Targeted disruption of the murine large nuclear KIAA1440/Ints1 protein causes growth arrest in early blastocyst stage embryos and eventual apoptotic cell death

Abstract

The KIAA1440 protein contains no significant domains that allow for a prediction of its function, despite the fact that it is an extremely large protein comprising 2222 amino acids. In our current study, we show that the developing KIAA1440 −/− mouse embryo in a pure ICR background arrests its growth at the early blastocyst stage, whereas the majority of the KIAA1440 −/− embryos of mixed genetic backgrounds do not progress beyond the morula stage, approximately 0.5 days earlier. KIAA1440 −/− embryos exhibited no abnormal localization of E-cadherin or β-catenin and no obvious compaction abnormalities at the morula stage. In addition, E3.5 KIAA1440 −/− embryos are not viable even in in vitro cultures. Both TUNEL and FAM-caspase-3/7 assays performed on these embryos consistently showed that E3.5 KIAA1440 −/− embryos had activated caspase-3/7, which then induced an apoptotic response predominantly within the inner cell mass of the blastocyst. Moreover, qRT-PCR analysis showed that KIAA1440 −/− embryos had increased levels of the unprocessed, primary U2 snRNA transcript but decreased levels of the mature U2 snRNA transcript compared to heterozygotes. The impaired processing of U2 snRNA and the predominantly nuclear localization of KIAA1440 protein is also very consistent with recently reported data showing that it is the largest subunit of the integrator complex, which mediates U1 and U2 snRNA 3′-end processing. Large nuclear KIAA1440/Ints1 is thus suggested to play non-redundant roles in the cell such as the formation of a scaffold for the assembly of the integrator complex.