ABSTRACT

Diabetes mellitus is caused by either insulin resistance or insulin deficiency. The pancreatic β cells are the primary producers of insulin. Large-scale CRISPR screens combined with single-cell RNA sequencing (scRNA-seq) on β cells has identified novel insulin regulators and revealed the presence of a highly complex inner network. Here, we performed pooled CRISPR delivery with single-cell transcriptome analysis on the MIN6 cell line, a pancreatic β-cell line. We have presented the scRNA-seq readout and demonstrated that the MIN6 cell line might develop genetic heterogeneity with increasing passage number based on GO and KEGG pathway analysis. Both computational and biological analyses revealed that the function of MIN6 cell lines could be divided into five clusters, including endocrine cells, basal cells, epithelial cells, and neuroendocrine cells. The fifth cluster was different from the other four clusters due to the differentially expressed insulin transcription and was called the lncRNA-enriched cluster. The experiments also confirmed that uncharacterized lncRNAs GM26917 and Cenpw were associated with insulin transcription. This study provides information that can be used to systematically characterize insulin regulator genes and other genes that control protein folding and vesicle trafficking.

摘要

糖尿病是由胰岛素抵抗或胰岛素缺乏引起的。胰腺β细胞是胰岛素的主要生产者。大规模 CRISPR 筛选与 β 细胞上的单细胞 RNA 测序 (scRNA-seq) 相结合，已经确定了新的胰岛素调节剂，并揭示了高度复杂的内部网络的存在。在这里，我们对胰腺 β 细胞系 MIN6 细胞系进行了合并的 CRISPR 递送和单细胞转录组分析。我们已经展示了 scRNA-seq 读数，并证明 MIN6 细胞系可能会根据 GO 和 KEGG 通路分析随着传代数的增加而发展出遗传异质性。计算和生物学分析均表明，MIN6细胞系的功能可分为五个簇，包括内分泌细胞、基底细胞、上皮细胞和神经内分泌细胞。由于胰岛素转录的差异表达，第五个簇与其他四个簇不同，被称为富含 lncRNA 的簇。实验还证实了未表征的 lncRNA GM26917 和Cenpw与胰岛素转录有关。该研究提供的信息可用于系统地表征胰岛素调节基因和其他控制蛋白质折叠和囊泡运输的基因。