_search) %>%
              pull(core_enrichment)
            data <- data %>%
              filter(new_column %in% selected_genes)
          }
          
          # Apply description search filter
          if (searchValues$description_search != "") {
            selected_genes <- searchValues$df_inverted %>%
              filter(Description == searchValues$description_search) %>%
              pull(core_enrichment)
            data <- data %>%
              filter(new_column %in% selected_genes)
          }
          
          data
        })
        
        # Render volcano plot based on filtered results
        output$volcanoPlot <- renderPlotly({
          data_res <- filteredRes()
          p_value <- input$pvalue #need to add slides here
          log2fc <- input$log2fc #need to add slider here
          
          p <- ggplot(data_res, aes(x = log2FoldChange, y = pvalue_log10, 
                                    text = paste("Gene:", new_column, "<br>Log2 Fold Change:", log2FoldChange, 
                                                 "<br>P-value:", pvalue, "<br>Significance:", significance, 
                                                 "<br>Differentially Expressed:", diffexpressed, "<br>-log10 Values:", pvalue_log10))) +
            geom_point(aes(color = log2FoldChange, shape = diffexpressed)) +
            geom_hline(yintercept = -log10(p_value), linetype = "dotted", color = "red") +
            geom_vline(xintercept = c(-log2fc, log2fc), linetype = "dotted", color = "darkblue") +
            xlim(-5, 5) +
            xlab("Log2 Fold Change") + 
            ylab("-log10(P-value)") +
            ggtitle("Volcano Plot") +
            scale_color_gradient2(low = "green", mid = "pink", high = "blue", midpoint = 0, 
                                  name = "Log2 Fold Change")  # Add a custom color scale for the color legends
          
          ggplotly(p, tooltip = "text")
        })
      })
      
      # Update search criteria based on user actions
      observeEvent(input$search_gene, {
        searchValues$gene_search <- input$gene_search
        searchValues$GO_search <- "All"
        searchValues$description_search <- ""
      })
      
      observeEvent(input$search_GO_term, {
        searchValues$GO_search <- input$GO_search
        searchValues$gene_search <- ""
        searchValues$description_search <- ""
      })
      
      observeEvent(input$search_description, {
        searchValues$description_search <- input$description_search
        searchValues$GO_search <- "All"
        searchValues$gene_search <- ""
      })
      
      observeEvent(input$visualize_gse, {
        library(ggplot2)
        library(ggtext)
        library(gridExtra)
        
        output$dotPlotTitle <- renderPlot({
          txt <- "Dot Plot with 10 Pathways"
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) +
            theme_void()
          print(title_plot)
        })
        
        output$dotPlot <- renderPlot({
          dot_plot <- dotplot(gse, showCategory = 10)
          print(dot_plot)
          
        })
        
        output$conceptNetworkTitle <- renderPlot({
          txt <- "Gene Concept Network"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$conceptNetwork <- renderPlot({
          gsex <- setReadable(gse, 'org.Hs.eg.db', 'ENTREZID')
          geneList <- gse@geneList
          
          p1 <- cnetplot(gsex, foldChange = geneList, max.overlaps = 100)
          p2 <- cnetplot(gsex, categorySize = "pvalue", foldChange = geneList, max.overlaps = 100)
          p3 <- cnetplot(gsex, foldChange = geneList, circular = TRUE, colorEdge = TRUE, max.overlaps = 100)
          
          maingene_plot <- cowplot::plot_grid(p1, p2, p3, ncol = 3, labels = LETTERS[1:3], rel_widths = c(.8, .8, 1.2))
          print(maingene_plot)
          
        })
        
        output$heatMapTitle <- renderPlot({
          txt <- "Heatmap-Like Functional Classification"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$heatMap <- renderPlot({
          
          gsex <- setReadable(gse, 'org.Hs.eg.db', 'ENTREZID')
          geneList <- gse@geneList
          
          p1 <- heatplot(gsex, showCategory=5)
          p2 <- heatplot(gsex, foldChange=geneList, showCategory=5)
          mainheatmap_plot <- cowplot::plot_grid(p1, p2, ncol=1, labels=LETTERS[1:2])
          
          print(mainheatmap_plot)
        })
        
        output$upsetPlotTitle <- renderPlot({
          txt <- "UpSet Plot"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$upsetPlot <- renderPlot({
          gse_df <- gse@result
          top_terms <- gse_df %>% arrange(pvalue) %>% head(10)
          top_gene_sets <- strsplit(top_terms$core_enrichment, "/")
          gene_sets_list <- lapply(top_gene_sets, function(x) unique(trimws(x)))
          
          gene_sets_df <- fromList(setNames(gene_sets_list, top_terms$ID))
          
