ClusterFoldSimilarity: comparing cell-groups from independent single-cell experiments
29 October 2024
Package
ClusterFoldSimilarity 1.2.0
1 Installation
The package can be installed using bioconductor install manager:
if (!require("BiocManager", quietly = TRUE))
install.packages("BiocManager")
BiocManager::install("ClusterFoldSimilarity")library(ClusterFoldSimilarity)2 Introduction
Comparing single-cell data across different datasets, samples and batches has demonstrated to be challenging. ClusterFoldSimilarity aims to solve the complexity of comparing different single-cell datasets by computing similarity scores between clusters (or user-defined groups) from any number of independent single-cell experiments, including different species and sequencing technologies. It accomplishes this by identifying analogous fold-change patterns across cell groups that share a common set of features (such as genes). Additionally, it selects and reports the top important features that have contributed to the observed similarity, serving as a tool for feature selection.
The output is a table that contains the similarity values for all the combinations of cluster-pairs from the independent datasets. ClusterFoldSimilarity also includes various plotting utilities to enhance the interpretability of the similarity scores.
2.0.1 Cross-species analysis and sequencing technologies (e.g.: Human vs Mouse, ATAC-Seq vs RNA-Seq)
ClusterFoldSimilarity is able to compare any number of independent experiments, including different organisms, making it useful for matching cell populations across different organisms, and thus, useful for inter-species analysis. Additionally, it can be used with single-cell RNA-Seq data, single-cell ATAC-Seq data, or more broadly, with continuous numerical data that shows changes in feature abundance across a set of common features between different groups.
2.0.2 Compatibility
It can be easily integrated on any existing single-cell analysis pipeline, and it is compatible with the most used single-cell objects: Seurat and SingleCellExperiment.
Parallel computing is available through the option parallel=TRUE which make use of BiocParallel.
3 Using ClusterFoldSimilarity to find similar clusters/cell-groups across datasets
Typically, ClusterFoldSimilarity will receive as input either a list of two or more Seurat or SingleCellExperiment objects.
ClusterFoldSimilarity will obtain the raw count data from these objects ( GetAssayData(assay, slot = "counts") in the case of Seurat, or counts() for SingleCellExperiment object), and group or cluster label information (using Idents() function from Seurat, or colLabels() for SingleCellExperiment ).
For the sake of illustration, we will employ the scRNAseq package, which contains numerous individual-cell datasets ready for download and encompassing samples from both human and mouse origins. In this example, we specifically utilize 2 human single-cell datasets obtained from the pancreas.
library(Seurat)
library(scRNAseq)
library(dplyr)
# Human pancreatic single cell data 1
pancreasMuraro <- scRNAseq::MuraroPancreasData(ensembl=FALSE)
pancreasMuraro <- pancreasMuraro[,rownames(colData(pancreasMuraro)[!is.na(colData(pancreasMuraro)$label),])]
colData(pancreasMuraro)$cell.type <- colData(pancreasMuraro)$label
rownames(pancreasMuraro) <- make.names(unlist(lapply(strsplit(rownames(pancreasMuraro), split="__"), function(x)x[[1]])), unique = TRUE)
singlecell1Seurat <- CreateSeuratObject(counts=counts(pancreasMuraro), meta.data=as.data.frame(colData(pancreasMuraro)))| Var1 | Freq |
|---|---|
| acinar | 219 |
| alpha | 812 |
| beta | 448 |
| delta | 193 |
| duct | 245 |
| endothelial | 21 |
| epsilon | 3 |
| mesenchymal | 80 |
| pp | 101 |
| unclear | 4 |
# Human pancreatic single cell data 2
pancreasBaron <- scRNAseq::BaronPancreasData(which="human", ensembl=FALSE)
colData(pancreasBaron)$cell.type <- colData(pancreasBaron)$label
rownames(pancreasBaron) <- make.names(rownames(pancreasBaron), unique = TRUE)
singlecell2Seurat <- CreateSeuratObject(counts=counts(pancreasBaron), meta.data=as.data.frame(colData(pancreasBaron)))| Var1 | Freq |
|---|---|
| acinar | 958 |
| activated_stellate | 284 |
| alpha | 2326 |
| beta | 2525 |
| delta | 601 |
| ductal | 1077 |
| endothelial | 252 |
| epsilon | 18 |
| gamma | 255 |
| macrophage | 55 |
| mast | 25 |
| quiescent_stellate | 173 |
| schwann | 13 |
| t_cell | 7 |
As we want to perform clustering analysis for later comparison of these cluster groups using ClusterFoldSimilarity, we first need to normalize and identify variable features for each dataset independently.
