Abstract
ABSTRACTThe mouse t haplotype, a variant 20 cM genomic region on Chromosome 17, harbors 16 embryonic control genes identified by recessive lethal mutations isolated from wild mouse populations. Due to technical constraints so far only one of these, the tw5 lethal, has been cloned and molecularly characterized. Here we report the molecular isolation of the tw18 lethal. Embryos carrying the tw18 lethal die from major gastrulation defects commencing with primitive streak formation at E6.5. We have used transcriptome and marker gene analyses to describe the molecular etiology of the tw18 phenotype. We show that both WNT and Nodal signal transduction are impaired in the mutant epiblast, causing embryonic patterning defects and failure of primitive streak and mesoderm formation. By using a candidate gene approach, gene knockout by homologous recombination and genetic rescue, we have identified the gene causing the tw18 phenotype as Ppp2r1a, encoding the PP2A scaffolding subunit PR65alpha. Our work highlights the importance of phosphatase 2A in embryonic patterning, primitive streak formation, gastrulation, and mesoderm formation downstream of WNT and Nodal signaling.
Highlights
Between 6-25% of individuals in wild house mouse populations carry a t haplotype, a variant form of the t complex spanning over 20 cM (∼40 Mbp) of Chromosome 17 (Chr 17) (Carroll et al, 2004; reviewed in Silver, 1985; Sugimoto, 2014)
Nodal and WNT signal transduction are impaired in homozygous tw18 embryos Prior to this study, tw18/tw18 mutant embryos have not been characterized at the molecular level
Embryos were dissected in two parts: the embryo proper consisting of epiblast and surrounding visceral endoderm (VE), which was used for RNA extraction, and the extraembryonic tissue, required for genotyping
Summary
Between 6-25% of individuals in wild house mouse populations carry a t haplotype, a variant form of the t complex spanning over 20 cM (∼40 Mbp) of Chromosome 17 (Chr 17) (Carroll et al, 2004; reviewed in Silver, 1985; Sugimoto, 2014) This region comprises many important developmental regulators, among them Pou5f1 (Oct4), Nkx, Axin and Brachyury (T ), as well as 16 recessive lethal mutations, known as t lethals, which are encoded by different t haplotypes and affect early embryonic development (Klein et al, 1984; Bennett, 1975; Sugimoto, 2014).
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