          # Create the UpSet plot
          upset_plot <- upset(gene_sets_df, 
                              sets = names(gene_sets_df), 
                              main.bar.color = "steelblue",
                              sets.bar.color = "darkred", 
                              order.by = "freq", 
                              matrix.color = "gray",
                              keep.order = TRUE)
          
          print(upset_plot)
        })
        
        output$pubmedPathwayPlotTitle <- renderPlot({
          txt <- "PubMed Pathway Enrichment"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$pubmedPathwayPlot <- renderPlot({
          results <- data.frame(Term = character(), Year = integer(), Count = integer(), stringsAsFactors = FALSE)
          
          terms <- tail(gse$Description, n = 10)
          
          results <- data.frame()
          titles_2024 <- data.frame()
          
          for (term in terms) {
            for (year in 2014:2024) {
              query <- paste(term, "[Title/Abstract] AND", year, "[PDAT]")
              
              # Count results for each term and year
              search_results <- entrez_search(db = "pubmed", term = query, retmax = 0)
              results <- rbind(results, data.frame(Term = term, Year = year, Count = search_results$count))
              
              # If the year is 2024, retrieve the first 10 article titles
              if (year == 2024) {
                search_results_2024 <- entrez_search(db = "pubmed", term = query, retmax = 10)
                if (search_results_2024$count > 0) {
                  article_ids <- search_results_2024$ids
                  articles <- entrez_fetch(db = "pubmed", id = article_ids, rettype = "abstract", retmode = "text")
                  titles <- sapply(strsplit(articles, "\n"), function(x) x[1])
                  titles_2024 <- reactive({rbind(titles_2024, data.frame(Term = term, Title = titles, stringsAsFactors = FALSE))})
                }
              }
            }
          }
          
          total_counts <- results %>%
            group_by(Year) %>%
            summarize(Total_Count = sum(Count), .groups = 'drop')
          
          results <- results %>%
            left_join(total_counts, by = "Year")
          
          results <- results %>%
            mutate(Ratio = Count / Total_Count)
          
          print(results)
          
          mainpubmed_plot <- ggplot(results, aes(x = Year, y = Ratio, color = Term)) +
            geom_line() +
            geom_point(size = 3, shape = 20, fill = "white", stroke = 1) +  # Bolded dots
            scale_x_continuous(limits = c(2013, 2025), breaks = seq(2013, 2025, by = 2.5)) +  # 2.5-year breaks
            labs(title = "Publication Ratio for Enriched Terms", x = "Year", y = "Publication Ratio") +
            theme_minimal()
          
          print(mainpubmed_plot)
        })
      })
      
    } else {
      showModal(modalDialog(
        title = "Access Denied",
        "Incorrect password. Please try again.",
        easyClose = TRUE,
        footer = NULL
      ))
    }
  })
}

shinyApp(ui = ui, server = server)
```

### Proteomics analysis
Analyzes proteomics data using DESeq2 and GSEA. Visualizes using volcano plot, and other plots to show the GSEA analysis results. Please make sure the uploaded data is in xlsx format, has the first column with the gene names, and the second column has the expanded name of each gene and there is an even number of data columns. The control condition should be first, the mutant condition second. 
```{r}

ui <- fluidPage(
  titlePanel("Interactive Volcano Plot with Gene and GO Term Search"),
  useShinyjs(),
  
  passwordInput("password", "Enter Password:", value = "", placeholder = "Password"),
  actionButton("submit_password", "Submit"),
  
  uiOutput("main_ui"),
  
  #if you define a side_ui --> absolute panel, it would have to be defined here
)

# Server logic
server <- function(input, output, session) {
  # Password handling
  correct_password <- "my_secret_password"
  
  observeEvent(input$submit_password, {
    if (input$password == correct_password) {
      showModal(modalDialog(
        title = "Access Granted",
        "Welcome! You can now search for genes and view the volcano plot.",
        easyClose = TRUE,
        footer = NULL
      ))
      
      # Hide the password input and button after validation
      shinyjs::hide("password")
      shinyjs::hide("submit_password")
      
