shRNA-mediated repression of deptor increases mTORC1 and mTORC2 activity, resulting in collagen I (α2) expression similar to TGFβ.

<p>(A–C and E) Human proximal tubular epithelial cells were transfected with two independent shRNAs against deptor (Deptor sh1 and Deptor sh2). The transfected cells were incubated with 2 ng/ml TGFβ for 24 hours. The cell lysates were immunoblotted with indicated antibodies. Quantifications of panels A–C are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0109608#pone.0109608.s004" target="_blank">Figs. S4A</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0109608#pone.0109608.s004" target="_blank">S4C</a>. (D) Human proximal tubular epithelial cells were transfected with two independent shRNAs against deptor (Deptor sh1 and Deptor sh2). The transfected cells were incubated with 2 ng/ml TGFβ for 24 hours. Total RNA was prepared and used for real time RT-PCR to detect collagen I (α2) mRNA as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0109608#s2" target="_blank">Materials and Methods</a>. Mean ± SE of triplicate measurements is shown. *p<0.01 vs control. Expression of Deptor in parallel samples is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0109608#pone.0109608.s005" target="_blank">Fig. S5A</a>. Quantification of Fig. 2E is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0109608#pone.0109608.s005" target="_blank">Fig. S5B</a>.