Note: these steps should be done tailored to each independent dataset, here we apply the same parameters for the sake of simplicity:
# Create a list with the unprocessed single-cell datasets
singlecellObjectList <- list(singlecell1Seurat, singlecell2Seurat)
# Apply the same processing to each dataset and return a list of single-cell analysis
singlecellObjectList <- lapply(X=singlecellObjectList, FUN=function(scObject){
scObject <- NormalizeData(scObject)
scObject <- FindVariableFeatures(scObject, selection.method="vst", nfeatures=2000)
scObject <- ScaleData(scObject, features=VariableFeatures(scObject))
scObject <- RunPCA(scObject, features=VariableFeatures(object=scObject))
scObject <- FindNeighbors(scObject, dims=seq(16))
scObject <- FindClusters(scObject, resolution=0.4)
})Once we have all of our single-cell datasets analyzed independently, we can compute the similarity values. clusterFoldSimilarity() takes as arguments:
scList: a list of single-cell objects (mandatory) either of classSeurator of classSingleCellExperiment.sampleNames: vector with names for each of the datasets. If not set the datasets will be named in the given order as: 1, 2, …, N.topN: the top n most similar clusters/groups to report for each cluster/group (default:1, the top most similar cluster). If set toInfit will return the values from all the possible cluster-pairs.topNFeatures: the top n features (e.g.: genes) that contribute to the observed similarity between the pair of clusters (default:1, the top contributing gene). If a negative number, the tool will report the n most dissimilar features.nSubsampling: number of subsamplings (1/3 of cells on each iteration) at group level for calculating the fold-changes (default:15). At start, the tool will report a message with the recommended number of subsamplings for the given data (average n of subsamplings needed to observe all cells).parallel: whether to use parallel computing with multiple threads or not (default:FALSE). If we want to use a specific single-cell experiment for annotation (from which we know a ground-truth label, e.g. cell type, cell cycle, treatment… etc.), we can use that label to directly compare the single-cell datasets.
Here we will use the annotated pancreas cell-type labels from the dataset 1 to illustrate how to match clusters to cell-types using a reference dataset:
# Assign cell-type annotated from the original study to the cell labels:
Idents(singlecellObjectList[[1]]) <- factor(singlecellObjectList[[1]][[]][, "cell.type"])
library(ClusterFoldSimilarity)
similarityTable <- clusterFoldSimilarity(scList=singlecellObjectList,
sampleNames=c("human", "humanNA"),
topN=1,
nSubsampling=24)
| similarityValue | w | datasetL | clusterL | datasetR | clusterR | topFeatureConserved | featureScore | |
|---|---|---|---|---|---|---|---|---|
| acinar | 50.38169 | 6.82 | human | acinar | humanNA | 2 | REG1A | 50.427213 |
| alpha | 34.65354 | 6.51 | human | alpha | humanNA | 3 | GCG | 34.122804 |
| beta | 33.92137 | 6.59 | human | beta | humanNA | 4 | INS | 30.337947 |
| delta | 29.38112 | 6.29 | human | delta | humanNA | 4 | PCSK1 | 7.573988 |
| duct | 33.42910 | 6.42 | human | duct | humanNA | 5 | SPP1 | 13.135512 |
| endothelial | 45.73913 | 7.01 | human | endothelial | humanNA | 9 | PLVAP | 18.286168 |
A data.frame with the results is returned containing:
similarityValue: The top similarity value calculated between datasetL:clusterL and datasetR.w: Weight associated with the similarity score value.datasetL: Dataset left, the dataset/sample which has been used to be compared.clusterL: Cluster left, the cluster source from datasetL which has been compared.datasetR: Dataset right, the dataset/sample used for comparison against datasetL.clusterR: Cluster right, the cluster target from datasetR which is being compared with the clusterL from datasetL.topFeatureConserved: The features (e.g.: genes, peaks…) that most contributed to the similarity between clusterL & clusterR.featureScore: The similarity score contribution for the specific topFeatureConserved (e.g.: genes, peaks…).