      # Main UI appears after password is correct
      output$main_ui <- renderUI({
        sidebarLayout(
          sidebarPanel(
            fluidRow(
              fileInput("file", "Choose XLSX File", multiple = FALSE, accept = c(".xlsx", "text/xlsx")),
              actionButton("analyze_button", "Analyze"),
              tags$hr(),
              textInput("gene_search", "Search for a gene or keyword (separate multiple genes with ';'):", ""),
              actionButton("search_gene", "Search Gene"),
              tags$hr(),
              selectInput("GO_search", "Select a GO term:", choices = NULL),
              actionButton("search_GO_term", "Search GO Term"),
              tags$hr(),
              selectInput("description_search", "Search for the name of a pathway:", choices = NULL),
              actionButton("search_description", "Search Description"),
              tags$hr(),
              sliderInput("pvalue", "P-value: ",
                          min = 0, max = 1,
                          value = 0.01, step = 0.00001),
              tags$hr(),
              sliderInput("log2fc", "Log2FoldChange: ", 
                          min = 0.0001, max = 100, 
                          value = 2.5, step = 0.05),
              tags$hr(), 
              actionButton("visualize_gse", "Visualize the GSEGO Results:")
            )
          ),
          mainPanel(
            plotlyOutput("volcanoPlot"),
            plotOutput("dotPlotTitle", width = "100%", height = "100px"),
            plotOutput("dotPlot", width = "100%", height = "1000px"), 
            plotOutput("conceptNetworkTitle", width = "100%", height = "100px"),
            plotOutput("conceptNetwork", width = "100%", height = "600px"),
            plotOutput("heatMapTitle", width = "100%", height = "100px"),
            plotOutput("heatMap", width = "100%", height = "400px"), 
            plotOutput("upsetPlotTitle", width = "100%", height = "100px"),
            plotOutput("upsetPlot", width = "100%", height = "1000px"),
            plotOutput("pubmedPathwayPlotTitle", width = "100%", height = "100px"),
            plotOutput("pubmedPathwayPlot", width = "100%", height = "1500px")
          )
        )
      })
      
      # Reactive values to store results and search criteria
      searchValues <- reactiveValues(
        gene_search = "",
        GO_search = "All",
        description_search = "",
        df_inverted = NULL  # Store df_inverted here
      )
      
      # Process uploaded file and perform DESeq2 analysis
      observeEvent(input$analyze_button, {
        req(input$file)  # Ensure a file is uploaded
        
        # Read in gene counts data
        genecounts <- tryCatch({
          read_excel(input$file$datapath, sheet = 1, col_names = TRUE)
        }, error = function(e) {
          showModal(modalDialog(title = "Error", "Could not read the Excel file.", easyClose = TRUE))
          return(NULL)
        })
        
        if (is.null(genecounts)) return(NULL)  # Stop further processing if reading failed
        
        genecounts <- as.data.frame(genecounts)
        rownames(genecounts) <- genecounts[, 1]
        genecounts <- genecounts[, -1]
        descriptions <<- data.frame(Description = genecounts[, 1])  
        rownames(descriptions) <- rownames(genecounts)
        genecounts <- genecounts[, -1]  
        num_samples <- ncol(genecounts)
        
        num_samples <- ncol(genecounts)
        
        # Check if the number of samples is even
        if (num_samples %% 2 != 0) {
          showModal(modalDialog(
            title = "Error",
            "The number of samples must be even for proper grouping.",
            easyClose = TRUE
          ))
          return(NULL)
        }
        
        # Create the condition data frame
        condition <- data.frame(genotype = rep(c('C', 'R'), each = num_samples / 2), row.names = colnames(genecounts))
        
        # Create DESeq2 dataset
        dds <- DESeqDataSetFromMatrix(countData = genecounts, colData = condition, design = ~genotype)
        de <- DESeq(dds)
        res_reactive <- reactiveVal()
        res_reactive(results(de))
        res <<- results(de)
        
        # Create additional columns for plotting
        res$pvalue_log10 <- -log10(res$pvalue)
        pvalue_threshold <- 0.05
        fold_change_threshold <- 2
        
        res$significance <- ifelse(res$pvalue < pvalue_threshold, "Significant", "Not Significant")
        res$new_column <- rownames(res)
        res$diffexpressed <- ifelse(res$log2FoldChange > 0, "UP", ifelse(res$log2FoldChange < 0, "DOWN", "NO_CHANGE"))
        
        # Generate gene list for GSEA
        organism = "org.Hs.eg.db"
        original_gene_list <- res$log2FoldChange
        names(original_gene_list) <- res$new_column
        gene_list <<- na.omit(original_gene_list)
        gene_list = sort(gene_list, decreasing = TRUE)
        
