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The Rab2A GTPase promotes breast cancer stem cells and tumorigenesis via Erk signaling activation.

Luo ML, Gong C, Chen CH, Hu H, Huang P, Zheng M, Yao Y, Wei S, Wulf G, Lieberman J, Zhou XZ, Song E, Lu KP - Cell Rep (2015)

Bottom Line: Mechanistically, Rab2A directly interacts with and prevents dephosphorylation/inactivation of Erk1/2 by the MKP3 phosphatase, resulting in Zeb1 upregulation and β-catenin nuclear translocation.Finally, Rab2A overexpression correlates with poor clinical outcome in breast cancer patients.Thus, Pin1/Rab2A/Erk drives BCSC expansion and tumorigenicity, suggesting potential drug targets.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine and Cancer Research Institute, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA; Breast Tumor Center, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120, China.

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Rab2A Interacts with and Prevents Inactivation of Erk1/2 by MKP3(A) Overexpressed Rab2A and its Q58H mutant co-localized with p-Erk1/2. Stable HMLE cells were serum starved and then treated with 10 ng/ml EGF for 5 min. Scale bar represents 10 μm.(B) Rab2A and its Q58H mutant co-localized with ERGIC53. Scale bar represents 20 μm.(C) Reciprocal coIP of endogenous Rab2A with Erk1/2. Lysates of HMLE cells were immunoprecipitated with Rab2A or Erk1/2 antibodies, followed by western blot for Rab2A and Erk1/2, respectively.(D) Rab2A immunoprecipitated with total Erk1/2 and p-Erk1/2 in HEK293 cells co-transfected with Flag-Rab2A and constitutive activated MEK1 (AcMEK1).(E) The consensus Erk docking motifs were found in Rab2A and several other Erk binding partners. + and φ represent basic and hydrophobic amino acids, respectively. X represents any amino acids.(F) Mutations in the Erk docking motif in Rab2A impaired its binding to Erk1/2.(G) Rab2A and MKP3 competed to bind Erk1/2. Lysates of 293T cells transfected with decreasing doses of myc-MKP3 and a constant dose of Flag-Rab2A were immunoprecipitated with M2 (Flag) antibody, followed by western blot for Erk1/2 and Flag-Rab2A.(H) Rab2A competed with MKP3 and kept Erk1/2 in the phosphorylated status. 293T cells were transfected to express Rab2A, MKP3, and AcMEK1, which induced p-Erk1/2 in serum-starved cells and was largely reversed by Myc-MKP3 expression, whereas Flag-Rab2A expression dose-dependently restored Erk1/2 phosphorylation.See also Figures S4 and S5.
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Figure 4: Rab2A Interacts with and Prevents Inactivation of Erk1/2 by MKP3(A) Overexpressed Rab2A and its Q58H mutant co-localized with p-Erk1/2. Stable HMLE cells were serum starved and then treated with 10 ng/ml EGF for 5 min. Scale bar represents 10 μm.(B) Rab2A and its Q58H mutant co-localized with ERGIC53. Scale bar represents 20 μm.(C) Reciprocal coIP of endogenous Rab2A with Erk1/2. Lysates of HMLE cells were immunoprecipitated with Rab2A or Erk1/2 antibodies, followed by western blot for Rab2A and Erk1/2, respectively.(D) Rab2A immunoprecipitated with total Erk1/2 and p-Erk1/2 in HEK293 cells co-transfected with Flag-Rab2A and constitutive activated MEK1 (AcMEK1).(E) The consensus Erk docking motifs were found in Rab2A and several other Erk binding partners. + and φ represent basic and hydrophobic amino acids, respectively. X represents any amino acids.(F) Mutations in the Erk docking motif in Rab2A impaired its binding to Erk1/2.(G) Rab2A and MKP3 competed to bind Erk1/2. Lysates of 293T cells transfected with decreasing doses of myc-MKP3 and a constant dose of Flag-Rab2A were immunoprecipitated with M2 (Flag) antibody, followed by western blot for Erk1/2 and Flag-Rab2A.(H) Rab2A competed with MKP3 and kept Erk1/2 in the phosphorylated status. 293T cells were transfected to express Rab2A, MKP3, and AcMEK1, which induced p-Erk1/2 in serum-starved cells and was largely reversed by Myc-MKP3 expression, whereas Flag-Rab2A expression dose-dependently restored Erk1/2 phosphorylation.See also Figures S4 and S5.