By default, clusterFoldSimilarity() will plot a graph network that visualizes the connections between the clusters from the different datasets using the similarity table that has been obtained. The arrows point in the direction of the similarity (datasetL:clusterL -> datasetR:clusterR); it can be useful for identifying relationships between groups of clusters and cell-populations that tend to be more similar. The graph plot can also be obtained by using the function plotClustersGraph() from this package, using as input the similarity table.
In this example, as we have information regarding cell-type labels, we can check how the cell types match by calculating the most abundant cell type on each of the similar clusters:
typeCount <- singlecellObjectList[[2]][[]] %>%
group_by(seurat_clusters) %>%
count(cell.type) %>%
arrange(desc(n), .by_group = TRUE) %>%
filter(row_number()==1)| seurat_clusters | cell.type | n | matched.type |
|---|---|---|---|
| 0 | alpha | 1308 | alpha |
| 1 | beta | 1022 | beta |
| 2 | acinar | 927 | acinar |
| 3 | alpha | 967 | alpha |
| 4 | beta | 958 | beta |
| 5 | ductal | 817 | duct |
| 6 | delta | 586 | delta |
| 7 | beta | 494 | beta |
| 8 | activated_stellate | 283 | mesenchymal |
| 9 | endothelial | 250 | endothelial |
| 10 | gamma | 211 | epsilon |
| 11 | ductal | 174 | acinar |
| 12 | macrophage | 55 | unclear |
| 13 | ductal | 77 | unclear |
3.1 Analyzing graph communities to identify super-groups of similar cell populations
To easily analyze and identify the similarities between the different datasets and cell-groups, we can find the communities that constitute the directed graph (cluster the nodes based on the graph´s closely-related elements).
We can make so by using the function findCommunitiesSimmilarity() from the ClusterFoldSimilarity package. It uses the InfoMap algorithm to find the best fitting communities. We just need the similarity table obtained from clusterFoldSimilarity() as explained on the previous section, the function will plot the graph with the communities and return a data frame containing the community that each sample & cluster/group belongs to.
cellCommunities <- findCommunitiesSimmilarity(similarityTable = similarityTable)
| sample | group | community | |
|---|---|---|---|
| human_group_acinar | human | acinar | 1 |
| human_group_alpha | human | alpha | 2 |
| human_group_beta | human | beta | 3 |
| human_group_delta | human | delta | 3 |
| human_group_duct | human | duct | 4 |
| human_group_endothelial | human | endothelial | 5 |
3.2 Retrieving the top-n similarities
If we suspect that clusters could be related with more than one cluster of other datasets, we can retrieve the top n similarities for each cluster:
# Retrieve the top 3 similar cluster for each of the clusters:
similarityTable3Top <- clusterFoldSimilarity(scList=singlecellObjectList,
topN=3,
sampleNames=c("human", "humanNA"),
nSubsampling=24)
| similarityValue | w | datasetL | clusterL | datasetR | clusterR | topFeatureConserved | featureScore | |
|---|---|---|---|---|---|---|---|---|
| acinar.3 | 50.00145 | 6.81 | human | acinar | humanNA | 2 | REG1A | 49.999513 |
| acinar.12 | 42.70464 | 6.65 | human | acinar | humanNA | 11 | REG3A | 40.070842 |
| acinar.6 | 27.00327 | 5.95 | human | acinar | humanNA | 5 | SERPINA3 | 14.315774 |
| alpha.4 | 34.50439 | 6.51 | human | alpha | humanNA | 3 | GCG | 33.757464 |
| alpha.1 | 31.33150 | 6.33 | human | alpha | humanNA | 0 | GCG | 29.997441 |
| alpha.5 | 29.70890 | 6.35 | human | alpha | humanNA | 4 | CPE | 7.782267 |
3.3 Obtaining the top-n feature markers
If we are interested on the features that contribute the most to the similarity, we can retrieve the top n features:
# Retrieve the top 5 features that contribute the most to the similarity between each pair of clusters:
similarityTable5TopFeatures <- clusterFoldSimilarity(scList=singlecellObjectList,
topNFeatures=5,
nSubsampling=24)
| similarityValue | w | datasetL | clusterL | datasetR | clusterR | topFeatureConserved | featureScore | |
