Mutation in the Trapα/Ssr1 Gene, Encoding Translocon-Associated Protein α, Results in Outflow Tract Morphogenetic Defects

Abstract

Translocon-associated protein complex (TRAP) is thought to be required for efficient protein-specific translocation across the endoplasmic reticulum membrane. We created a mutation in the Trapα gene that leads to the synthesis of a truncated TRAPα protein fused to ShBle-β-galactosidase. Analysis of Trapα cDNAs reveals that among three different messenger RNAs expressed in the mouse, one of them encodes a slightly larger protein that differs in its C-terminal end. This mRNA, specific for skeletal muscle and heart, is only expressed after birth. Homozygous Trapα mutant pups die at birth, likely as a result of severe cardiac defects. Indeed, the septation of the proximal part of the outflow tract is absent, resulting in a double-outlet right ventricle. Studies of protein secretion in transfected embryonic fibroblasts reveal that the TRAP complex does not function properly in homozygous mutant cells and confirm, in vivo, the involvement of TRAP in substrate-specific translocation. Our results provide the first in vivo demonstration that a member of the TRAP complex plays a crucial role in mammalian heart development and suggest that TRAPα could be involved in translocation of factors necessary for maturation of endocardial cushions.

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We acknowledge Tom A. Rapoport for the gift of the C-terminal TRAPα antibody and Claire Soudais for the IFN-γ cDNA. We are very grateful to Robert Kelly, Stéphane Zaffran, and Fanny Bajolle, for precious advice and helpful discussions. We thank Laurence Fiette and Yves Pierre for technical advice. The monoclonal antibodies 40-1A and MF20, developed by J. R. Sanes and D. A. Fischman, respectively, were obtained from the DSHB, University of Iowa, Iowa City. We thank M. Buckingham, R. Kelly, and J.-L. Popot for critical reading of the manuscript.

This work was supported by the Pasteur Institute and CNRS.