The mouse amyloid precursor-like protein 2 (APLP2) belongs to the Alzheimer peptide precursor family. A possible role in pre-implantation development had been suggested previously, and was investigated further by creating a large deletion in the genomic locus. While heterozygous mice developed normally, homozygous embryos were arrested before reaching the blastocyst stage. One-cell embryos which contained protein of maternal origin underwent a limited number of cleavages. The progressive disappearance of the protein at stages 4 and beyond correlated with the appearance of extensive cytopathological effects. Nuclear DNA contents of the arrested embryos departed widely from the normal 2-4C value, thus suggesting a role for the protein in replication and/or segregation of the embryonic genome. Embryonic mortality was not due to the untimely initiation of programmed cell death, and it occurred before the stage at which apoptotic cells normally appear. The same abnormal distribution of DNA contents was seen in primary cultures of Aplp2 +/- embryonic fibroblasts following transfection of an expression vector for Aplp2 antisense RNA with green fluorescent protein (GFP) expressed from a co-transfected construct. Daughter cells derived from a GFP-positive cell showed abnormal DNA contents both >4C and <2C, thus indicating a role for the protein in the mitotic segregation of the genome and establishment of the proper nuclear structure.