A CANDIDATE GENE FOR THE AMNIONLESS GASTRULATION STAGE MOUSE MUTATIONENCODES A TRAF-RELATED PROTEIN

We report the identification of a new recessive prenatal lethal insert ional mutation, amnionless (amn). amn mutant embryos first appear abno rmal during the Early Streak stage, between E6.5 and E7.0, when they i nitiate mesoderm production. Subsequently, the amn mutants become deve lopmentally arrested between the Mid and Late Streak stages of gastrul ation and they die and are resorbed between E9.5 and E10.5. While extr aembryonic structures, including the chorion, yolk sac blood islands, and allantois appear to develop normally, the small embryonic ectoderm remains undifferentiated and generates no amnion. In addition, the em bryonic mesoderm that is produced does not become organized into node, notochord, and somites and there is no morphological evidence of neur al induction. Interspecific backcross and fluorescence in situ hybridi zation analyses map the transgene insertion, and thus the amn mutation , to the distal region of mouse chromosome 12, which has synteny with human chromosome 14q32. A gene encoding a 7.5-kb transcript has been i dentified at a junction between the integrated transgene and host chro mosome 12 sequences that meets three criteria expected of a candidate amn gene. This gene maps to the site of transgene insertion; it is tra nscribed during gastrulation, and its expression is disrupted in amn m utant embryos. Nucleotide sequencing studies show that the 567 amino a cid protein encoded by the 7.5-kb transcript is a member of the newly defined family of putative signal transducing proteins, TRAFs, that as sociate with the cytoplasmic domains of members of the tumor necrosis factor (TNF) receptor superfamily. Thus, we have named the gene encodi ng the 7.5-kb transcript TRAFamn. TRAFamn is identical to a recently r eported protein (CD40bp, CAP-1, CRAF1, LAP1) that can bind the cytopla smic domains of CD40 and the lymphotoxin beta receptor (LT beta R), bo th of which are known members of the TNF receptor superfamily. The imp lications of these findings regarding a possible role for the TNF rece ptor superfamily during gastrulation are discussed. (C) 1996 Academic Press, Inc.