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. 2022 Jun 10;298(7):102121. doi: 10.1016/j.jbc.2022.102121

Figure 9.

Figure 9

RasGRP3 partially mediates PKCα-induced antiproliferative signaling.A, loss of RasGRP3 significantly inhibits PKCα-induced activation of ERK and upregulation of p21Cip1. IEC-18 cells were transfected with nontargeting siRNA (NT) or ON-TARGETplus SMARTpool siRNA targeting RasGRP3 (Dharmacon) as indicated. After 48 h, cells were treated with 100 nM PMA for 15 min (i) or 2 h (ii) and analyzed for the indicated proteins by Western blotting. iii, densitometric quantification of p21Cip1, cyclin D1, and Id1 expression (normalized to β-actin) from three independent experiments. ∗p = 0.03. iv, cells were treated as in (A(i)) except that the siRNA targeting RasGRP3 was from Invitrogen (Silencer Select predesigned siRNA Rasgrp3). Data are from single immunoblots; the vertical line indicates rearrangement of the blot for clarity. B, IEC-18 cells were transfected with nontargeting siRNA (NT) or siRNA targeting RasGRP3 (ON-TARGETplus SMARTpool, Dharmacon). After 24 h, cells were treated with 100 nM PMA for 6 h, as indicated, and the percentage of cells in S-phase was assessed by flow cytometry. ∗∗p = 0.003, n = 4. PMA, phorbol 12-myristate 13-acetate.