The RAS Guanyl Nucleotide-releasing Protein RasGRP1 Is Involved in Lymphatic Development in Zebrafish

Fulong Wang,Heng Huang,Lin Wang,Rui Zhang,Ting Jin,Yan Chen,Zhenzhen Wang,Yi Pan

doi:10.1074/jbc.m112.418202

Abstract

The molecular basis of the lymphatic development remains largely unknown. Using zebrafish as a model, we discovered a novel role for the Ras guanine-releasing protein 1 (RasGRP1), a protein involved in Ras activation in lymphangiogenesis. Secondary lymphatic sprouts from the posterior cardinal vein give rise to thoracic duct which is the first lymphatic vessel in zebrafish. Knockdown of rasgrp1 by injecting morpholino in zebrafish embryos impaired formation of thoracic duct accompanied by pericardial and truck edema, whereas blood vessel development of the embryos was largely unaffected. In rasgrp1-knockdown embryos, the number of sprouts producing the string of parachordal lymphangioblast cells was reduced. Meanwhile the total number of the secondary sprouts was not changed. As a result, the number of venous intersegmental vessels was increased, whereas the number of lymphatic vessel was reduced at a later stage. The lymphatic developmental defects caused by rasgrp1 knockdown could be rescued by ectopic expression of a constitutively active HRas. Further analysis revealed that RasGRP1 knockdown could synergize with flt4/vegfr3 knockdown to induce defects in lymphangiogenesis. Taken together, this finding demonstrates a critical role for RasGRP1 in lymphatic development in zebrafish.

Highlights

Ras guanine-releasing protein 1 (RasGRP1) is a protein involved in Ras activation, but its physiological function is largely unknown
Using zebrafish as a model, we discovered a novel role for the Ras guanine-releasing protein 1 (RasGRP1), a protein involved in Ras activation in lymphangiogenesis
The signal of rasgrp1 expression was mostly lost in dorsal aorta (DA) but still persistent in posterior cardinal vein (PCV) and Pl regions at 36 hpf (Fig. 1A), at a time when the lymphatic precursors begin to migrate toward the horizontal myoseptum region from the PCV [9]

Summary

Background

RasGRP1 is a protein involved in Ras activation, but its physiological function is largely unknown. Results: Knockdown of RasGRP1 impairs development of the lymphatic system in zebrafish embryos, shown as defective thoracic duct formation, pericardial and truck edema, and altered sprouts of lymphatic vessels. Using zebrafish as a model, we discovered a novel role for the Ras guanine-releasing protein 1 (RasGRP1), a protein involved in Ras activation in lymphangiogenesis. Knockdown of rasgrp by injecting morpholino in zebrafish embryos impaired formation of thoracic duct accompanied by pericardial and truck edema, whereas blood vessel development of the embryos was largely unaffected. Further analysis revealed that RasGRP1 knockdown could synergize with flt4/vegfr knockdown to induce defects in lymphangiogenesis Taken together, this finding demonstrates a critical role for RasGRP1 in lymphatic development in zebrafish. We analyzed the potential function of RasGRP1 during zebrafish development and discovered a novel function of RasGRP1 in early lymphangiogenesis

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION