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The mutants showed a lethal phenotype similar to the previously reported mua-3 mutants, confirming that the lethality is due to a mutation in mua-3 (Bercher et al. 2001).
The lethality is due to defects in the endoderm derivatives, as well as in trophoblast cell proliferation [10], [11].
This gob-1 lethality is completely suppressed when the upstream trehalose-6-phosphate synthase genes are deleted, indicating that the lethality is due to toxic build-up of the intermediate trehalose-6-phosphate [63].
We have obtained significant amount of data supporting that the lethality is due to an abrogated differentiation of mESCs, which is involved in placenta development (Kim et al., manuscript in review).
To determine whether the homozygous lethality is due to the defects in gametes or embryos, reciprocal crosses were performed between WT and Ospie1-1/+, and between WT Ospie1-2/+.
Homozygous Tek-Cre is expected to occur with 25% frequency and result in complete lethality before E7.5, however, in CKO × CKO matings more than 40% of embryos die at this stage, suggesting that the remaining ∼15% on top of the Tek-Cre-induced lethality is due to the deletion of the Ate1-floxed allele (denoted Δ in the tables and in Figure 6).
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It was not clear whether lethality was due to absence of VPg3 or to absence of the proteolytic cleavage site between VPg3 and 3C [11].
To test the possibility that this embryonic lethality was due to the inability of CKO females to carry a healthy pregnancy to term (due to angiogenic defects that could interfere with normal blood supply to the fetuses), we transferred pre-implantation embryos from CKO × CKO matings into wild type females and analyzed the lethality of the resulting progeny.
We therefore concluded that the lethality was due to Scn4a allelic non-complementation.
In all cases, viable Top2 m/ Top2 Df35; P[Top2-w +]/+ adults were obtained (Table S3), confirming that lethality was due to mutations in Top2.
On the basis of these analyses, a model has emerged wherein NMD-mutant phenotypes, including lethality, are due to misregulation of specific native targets (Hwang and Maquat 2011; Palacios 2013).
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