The Wnt-specific astacin proteinase HAS-7 restricts head organizer formation in Hydra

Berenice Ziegler,Sumit Kumar,Svenja Kling,Maike Fath,Benjamin Trageser,Thomas W Holstein,Walter Stöcker,Jörg Stetefeld,Moritz Mercker,Martina Schnölzer,Suat Özbek,Irene Yiallouros,Markus Hartl,Uwe Warnken,Anna Marciniak-Czochra

doi:10.1186/s12915-021-01046-9

Abstract

BackgroundThe Hydra head organizer acts as a signaling center that initiates and maintains the primary body axis in steady state polyps and during budding or regeneration. Wnt/beta-Catenin signaling functions as a primary cue controlling this process, but how Wnt ligand activity is locally restricted at the protein level is poorly understood. Here we report a proteomic analysis of Hydra head tissue leading to the identification of an astacin family proteinase as a Wnt processing factor.ResultsHydra astacin-7 (HAS-7) is expressed from gland cells as an apical-distal gradient in the body column, peaking close beneath the tentacle zone. HAS-7 siRNA knockdown abrogates HyWnt3 proteolysis in the head tissue and induces a robust double axis phenotype, which is rescued by simultaneous HyWnt3 knockdown. Accordingly, double axes are also observed in conditions of increased Wnt activity as in transgenic actin::HyWnt3 and HyDkk1/2/4 siRNA treated animals. HyWnt3-induced double axes in Xenopus embryos could be rescued by coinjection of HAS-7 mRNA. Mathematical modelling combined with experimental promotor analysis indicate an indirect regulation of HAS-7 by beta-Catenin, expanding the classical Turing-type activator-inhibitor model.ConclusionsWe show the astacin family protease HAS-7 maintains a single head organizer through proteolysis of HyWnt3. Our data suggest a negative regulatory function of Wnt processing astacin proteinases in the global patterning of the oral-aboral axis in Hydra.

Highlights

The Hydra head organizer acts as a signaling center that initiates and maintains the primary body axis in steady state polyps and during budding or regeneration
While unspecific proteolysis of HyWnt3-His was evident after 24 h incubation in Phosphate-buffered saline (PBS), the recombinant protein stayed remarkably stable in lysates of tentacles and the lower body part
In head lysate (HL) samples supplemented with broad-spectrum metalloproteinase inhibitors like EDTA and 1,10-Phenanthroline or the specific matrix metalloproteinase inhibitor Batimastat, HyWnt3His processing was completely blocked in the given time frame, indicating that metalloproteinases could be responsible for the observed activity

Summary

Introduction

The Hydra head organizer acts as a signaling center that initiates and maintains the primary body axis in steady state polyps and during budding or regeneration. Wnt/beta-Catenin signaling functions as a primary cue controlling this process, but how Wnt ligand activity is locally restricted at the protein level is poorly understood. The role of Wnt/beta-Catenin signaling in the axial patterning of cnidarians has been extensively studied in the freshwater polyp Hydra [4,5,6,7], which has a single oral-aboral body axis. A similar case for morphogen inactivation has been proposed for TIKI1, a highly conserved metalloproteinase expressed in the Xenopus organizer and shown to antagonize Wnt function by cleaving eight amino-terminal residues of Wnt3a [14]

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Results

Conclusion