Nup62 is involved in reduction of nuclear SREBPs by excess ORP8.

Abstract

<p><b>A</b> HuH7 cells were transfected with empty vector plasmid (Mock), <i>ORP8</i> cDNA (ORP8), non-targeting siRNA (siNT) or siNup62 as indicated, followed by preparation of total protein specimens and nuclear fractions, and Western blot analysis thereof with antibodies against SREBP-1 and SREBP-2. Nuclear SREBPs (nSREBPs) decreased in cells overexpressing ORP8 (the top panels), while no change in nSREBPs was seen in cells transfected with siNup62 (the two bottom panels). <b>B</b> HuH7 cells were transfected with combinations of siRNAs and plasmids as indicated at the top. Nuclear fractions Western blotted for SREBP-1 and SREBP-2, and total protein specimens (40 µg/lane) blotted for Nup62, ORP8, and β-actin are shown at the bottom. <b>C,D</b> Quantification of the relative levels of nSREBP-1 and nSREBP-2 after the indicated combined transfections. <b>E</b> Effect of the combined transfections on the mRNA levels of SREBP target genes (identified at the bottom); qPCR analysis. The results represent mean ± s.e.m. (n = 4, *p<0.05, **p<0.01, t-test; difference to values of the siNT and Mock-transfected control, which was set at 1).</p