|---|---|---|---|---|---|---|---|---|
| acinar.11 | 50.59540 | 6.84 | 1 | acinar | 2 | 2 | REG1A | 51.04858 |
| acinar.12 | 50.59540 | 6.84 | 1 | acinar | 2 | 2 | CTRB2 | 47.57484 |
| acinar.13 | 50.59540 | 6.84 | 1 | acinar | 2 | 2 | REG1B | 46.55425 |
| acinar.14 | 50.59540 | 6.84 | 1 | acinar | 2 | 2 | PRSS1 | 45.54733 |
| acinar.15 | 50.59540 | 6.84 | 1 | acinar | 2 | 2 | CELA3A | 41.50131 |
| alpha.16 | 34.52617 | 6.50 | 1 | alpha | 2 | 3 | GCG | 33.61875 |
| alpha.17 | 34.52617 | 6.50 | 1 | alpha | 2 | 3 | TTR | 22.43977 |
| alpha.18 | 34.52617 | 6.50 | 1 | alpha | 2 | 3 | CHGB | 13.33365 |
| alpha.19 | 34.52617 | 6.50 | 1 | alpha | 2 | 3 | PCSK2 | 11.68422 |
| alpha.20 | 34.52617 | 6.50 | 1 | alpha | 2 | 3 | TM4SF4 | 11.57276 |
3.4 Retrieving all the similarity values and plotting a similarity heatmap
Sometimes it is useful to retrieve all the similarity values for downstream analysis (e.g. identify more than one cluster that is similar to a cluster of interest, finding the most dissimilar clusters, etc). To obtain all the values, we need to specify topN=Inf.
By default, clusterFoldSimilarity creates a heatmap plot with the computed similarity values (from the perspective of the first dataset found on scList; to modify this plot see the following section). The top 2 similarities for each group within dataset 1 (heatmap row-wise) are highlighted with colored borders.
similarityTableAllValues <- clusterFoldSimilarity(scList=singlecellObjectList,
sampleNames=c("human", "humanNA"),
topN=Inf)
dim(similarityTableAllValues)## [1] 280 8For downstream analysis of the similarities, it can be convenient to create a matrix with all the scores from the comparison of two datasets:
library(dplyr)
dataset1 <- "human"
dataset2 <- "humanNA"
similarityTable2 <- similarityTableAllValues %>%
filter(datasetL == dataset1 & datasetR == dataset2) %>%
arrange(desc(as.numeric(clusterL)), as.numeric(clusterR))
cls <- unique(similarityTable2$clusterL)
cls2 <- unique(similarityTable2$clusterR)
similarityMatrixAll <- t(matrix(similarityTable2$similarityValue, ncol=length(unique(similarityTable2$clusterL))))
rownames(similarityMatrixAll) <- cls
colnames(similarityMatrixAll) <- cls2| 0 | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 | 13 | |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| acinar | -10.56 | 31.63 | 23.51 | 23.94 | -9.63 | -12.46 | 0.39 | -11.83 | 23.42 | -15.30 | -11.05 | 27.54 | 31.27 | 26.93 |
| alpha | -11.10 | -11.83 | -7.44 | -9.94 | 22.68 | -16.51 | 50.04 | -11.42 | -11.75 | -12.91 | 27.40 | -7.00 | -26.26 | 16.02 |
| beta | 7.82 | -16.39 | -16.08 | 34.82 | 25.82 | 26.04 | -13.54 | -15.79 | 24.00 | -17.11 | 26.30 | -15.08 | -17.21 | 29.94 |
| delta | 33.72 | 29.42 | -15.33 | -14.81 | 18.10 | -15.09 | 24.77 | -15.12 | 26.46 | -15.52 | -15.01 | -13.63 | 33.40 | 15.63 |
| duct | -10.12 | 15.29 | -14.02 | 29.98 | -15.15 | 26.00 | 25.36 | 28.94 | -12.27 | -13.34 | 20.92 | -14.01 | 23.99 | -12.13 |
| endothelial | -18.84 | 28.12 | 32.98 | 28.61 | -15.43 | -14.51 | 23.54 | -15.93 | 23.74 | -11.68 | -7.63 | -17.63 | -15.68 | -14.81 |
| epsilon | 13.63 | 33.85 | -14.68 | 48.55 | -15.65 | -6.25 | -10.02 | -17.64 | -15.47 | -14.82 | 10.31 | 45.71 | -10.17 | 32.65 |
| mesenchymal | -15.68 | 20.14 | -17.78 | 25.53 | 22.13 | 24.26 | -13.87 | -13.53 | 30.10 | -15.97 | 26.91 | -8.09 | 42.36 | -19.38 |
| pp | -19.15 | -17.71 | 29.31 | -1.01 | -15.72 | -6.95 | -12.08 | 30.60 | -10.35 | -13.69 | -13.27 | -11.84 | 14.49 | 24.68 |
| unclear | 26.43 | 22.03 | -13.59 | 31.11 | 22.19 | -20.62 | -19.66 | -17.55 | 30.26 | 17.34 | 23.79 | 3.76 | -17.56 | 41.91 |
4 Using ClusterFoldSimilarity across species and numerous datasets:
ClusterFoldSimilarity can compare any number of independent studies, including different organisms, making it useful for inter-species analysis. Also, it can be used on different sequencing data technologies: e.g.: compare single-cell ATAC-Seq VS RNA-seq.
