Hyaluronic Acid Receptor Stabilin-2 Regulates Erk Phosphorylation and Arterial - Venous Differentiation in Zebrafish

Megan S Rost,Saulius Sumanas,Vladimir Korzh

doi:10.1371/journal.pone.0088614

Megan S Rost, Saulius Sumanas + Show 1 more

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https://doi.org/10.1371/journal.pone.0088614

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Abstract

The hyaluronic acid receptor for endocytosis Stabilin-2/HARE mediates systemic clearance of multiple glycosaminoglycans from the vascular and lymphatic circulations. In addition, recent in vitro studies indicate that Stab2 can participate in signal transduction by interacting with hyaluronic acid (HA), which results in Erk phosphorylation. However, it is not known whether Stab2 function or HA-Stab2 signaling play any role in embryonic development. Here we show that Stab2 functions in a signal transduction pathway regulating arterial-venous differentiation during zebrafish embryogenesis. Stab2 morpholino knockdown embryos (morphants) display an absence of intersegmental vessels and defects in the axial vessel formation. In addition, Stab2 morphants show defects in arterial-venous differentiation including the expansion of venous marker expression. Simultaneous knockdown of Stabilin-2 and Has2, an HA synthetase, results in a synergistic effect, arguing that HA and Stab2 interact during vasculature formation. Stab2 morphants display reduced Erk phosphorylation in the arterial progenitors, which is a known transducer of VEGF signaling, previously associated with arterial-venous differentiation. In addition, VEGF signaling acts as a negative feedback loop to repress stab2 expression. These results argue that Stab2 is involved in a novel signaling pathway that plays an important role in regulating Erk phosphorylation and establishing arterial-venous identity.

Highlights

Vascular endothelial cells establish their arterial-venous identities prior to the initiation of circulation [1,2]
intersegmental vessels (ISVs) formation was unaffected by injecting a standard control morpholino at similar concentrations which argues for the specificity of the observed defects in Stab2 morphants (Fig. S1B)
Control venous flt4 expression was expanded in DAPT-treated embryos as reported previously [11] (Fig. 7I–J). These results show that Vegf but not Notch signaling negatively regulates stab2 expression. These results demonstrate that Stabilin-2 function is required for arterial-venous differentiation in the major axial vessels during zebrafish embryogenesis

Summary

Introduction

Vascular endothelial cells establish their arterial-venous identities prior to the initiation of circulation [1,2]. Multiple arterial and venous specific markers become preferentially enriched in the DA or the PCV by 24 hpf, prior to the initiation of circulation [6]. These include arterialspecific ephrin B2A (efnb2A) [6], aquaporin8a (aqp8a) [7], gridlock/hey (grl) [8] and claudin5b (cldn5b) [9] and venous specific eph receptor B4a (ephb4a) [6], fms-related tyrosine kinase 4 (flt4) [7], mannose-receptor C1 (mrc1) [10], stabilin-2 (stab2) [10] and stabilin1-like (stab1l) [11]

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