        # Perform GO enrichment analysis
        gse <<- gseGO(geneList = gene_list, 
                      ont = "ALL", 
                      keyType = "SYMBOL", 
                      minGSSize = 3, 
                      maxGSSize = 800, 
                      pvalueCutoff = 0.05, 
                      verbose = TRUE, 
                      OrgDb = organism, 
                      pAdjustMethod = "none")
        
        # Store inverted results for GO terms in reactive values
        searchValues$df_inverted <- gse@result %>% separate_rows(core_enrichment, sep = "/")
        
        # Update GO term and description choices in UI
        updateSelectInput(session, "GO_search", choices = unique(searchValues$df_inverted$ID))
        updateSelectInput(session, "description_search", choices = unique(searchValues$df_inverted$Description))
        
        # Reactive filtering of results based on user input
        filteredRes <- reactive({
          data <- as.data.frame(res)
          
          # Apply gene search filter
          if (searchValues$gene_search != "") {
            genes <- strsplit(searchValues$gene_search, ";")[[1]]
            genes <- trimws(genes)
            data <- data %>%
              filter(rowSums(sapply(genes, function(gene) grepl(gene, new_column, ignore.case = TRUE))) > 0)
          }
          
          # Apply GO term filter
          if (searchValues$GO_search != "All") {
            selected_genes <- searchValues$df_inverted %>%
              filter(ID == searchValues$GO_search) %>%
              pull(core_enrichment)
            data <- data %>%
              filter(new_column %in% selected_genes)
          }
          
          # Apply description search filter
          if (searchValues$description_search != "") {
            selected_genes <- searchValues$df_inverted %>%
              filter(Description == searchValues$description_search) %>%
              pull(core_enrichment)
            data <- data %>%
              filter(new_column %in% selected_genes)
          }
          
          data
        })
        
        # Render volcano plot based on filtered results
        output$volcanoPlot <- renderPlotly({
          data_res <- filteredRes()
          p_value <- input$pvalue #need to add slides here
          log2fc <- input$log2fc #need to add slider here
          
          p <- ggplot(data_res, aes(x = log2FoldChange, y = pvalue_log10, 
                                    text = paste("Gene:", new_column, "<br>Log2 Fold Change:", log2FoldChange, 
                                                 "<br>P-value:", pvalue, "<br>Significance:", significance, 
                                                 "<br>Differentially Expressed:", diffexpressed, "<br>-log10 Values:", pvalue_log10))) +
            geom_point(aes(color = log2FoldChange, shape = diffexpressed)) +
            geom_hline(yintercept = -log10(p_value), linetype = "dotted", color = "red") +
            geom_vline(xintercept = c(-log2fc, log2fc), linetype = "dotted", color = "darkblue") +
            xlim(-5, 5) +
            xlab("Log2 Fold Change") + 
            ylab("-log10(P-value)") +
            ggtitle("Volcano Plot") +
            scale_color_gradient2(low = "green", mid = "pink", high = "blue", midpoint = 0, 
                                  name = "Log2 Fold Change")  # Add a custom color scale for the color legends
          
          ggplotly(p, tooltip = "text")
        })
      })
      
      # Update search criteria based on user actions
      observeEvent(input$search_gene, {
        searchValues$gene_search <- input$gene_search
        searchValues$GO_search <- "All"
        searchValues$description_search <- ""
      })
      
      observeEvent(input$search_GO_term, {
        searchValues$GO_search <- input$GO_search
        searchValues$gene_search <- ""
        searchValues$description_search <- ""
      })
      
      observeEvent(input$search_description, {
        searchValues$description_search <- input$description_search
        searchValues$GO_search <- "All"
        searchValues$gene_search <- ""
      })
      
      observeEvent(input$visualize_gse, {
        library(ggplot2)
        library(ggtext)
        library(gridExtra)
        
        output$dotPlotTitle <- renderPlot({
          txt <- "Dot Plot with 10 Pathways"
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) +
            theme_void()
          print(title_plot)
        })
        
        output$dotPlot <- renderPlot({
          dot_plot <- dotplot(gse, showCategory = 10)
          print(dot_plot)
        })
        
        output$conceptNetworkTitle <- renderPlot({
          txt <- "Gene Concept Network"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$conceptNetwork <- renderPlot({
          gsex <- setReadable(gse, 'org.Hs.eg.db', 'ENTREZID')
          geneList <- gse@geneList
          
          p1 <- cnetplot(gsex, foldChange = geneList, max.overlaps = 100)
          p2 <- cnetplot(gsex, categorySize = "pvalue", foldChange = geneList, max.overlaps = 100)
          p3 <- cnetplot(gsex, foldChange = geneList, circular = TRUE, colorEdge = TRUE, max.overlaps = 100)
          
          maingene_plot <- cowplot::plot_grid(p1, p2, p3, ncol = 3, labels = LETTERS[1:3], rel_widths = c(.8, .8, 1.2))
          print(maingene_plot)
          
