Fig. 3.

SRRM2 deficiency destabilizes PQBP1. a Interactome database (String ver.10.5) predicted the interaction of PQBP1 and SC35 with SRRM2. b Immunoprecipitation revealed the interaction between endogenous SRRM2 and PQBP1 or between SRRM2 and SC35. c SiRNA-mediated SRRM2 knockdown reduced PQBP1 in primary cortical neurons prepared from E15 mouse embryos (white arrow). d Expression of EGFP-wild-type or 1068A-mutant SRRM2 recovered PQBP1, whereas EGFP-1068D-mutant or 1068E-mutant SRRM2 did not recover PQBP1 sufficiently. e Western blot analysis confirmed SRRM2 knockdown by siRNA and the accompanying decrease of PQBP1 in HeLa cells. f Western blot analysis of whole cerebral cortex lysates from 5xFAD mice and sibling mice (B6/SJL) at each age revealed that PQBP1 decreased during aging and the decrease occurred more rapidly in 5xFAD mice. g Immunohistochemistry was performed with parietal lobe tissues of human AD patients with PS1 Met146Leu mutation (N = 3) and non-neurological disease controls (N = 3). Reduction of SRRM2 and PQBP1 was obvious in AD patients, and representative images of cortical neurons are shown. h Western blot analysis was performed with the same AD and non-neurological disease patients. The right graphs show the quantitative analysis of β-actin-corrected PQBP1 and SRRM2 signals (N = 3, *p < 0.05, Student’s t-test). i Human iPS cells carrying the APP KM670/671NL heterozygous or homozygous mutation generated by Crispr-cas9 based genome editing were used to show that SRRM2 and PQBP1 are lower in the nuclei and that pSer1068-SRRM2 is increased in the cytoplasm of differentiated neurons. Cytoplasmic levels of TCP1α in human iPS cells carrying the APP KM670/671NL mutation and in human normal iPS cell-derived neurons were similar. j Western blot analysis of neurons differentiated from human iPS cells carrying the APP KM670/671NL heterozygous or homozygous mutation. Right panels show quantification of the band intensities