Transcriptomic analysis of P0 mouse lung lymphatic endothelial cells

Fluid clearance mediated by lymphatic vessels is known to be essential for lung inflation and gas exchange function during the transition from prenatal to postnatal life, yet the molecular mechanisms that regulate lymphatic function remain unclear. Here, we profiled the molecular features of lymphatic endothelial cells (LECs) in embryonic and postnatal day (P) 0 lungs by single-cell RNA-seq analysis. We identified that the expression of c-JUN is transiently upregulated in P0 LECs. Conditional knockout of Jun in LECs impairs the opening of lung lymphatic vessels at birth, leading to fluid retention in the lungs and neonatal death. We further demonstrated that increased mechanical pressure induces the expression of c-JUN in LECs. c-JUN regulates the opening of lymphatic vessels by modulating remodeling of the actin cytoskeleton in LECs. Our study established the essential regulatory function of c-JUN-mediated transcriptional responses in facilitating lung lymphatic fluid clearance at birth. Overall design: Control and Jun knockout LECs from Prox1-GFP mouse lungs were isolated at P0 by FACS. LECs isolated from three replicates of each group were identically mixed and then processed to RNA amplification following the protocol of Arcturus RiboAmp HS PLUS RNA ampli?cation kit before RNA sequencing. Datasets were generated named as "Jun-Ctrl", and "Jun-cKO".