Calcium-dependent transcriptional profiles of human pancreatic islet cells reveal functional diversity in islet subpopulations

Aims/hypothesis Pancreatic islets depend on cytosolic calcium to trigger the secretion of glucoregulatory hormones and regulate the transcription of genes important for the response to stimuli. To date, there has not been an attempt to profile calcium-regulated gene expression in all islet cell types. Our aim was to construct a large single-cell transcriptomic dataset from human islets exposed to conditions that would acutely induce or inhibit intracellular calcium signalling, while preserving biological heterogeneity. Methods We exposed intact human islets from three donors to the following conditions: (1) 2.8 mM glucose; (2) 25 mM glucose and 40 mM KCl to maximally stimulate calcium signalling; and (3) 25 mM glucose, 40 mM KCl and 5 mM EGTA (calcium chelator) to inhibit calcium signalling, for 1 hour. We sequenced 43,909 cells from all islet cell types, and further subsetted the cells to form an endocrine cell-specific dataset of 32,486 cells expressing INS , GCG , SST or PPY . We compared transcriptomes across conditions to determine the differentially expressed calcium-regulated genes in each endocrine cell type, and in each endocrine cell subcluster of alpha and beta cells. Results Based on the number of calcium-regulated genes, we found that each alpha and beta cell cluster had a different magnitude of calcium response. We also showed that a polyhormonal cluster expressing INS, GCG , and SST is defined by calcium-regulated genes specific to this cluster. Finally, we identified the gene PCDH7 from the beta cell clusters that had the highest number of calcium-regulated genes, and showed that cells expressing cell surface PCDH7 protein have enhanced glucose-stimulated insulin secretory function. Conclusions Here we use our single-cell dataset to show that human islets have cell-type-specific calcium-regulated gene expression profiles, some of them specific to subpopulations. In our dataset, we identify PCDH7 as a novel marker of beta cells having an increased number of calcium-regulated genes and enhanced insulin secretory function. Data availability A searchable and user-friendly format of the data in this study, specifically designed for rapid mining of single-cell RNA sequencing data, is available at . The raw data files are available at NCBI Gene Expression Omnibus (GSE196715). ### Competing Interest Statement The authors have declared no competing interest. * α : Alpha β : Beta δ : Delta CAMK : Calmodulin-dependent protein kinase CaN : Calcineurin CREB : cAMP response element-binding protein DEG : Differentially expressed genes GCG : Glucagon GSIS : Glucose-stimulated insulin secretion IEG : Immediate early genes INS : Insulin KRBH : Krebs-Ringer Bicarbonate HEPES NFAT : Nuclear factor of activated T cells OxPhos : Oxidative phosphorylation PCDH7 : Protocadherin 7 PP : Pancreatic polypeptide qPCR : Quantitative PCR scRNA-seq : Single-cell RNA sequencing SST : Somatostatin TCA : Tricarboxylic acid UMAP : Uniform Manifold Approximation and Projection