The endothelial-enriched LINC00607 mediates alternative splicing of the VEGF receptor FLT1

F. Lam (Frankfurt am Main)¹, J. Oo (Frankfurt am Main)¹, T. Warwick (Frankfurt am Main)¹, S. Günther (Bad Nauheim)², J. Izquierdo Ponce (Frankfurt am Main)¹, R. Dechend (Berlin)³, F. Louwen (Frankfurt am Main)⁴, M. Schulz (Frankfurt am Main)⁵, I. Wittig (Frankfurt am Main)⁶, R. P. Brandes (Frankfurt am Main)¹, M. S. Leisegang (Frankfurt am Main)^1
1Universitätsklinikum Frankfurt Institut für Kardiovaskuläre Physiologie Frankfurt am Main, Deutschland; ²Max-Planck-Institut für Herz- und Lungenforschung Bad Nauheim, Deutschland; ³HELIOS Klinikum Berlin-Buch Klinik und Poliklinik für Kardiologie und Nephrologie Berlin, Deutschland; ⁴Universitätsklinik Frankfurt Prenatal Medicine and Obstetrics Frankfurt am Main, Deutschland; ⁵Goethe Universität Frankfurt am Main Institute of Cardiovascular Regeneration Frankfurt am Main, Deutschland; ⁶Institute for Cardiovascular Physiology Functional Proteomics Group Frankfurt am Main, Deutschland

Background: Endothelial lncRNAs, their vascular functions and contributions for disease development are only poorly understood. We have previously shown that the highly expressed and endothelial-enriched lncRNA LINC00607 maintains endothelial homeostasis especially in response to VEGF-A signaling through binding to the chromatin remodeling protein BRG1. Here we investigated the role of LINC00607 in alternative splicing regulation on the example of the VEGF receptor FLT1.

Result: Differential splicing analysis of RNA-Seq in HUVEC revealed many alternative splice events upon loss of LINC00607. Among them are highly significantly spliced transcripts such as the VEGF receptor FLT1 (fms related receptor tyrosine kinase 1). The differential splicing of FLT1 produces the anti-angiogenic soluble isoform of FLT1 (sFLT1). The anti-angiogenic capacity of sFLT1, as seen in LINC00607 KO cells in a spheroid outgrowth assay, could be rescued through addition of a sFLT1 neutralizing antibody. sFLT1 is a clinical marker in the pregnancy disorder preeclampsia (PE). We identified that LINC00607 is highly expressed in the extravillous trophoblasts and syncyciotrophoblast of the placenta. RT-qPCR, RNAscope and single-cell RNA-sequencing data of healthy versus PE placenta showed differential expression of LINC00607 in PE. LINC00607 expression is increased upon DMOG stimulation in organotypic cell culture of term placenta. Interestingly, the LINC00607 binding protein BRG1 was found to bind to the FLT1 intron nearby the alternatively spliced critical exons. LINC00607 might qualify to be an additional marker for the onset and manifestation of the pregnancy disorder.

Conclusions:  LINC00607 maintains constitutive FLT1 splicing in endothelial cells and the placenta and has the potential to serve as a biomarker in PE.