In this example, we are going to add a pancreas single-cell dataset from Mouse to the 2 existing ones from Human that we have processed in the previous steps.
# Mouse pancreatic single cell data
pancreasBaronMM <- scRNAseq::BaronPancreasData(which="mouse", ensembl=FALSE)| Var1 | Freq |
|---|---|
| B_cell | 10 |
| T_cell | 7 |
| activated_stellate | 14 |
| alpha | 191 |
| beta | 894 |
| delta | 218 |
| ductal | 275 |
| endothelial | 139 |
| gamma | 41 |
| immune_other | 8 |
| macrophage | 36 |
| quiescent_stellate | 47 |
| schwann | 6 |
colData(pancreasBaronMM)$cell.type <- colData(pancreasBaronMM)$label
# Translate mouse gene ids to human ids
# *for the sake of simplicity we are going to transform to uppercase all mouse gene names
rownames(pancreasBaronMM) <- make.names(toupper(rownames(pancreasBaronMM)), unique=TRUE)
# Create seurat object
singlecell3Seurat <- CreateSeuratObject(counts=counts(pancreasBaronMM), meta.data=as.data.frame(colData(pancreasBaronMM)))
# We append the single-cell object to our list
singlecellObjectList[[3]] <- singlecell3SeuratNow, we process the new single-cell dataset from mouse, and we calculate the similarity scores between the 3 independent datasets.
scObject <- singlecellObjectList[[3]]
scObject <- NormalizeData(scObject)
scObject <- FindVariableFeatures(scObject, selection.method="vst", nfeatures=2000)
scObject <- ScaleData(scObject, features=VariableFeatures(scObject))
scObject <- RunPCA(scObject, features=VariableFeatures(object=scObject))
scObject <- FindNeighbors(scObject, dims=seq(16))
scObject <- FindClusters(scObject, resolution=0.4)
singlecellObjectList[[3]] <- scObjectThis time we will make use of the option parallel=TRUE. We can set the specific number of CPUs to use using BiocParallel::register()
# We use the cell labels as a second reference, but we can also use the cluster labels if our interest is to match clusters
Idents(singlecellObjectList[[3]]) <- factor(singlecellObjectList[[3]][[]][,"cell.type"])
# We subset the most variable genes in each experiment
singlecellObjectListVariable <- lapply(singlecellObjectList, function(x){x[VariableFeatures(x),]})
# Setting the number of CPUs with BiocParallel:
BiocParallel::register(BPPARAM = BiocParallel::MulticoreParam(workers = 6))
similarityTableHumanMouse <- clusterFoldSimilarity(scList=singlecellObjectListVariable,
sampleNames=c("human", "humanNA", "mouse"),
topN=1,
nSubsampling=24,
parallel=TRUE)
We can compute and visualize with a heatmap all the similarities for each cluster/group of cells from the 3 datasets using topN=Inf. Additionally, we can use the function similarityHeatmap() from this package to plot the heatmap with the datasets in a different order, or just plot the 2 datasets we are interested in. The top 2 similarities are highlighted to help visualizing the best matching groups.
