2217-LB: Single-Cell Analysis Reveals Pbx4 as a Key Regulator of Pancreatic β-Cell Proliferation in Pregnant Mice

Introduction and Objective: During pregnancy, the pancreatic β-cell mass expands to meet the increased metabolic demands, and it subsequently regresses in the postpartum period as maternal glucose homeostasis returns to normal. This study aims to investigate the regulatory factors driving β-cell expansion during pregnancy in mice. Methods: Islets were isolated from non-pregnant, pregnant (day 14.5), and postpartum (day 4) mice. Single-cell RNA sequencing and bioinformatics identified proliferative phase β-cells. SCENIC analysis was employed to identify the transcription factors most active in proliferating β-cells. In vitro functional assays were conducted in mouse pancreatic β-cell lines to validate the transcription factor Pbx4 drives the proliferation of pancreatic β-cells by regulating the expression of the cell cycle protein Cdk1. Results: Our single-cell transcriptomic analysis identified five functionally distinct β-cell subpopulations, with a particular focus on the proliferative population marked by Mki67. Using the SCENIC method, we identified specific transcription factors regulating this population, among which Pbx4 exhibited the strongest transcriptional activity and specificity. Overexpression of Pbx4 in mouse β-cell lines significantly upregulated Cdk1 expression, and chromatin immunoprecipitation assays further confirmed that Pbx4 directly binds to the promoter region of the Cdk1 gene. The dual-luciferase reporter assay demonstrated that overexpression of Pbx4 significantly upregulated the transcriptional activity of Cdk1, indicating that Pbx4 regulates Cdk1 to promote β-cell proliferation. Conclusion: Our data indicates that the transcription factor Pbx4 is a key driver of β-cell expansion during pregnancy, regulating the transcriptional activity of the essential cell cycle factor Cdk1 to promote pancreatic β-cell proliferation. Disclosure S. Tian: None. J. Wang: None. J. Li: None. Y. Li: None. X. Li: None. W.W. Li: None. H. Zhang: None.