        })
        
        output$heatMapTitle <- renderPlot({
          txt <- "Heatmap-Like Functional Classification"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$heatMap <- renderPlot({
          
          gsex <- setReadable(gse, 'org.Hs.eg.db', 'ENTREZID')
          geneList <- gse@geneList
          
          p1 <- heatplot(gsex, showCategory=5)
          p2 <- heatplot(gsex, foldChange=geneList, showCategory=5)
          mainheatmap_plot <- cowplot::plot_grid(p1, p2, ncol=1, labels=LETTERS[1:2])
          
          print(mainheatmap_plot)
        })
        
        output$upsetPlotTitle <- renderPlot({
          txt <- "UpSet Plot"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$upsetPlot <- renderPlot({
          gse_df <- gse@result
          top_terms <- gse_df %>% arrange(pvalue) %>% head(10)
          top_gene_sets <- strsplit(top_terms$core_enrichment, "/")
          gene_sets_list <- lapply(top_gene_sets, function(x) unique(trimws(x)))
          
          gene_sets_df <- fromList(setNames(gene_sets_list, top_terms$ID))
          
          # Create the UpSet plot
          upset_plot <- upset(gene_sets_df, 
                              sets = names(gene_sets_df), 
                              main.bar.color = "steelblue",
                              sets.bar.color = "darkred", 
                              order.by = "freq", 
                              matrix.color = "gray",
                              keep.order = TRUE)
          
          print(upset_plot)
        })
        
        output$pubmedPathwayPlotTitle <- renderPlot({
          txt <- "PubMed Pathway Enrichment"
          
          title_plot <- ggplot() +
            geom_textbox(
              aes(x = 0, y = 0, label = txt),
              size = 18 / .pt,
              width = unit(6, "inches")
            ) + 
            theme_void()
          
          print(title_plot)
        })
        
        output$pubmedPathwayPlot <- renderPlot({
          results <- data.frame(Term = character(), Year = integer(), Count = integer(), stringsAsFactors = FALSE)
          
          terms <- tail(gse$Description, n = 10)
          
          results <- data.frame()
          titles_2024 <- data.frame()
          
          for (term in terms) {
            for (year in 2014:2024) {
              query <- paste(term, "[Title/Abstract] AND", year, "[PDAT]")
              
              # Count results for each term and year
              search_results <- entrez_search(db = "pubmed", term = query, retmax = 0)
              results <- rbind(results, data.frame(Term = term, Year = year, Count = search_results$count))
              
              # If the year is 2024, retrieve the first 10 article titles
              if (year == 2024) {
                search_results_2024 <- entrez_search(db = "pubmed", term = query, retmax = 10)
                if (search_results_2024$count > 0) {
                  article_ids <- search_results_2024$ids
                  articles <- entrez_fetch(db = "pubmed", id = article_ids, rettype = "abstract", retmode = "text")
                  titles <- sapply(strsplit(articles, "\n"), function(x) x[1])
                  titles_2024 <- reactive({rbind(titles_2024, data.frame(Term = term, Title = titles, stringsAsFactors = FALSE))})
                }
              }
            }
          }
          
          total_counts <- results %>%
            group_by(Year) %>%
            summarize(Total_Count = sum(Count), .groups = 'drop')
          
          results <- results %>%
            left_join(total_counts, by = "Year")
          
          results <- results %>%
            mutate(Ratio = Count / Total_Count)
          
          print(results)
          
          mainpubmed_plot <- ggplot(results, aes(x = Year, y = Ratio, color = Term)) +
            geom_line() +
            geom_point(size = 3, shape = 20, fill = "white", stroke = 1) +  # Bolded dots
            scale_x_continuous(limits = c(2013, 2025), breaks = seq(2013, 2025, by = 2.5)) +  # 2.5-year breaks
            labs(title = "Publication Ratio for Enriched Terms", x = "Year", y = "Publication Ratio") +
            theme_minimal()
          
          print(mainpubmed_plot)
        })
      })
      
    } else {
      showModal(modalDialog(
        title = "Access Denied",
        "Incorrect password. Please try again.",
        easyClose = TRUE,
        footer = NULL
      ))
    }
  })
}

shinyApp(ui = ui, server = server)
```