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. 2018 Jun 22;7(7):65. doi: 10.3390/cells7070065

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© 2018 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).

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Sorbitol does not interfere with phosphorylation of MK2 targets. (A) U2OS cells were treated with anisomycin or sorbitol (1 h) in the indicated combinations. Cells were immunostained with antibodies against CEP131 and counterstained for nuclear content with DAPI. PCNT; Pericentrin. Scale bars, 10 μm. (B) Quantification of (A). At least 100 cells were scored per condition in 3 independent experiments. Bars indicate the mean +/− SEM. p-values were calculated from a one-way ANOVA using Dunnett’s correction for multiple testing. ****, p < 0.0001, n.s.; not significant. CS; centriolar satellites. (C) U2OS cells were pretreated with an MK2 inhibitor (MK2i—1 h) and incubated in the presence of sorbitol and anisomycin as indicated. Lysates were analysed by immunoblotting with antibodies against phospho-p38, total p38, MK2, NOGO-B, phospho-HSP27, phospho-SRF and p150 (loading control). (D) Lysates from cells treated as in (c) were incubated with recombinant GST or GST-14-3-3 and subjected to GST pulldown (PD). Pulldown material and whole cell extracts (WCE) were analysed by immunoblotting with antibodies against CEP131, MK2 and GST.

Sorbitol does not interfere with phosphorylation of MK2 targets. (A) U2OS cells were treated with anisomycin or sorbitol (1 h) in the indicated combinations. Cells were immunostained with antibodies against CEP131 and counterstained for nuclear content with DAPI. PCNT; Pericentrin. Scale bars, 10 μm. (B) Quantification of (A). At least 100 cells were scored per condition in 3 independent experiments. Bars indicate the mean +/− SEM. p-values were calculated from a one-way ANOVA using Dunnett’s correction for multiple testing. ****, p < 0.0001, n.s.; not significant. CS; centriolar satellites. (C) U2OS cells were pretreated with an MK2 inhibitor (MK2i—1 h) and incubated in the presence of sorbitol and anisomycin as indicated. Lysates were analysed by immunoblotting with antibodies against phospho-p38, total p38, MK2, NOGO-B, phospho-HSP27, phospho-SRF and p150 (loading control). (D) Lysates from cells treated as in (c) were incubated with recombinant GST or GST-14-3-3 and subjected to GST pulldown (PD). Pulldown material and whole cell extracts (WCE) were analysed by immunoblotting with antibodies against CEP131, MK2 and GST.