Figure 1 | Scientific Reports

Figure 1

From: VRK3-mediated nuclear localization of HSP70 prevents glutamate excitotoxicity-induced apoptosis and Aβ accumulation via enhancement of ERK phosphatase VHR activity

Figure 1

HSP70 interacts with and enhances VHR phosphatase activity.

(a) Endogenous VHR and VRK3 immunoprecipitated with an antibody against anti-FLAG in SH-SY5Y cell lysates expressing FLAG- HSP70. (b) Endogenous HSP70 immunoprecipitated with an antibody against anti-HA in glutamate-treated SH-SY5Y cell lysates expressing HA-VHR. (c) Endogenous HSP70 immunoprecipitated with endogenous VHR in glutamate-treated SH-SY5Y cell lysates. (d) FLAG-tagged HSP70 and HA-tagged VHR were mainly co-localized in the nucleus of SH-SY5Y cells treated with glutamate. Scale bar, 20 μm. (e) Domain structure of HSP70 and schematic representation of HSP70 fragments. (f) HSP70 bound to VHR with its ATPase domain and a functionally unknown region (C-terminal region). (g) A 2-fold increase in VHR phosphatase activity by HSP70. In vitro VHR phosphatase assay was performed using HSP70 with pNPP. Values are shown as mean ± standard deviation (s.d.), n = 3. Student’s t-tests, ***P < 0.001. (h) HSP70 enhanced VHR phosphatase activity in a chaperone activity-independent manner. In vitro VHR phosphatase assay was performed using HSP70 with recombinant p-ERK2. (i) Quantification of the percentage of p-ERK2 versus total ERK2 as in (h). Values are normalized to p-ERK2-only control and shown as mean ± s.d., n = 3. Student’s t-tests, *P < 0.05, **P < 0.01.

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