PI3K-AKT signaling stabilizes a set of KSRP-interacting mRNAs and increases their expression

<p><b>Copyright information:</b></p><p>Taken from "Identification of a set of KSRP target transcripts upregulated by PI3K-AKT signaling"</p><p>http://www.biomedcentral.com/1471-2199/8/28</p><p>BMC Molecular Biology 2007;8():28-28.</p><p>Published online 16 Apr 2007</p><p>PMCID:PMC1858702.</p><p></p> (A) Either mock-αT3-1 or αT3-1-myrAKT1 cells were lysed and total extracts were immunoprecipitated (Ip) with either anti-AKT antibody or control IgG (cIgG). Pellets were incubated (20 min at 30°C) with histone 2B (H2B) in kinase buffer in the presence of γ[P]ATP under gentle shaking. Labeled proteins were separated by SDS-PAGE and detected by autoradiography. (B) Expression of KSRP-interacting mRNAs and β2-MG (control transcript), monitored by RT-PCR, in either mock-αT3-1 or αT3-1-myrAKT1 cells. (C) Semi quantitative RT-PCR analysis of both KSRP-interacting mRNAs and β2-MG (control transcript) in either mock-αT3-1 (red lines) or αT3-1-myrAKT1 (blue lines). Total RNA was isolated at the indicated times after addition of Actinomycin D. The amount of each transcript was quantitated by densitometry and plotted using a linear regression program. The values shown are averages (± SEM) of three independent experiments performed in duplicates. A quantitation of the transcripts' t(1/2) is presented in Additional file

KSRP associates with AUF1p45 and hnRNPA1 in cytoplasmic extracts of aT3-1 cells

<p><b>Copyright information:</b></p><p>Taken from "Identification of a set of KSRP target transcripts upregulated by PI3K-AKT signaling"</p><p>http://www.biomedcentral.com/1471-2199/8/28</p><p>BMC Molecular Biology 2007;8():28-28.</p><p>Published online 16 Apr 2007</p><p>PMCID:PMC1858702.</p><p></p> (A) S100 extracts from αT3-1 cells were subjected to gel filtration chromatography on a Superose 6 column. Aliquots of the eluted fractions were analyzed by Western blotting using the indicated antibodies. (B) RNase A-treated S100 extracts from αT3-1 cells were immunoprecipitated with preimmune (lane 2) or anti-KSRP (lane 3) sera and analyzed by immunoblotting with either anti-AUF1 (top) or anti-HnRNPA1 (bottom) antibodies. The arrows mark the position of either AUF1p45 or hnRNPA1, while the asterisk marks a anti-AUF1 cross-reacting band. (C) GST-pulldown of either endogenous AUF1p45 (top) or endogenous hnRNPA1 (bottom) from S100 extracts of αT3-1 cells using either control GST or GST-KSRP. Proteins were analyzed by immunoblotting using the indicated antibodies. The arrows mark the position of either AUF1p45 or hnRNPA1