similarityTableHumanMouseAll <- clusterFoldSimilarity(scList=singlecellObjectListVariable,
sampleNames=c("human", "humanNA", "mouse"),
topN=Inf,
nSubsampling=24,
parallel=TRUE)
As the similarity values might not be symmetric (e.g. a cluster A from D1 showing the top similarity to B from D2, might not be the top similar cluster to B from D2), we can select which dataset to plot in the Y-axis:
ClusterFoldSimilarity::similarityHeatmap(similarityTable=similarityTableHumanMouseAll, mainDataset="humanNA")
Additionally, we can turn-off the highlight using highlightTop=FALSE
# Turn-off the highlighting:
ClusterFoldSimilarity::similarityHeatmap(similarityTable=similarityTableHumanMouseAll, mainDataset="humanNA", highlightTop=FALSE)
5 Similarity score calculation
ClusterFoldSimilarity does not need to integrate the data, or apply any batch correction techniques across the datasets that we aim to analyze, which makes it less prone to data-loss or noise. The similarity value is based on the fold-changes between clusters/groups of cells defined by the user. These fold-changes from different independent datasets are first computed using a Bayesian approach, we calculate this fold-change distribution using a permutation analysis that shrink the fold-changes with no biological meaning. These differences in abundance are then combined using a pairwise dot product approach, after adding these feature contributions and applying a fold-change concordance weight, a similarity value is obtained for each of the clusters of each of the datasets present.
Session information
sessionInfo()## R version 4.4.1 (2024-06-14)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.1 LTS
##
## Matrix products: default
## BLAS: /home/biocbuild/bbs-3.20-bioc/R/lib/libRblas.so
## LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.12.0
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## locale:
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## [7] LC_PAPER=en_US.UTF-8 LC_NAME=C
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## [11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
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## time zone: America/New_York
## tzcode source: system (glibc)
##
## attached base packages:
## [1] stats4 stats graphics grDevices utils datasets methods
## [8] base
##
## other attached packages:
## [1] dplyr_1.1.4 scRNAseq_2.19.1
## [3] SingleCellExperiment_1.28.0 SummarizedExperiment_1.36.0
## [5] Biobase_2.66.0 GenomicRanges_1.58.0
## [7] GenomeInfoDb_1.42.0 IRanges_2.40.0
## [9] S4Vectors_0.44.0 BiocGenerics_0.52.0
## [11] MatrixGenerics_1.18.0 matrixStats_1.4.1
## [13] Seurat_5.1.0 SeuratObject_5.0.2
## [15] sp_2.1-4 ClusterFoldSimilarity_1.2.0
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## [118] UCSC.utils_1.2.0 shiny_1.9.1 farver_2.1.2
## [121] jquerylib_0.1.4 zoo_1.8-12 jsonlite_1.8.9
## [124] BiocParallel_1.40.0 RCurl_1.98-1.16 magrittr_2.0.3
## [127] kableExtra_1.4.0 GenomeInfoDbData_1.2.13 dotCall64_1.2
## [130] patchwork_1.3.0 Rhdf5lib_1.28.0 munsell_0.5.1
## [133] Rcpp_1.0.13 reticulate_1.39.0 stringi_1.8.4
## [136] alabaster.schemas_1.6.0 zlibbioc_1.52.0 MASS_7.3-61
## [139] AnnotationHub_3.14.0 plyr_1.8.9 parallel_4.4.1
## [142] listenv_0.9.1 ggrepel_0.9.6 deldir_2.0-4
## [145] Biostrings_2.74.0 splines_4.4.1 tensor_1.5
## [148] igraph_2.1.1 spatstat.geom_3.3-3 RcppHNSW_0.6.0
## [151] reshape2_1.4.4 BiocVersion_3.20.0 XML_3.99-0.17
## [154] evaluate_1.0.1 BiocManager_1.30.25 httpuv_1.6.15
## [157] RANN_2.6.2 tidyr_1.3.1 purrr_1.0.2
## [160] polyclip_1.10-7 future_1.34.0 scattermore_1.2
## [163] alabaster.sce_1.6.0 ggplot2_3.5.1 xtable_1.8-4
## [166] restfulr_0.0.15 AnnotationFilter_1.30.0 RSpectra_0.16-2
## [169] later_1.3.2 viridisLite_0.4.2 tibble_3.2.1
## [172] memoise_2.0.1 AnnotationDbi_1.68.0 GenomicAlignments_1.42.0
## [175] cluster_2.1.6 globals_0.16.3