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. 2014 Oct 30;1(1):16–28. doi: 10.1016/j.ebiom.2014.10.019

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© 2014 The Authors

This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/3.0/).

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Fig. 8 — siRNA-mediated depletion of Native SYK prevents nocodazole-induced S216-phosphorylation of native CDC25C in human cells. [a–c] Anti-CDC25C(pS216), anti-CDC25C, and anti-GRP78 Western blot analysis of whole cell lysates from 293T cells treated with medium only (CON), SYK-siRNA or scr-siRNA that was used as a control. NOC treatment (0.1 μg/mL) for 24 h induced S216 phosphorylation of native CDC25C (Panel a, Lane 2 vs. Lane 1) without a change in the levels of CDC25C or GRP78 proteins (Lane 2 vs. 1, Panels b & c). SYK-siRNA abrogated NOC-induced S216 phosphorylation of CDC25C (Panel a, Lane 4 vs. Lane 3). By contrast, scr-siRNA did not block NOC-induced S216 phosphorylation of CDC25C (Panel a, Lane 6 vs. Lane 5). [d & e] Depicted are the results of a repeat analysis of SYK-siRNA effects on NOC-induced CDC25C phosphorylation on S216. Anti-CDC25C(pS216) (Panel d) and anti-cdc2/Cdk1 (Panel e) Western blot analysis of whole cell lysates from 293T cells treated with medium only (CON), SYK-siRNA or scr-siRNA that was used as a control showed that NOC treatment induced S216 phosphorylation of native CDC25C (Panel d, Lane 2 vs. Lane 1) without a change in the levels of Cdc2/Cdk1 (Panel e, Lane 2 vs. Lane 1). SYK siRNA abrogated NOC-induced S216 phosphorylation of CDC25C (Panel d, Lane 4 vs. Lane 3). By contrast, scr-siRNA did not block NOC-induced S216 phosphorylation of CDC25C (Panel d, Lane 6 vs. Lane 5). In a-e, each siRNA was used at a 50 nM concentration.