CRISPR-mediated deletion of the PECAM-1 cytoplasmic domain increases receptor lateral mobility and strengthens endothelial cell junctional integrity

LIFE SCIENCES

卷:193

Article

Liao, Danying^1,2  Mei, Heng²  Hu, Yu²  Newman, Debra K.^1,3,4,6  Newman, Peter J.^1,2,3,5,6 

[1] Blood Ctr Wisconsin, Blood Res Inst, 8727 Watertown Plank Rd, Milwaukee, WI 53266 USA

[2] Huazhong Univ Sci & Technol, Tongji Med Coll, Union Hosp, Inst Hematol, Wuhan, Hubei, Peoples R China

[3] Med Coll Wisconsin, Dept Pharmacol, Milwaukee, WI 53226 USA

[4] Med Coll Wisconsin, Dept Microbiol, Milwaukee, WI 53226 USA

[5] Med Coll Wisconsin, Dept Cell Biol Neurobiol & Anat, Milwaukee, WI 53226 USA

[6] Med Coll Wisconsin, Cardiovasc Ctr, Milwaukee, WI 53226 USA

关键词: PECAM-1;    Glycosylation;    Sialic acid;    Endothelial cell;    Adhesion;    Permeability;    Vascular biology;   

DOI  :  10.1016/j.lfs.2017.11.002

来源: Elsevier

【 摘 要 】

Aims: PECAM-1 is an abundant endothelial cell surface receptor that becomes highly enriched at endothelial cell-cell junctions, where it functions to mediate leukocyte transendothelial migration, sense changes in shear and flow, and maintain the vascular permeability barrier. Homophilic interactions mediated by the PECAM-1 extracellular domain are known to be required for PECAM-1 to perform these functions; however, much less is understood about the role of its cytoplasmic domain in these processes. Main methods: CRISPR/Cas9 gene editing technology was employed to generate human endothelial cell lines that either lack PECAM-1 entirely, or express mutated PECAM-1 missing the majority of its cytoplasmic domain (Delta CD-PECAM-1). The endothelial barrier function was evaluated by Electric Cell-substrate Impedance Sensing, and molecular mobility was assessed by fluorescence recovery after photobleaching. Key findings: We found that Delta CD-PECAM-1 concentrates normally at endothelial cell junctions, but has the unexpected property of conferring increased baseline barrier resistance, as well as a more rapid rate of recovery of vascular integrity following thrombin-induced disruption of the endothelial barrier. Fluorescence recovery after photobleaching analysis revealed that Delta CD-PECAM-1 exhibits increased mobility within the plane of the plasma membrane, thus allowing it to redistribute more rapidly back to endothelial cell-cell borders to reform the vascular permeability barrier. Significance: The PECAM-1 cytoplasmic domain plays a novel role in regulating the rate and extent of vascular permeability following thrombotic or inflammatory challenge.

【 授权许可】

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