title: Laminar flow downregulates Notch activity to promote lymphatic sprouting.
abstract: The major function of the lymphatic system is to drain interstitial fluid from tissue. Functional drainage causes increased fluid flow that triggers lymphatic expansion, which is conceptually similar to hypoxia-triggered angiogenesis. Here, we have identified a mechanotransduction pathway that translates laminar flow-induced shear stress to activation of lymphatic sprouting. While low-rate laminar flow commonly induces the classic shear stress responses in blood endothelial cells and lymphatic endothelial cells (LECs), only LECs display reduced Notch activity and increased sprouting capacity. In response to flow, the plasma membrane calcium channel ORAI1 mediates calcium influx in LECs and activates calmodulin to facilitate a physical interaction between Krüppel-like factor 2 (KLF2), the major regulator of shear responses, and PROX1, the master regulator of lymphatic development. The PROX1/KLF2 complex upregulates the expression of DTX1 and DTX3L. DTX1 and DTX3L, functioning as a heterodimeric Notch E3 ligase, concertedly downregulate NOTCH1 activity and enhance lymphatic sprouting. Notably, overexpression of the calcium reporter GCaMP3 unexpectedly inhibited lymphatic sprouting, presumably by disturbing calcium signaling. Endothelial-specific knockouts of Orai1 and Klf2 also markedly impaired lymphatic sprouting. Moreover, Dtx3l loss of function led to defective lymphatic sprouting, while Dtx3l gain of function rescued impaired sprouting in Orai1 KO embryos. Together, the data reveal a molecular mechanism underlying laminar flow-induced lymphatic sprouting.
authors.forename: Dongwon
authors.initials: D
authors.lastname: Choi
authors.forename: Eunkyung
authors.initials: E
authors.lastname: Park
authors.forename: Eunson
authors.initials: E
authors.lastname: Jung
authors.forename: Young Jin
authors.initials: YJ
authors.lastname: Seong
authors.forename: Jaehyuk
authors.initials: J
authors.lastname: Yoo
authors.forename: Esak
authors.initials: E
authors.lastname: Lee
authors.forename: Mingu
authors.initials: M
authors.lastname: Hong
authors.forename: Sunju
authors.initials: S
authors.lastname: Lee
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Ishida
authors.forename: James
authors.initials: J
authors.lastname: Burford
authors.forename: Janos
authors.initials: J
authors.lastname: Peti-Peterdi
authors.forename: Ralf H
authors.initials: RH
authors.lastname: Adams
authors.forename: Sonal
authors.initials: S
authors.lastname: Srikanth
authors.forename: Yousang
authors.initials: Y
authors.lastname: Gwack
authors.forename: Christopher S
authors.initials: CS
authors.lastname: Chen
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Vogel
authors.forename: Chester J
authors.initials: CJ
authors.lastname: Koh
authors.forename: Alex K
authors.initials: AK
authors.lastname: Wong
authors.forename: Young-Kwon
authors.initials: YK
authors.lastname: Hong
mesh_terms: D000818:Animals; D001783:Blood Flow Velocity; D020013:Calcium Signaling / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation / Q000502:physiology*; D042783:Endothelial Cells / Q000166:cytology; D057809:HEK293 Cells; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000378:metabolism; D042583:Lymphangiogenesis / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D000071740:ORAI1 Protein / Q000235:genetics / Q000378:metabolism; D051881:Receptor, Notch1 / Q000096:biosynthesis* / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism; D000098551:Prospero-Related Homeobox 1 Protein
chemical_list: D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C096161:KLF2 protein, human; C498314:Klf2 protein, mouse; D051741:Kruppel-Like Transcription Factors; C497761:NOTCH1 protein, human; C497762:Notch1 protein, mouse; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; D051881:Receptor, Notch1; D025521:Tumor Suppressor Proteins; D000098551:Prospero-Related Homeobox 1 Protein; C583410:DTX1 protein, human; C475669:DTX3L protein, human; C000606502:DTX3L protein, mouse; C104035:Dtx1 protein, mouse; D044767:Ubiquitin-Protein Ligases