Mentions: To elucidate how Rab2A overexpression or its Q58H mutation activates Erk1/2, we first examined whether Rab2A co-localized with Erk1/2. Overexpressing WT Rab2A not only activated Erk1/2 but also surprisingly colocalized with activated Erk1/2 at the perinuclear region at 5 min (Figure 4A) and 1 hr (Figure S4A) after EGF stimulation. Overexpressing Rab2A Q58H at levels similar to the endogenous level also activated and colocalized with Erk1/2 (Figures 4A and S4A). Importantly, Rab2A and its Q58H mutant colocalized with Erk1/2 at the ERGIC, as assayed by ERGIC53 staining (Figure 4B). To examine whether Rab2A’s vesicular trafficking function is associated with Erk activation, we used brefeldin A (BFA) to block the trafficking from the ERGIC to ER. As shown previously (Hauri et al., 2000), BFA treatment damaged the ERGIC structure (Figure S4B) but did not obviously affect Erk phosphorylation (Figure S4C), suggesting that Erk1/2 activation is likely to be independent of Rab2A’s trafficking function.


The Rab2A GTPase promotes breast cancer stem cells and tumorigenesis via Erk signaling activation.

Luo ML, Gong C, Chen CH, Hu H, Huang P, Zheng M, Yao Y, Wei S, Wulf G, Lieberman J, Zhou XZ, Song E, Lu KP - Cell Rep (2015)

Rab2A Interacts with and Prevents Inactivation of Erk1/2 by MKP3(A) Overexpressed Rab2A and its Q58H mutant co-localized with p-Erk1/2. Stable HMLE cells were serum starved and then treated with 10 ng/ml EGF for 5 min. Scale bar represents 10 μm.(B) Rab2A and its Q58H mutant co-localized with ERGIC53. Scale bar represents 20 μm.(C) Reciprocal coIP of endogenous Rab2A with Erk1/2. Lysates of HMLE cells were immunoprecipitated with Rab2A or Erk1/2 antibodies, followed by western blot for Rab2A and Erk1/2, respectively.(D) Rab2A immunoprecipitated with total Erk1/2 and p-Erk1/2 in HEK293 cells co-transfected with Flag-Rab2A and constitutive activated MEK1 (AcMEK1).(E) The consensus Erk docking motifs were found in Rab2A and several other Erk binding partners. + and φ represent basic and hydrophobic amino acids, respectively. X represents any amino acids.(F) Mutations in the Erk docking motif in Rab2A impaired its binding to Erk1/2.(G) Rab2A and MKP3 competed to bind Erk1/2. Lysates of 293T cells transfected with decreasing doses of myc-MKP3 and a constant dose of Flag-Rab2A were immunoprecipitated with M2 (Flag) antibody, followed by western blot for Erk1/2 and Flag-Rab2A.(H) Rab2A competed with MKP3 and kept Erk1/2 in the phosphorylated status. 293T cells were transfected to express Rab2A, MKP3, and AcMEK1, which induced p-Erk1/2 in serum-starved cells and was largely reversed by Myc-MKP3 expression, whereas Flag-Rab2A expression dose-dependently restored Erk1/2 phosphorylation.See also Figures S4 and S5.
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Figure 4: Rab2A Interacts with and Prevents Inactivation of Erk1/2 by MKP3(A) Overexpressed Rab2A and its Q58H mutant co-localized with p-Erk1/2. Stable HMLE cells were serum starved and then treated with 10 ng/ml EGF for 5 min. Scale bar represents 10 μm.(B) Rab2A and its Q58H mutant co-localized with ERGIC53. Scale bar represents 20 μm.(C) Reciprocal coIP of endogenous Rab2A with Erk1/2. Lysates of HMLE cells were immunoprecipitated with Rab2A or Erk1/2 antibodies, followed by western blot for Rab2A and Erk1/2, respectively.(D) Rab2A immunoprecipitated with total Erk1/2 and p-Erk1/2 in HEK293 cells co-transfected with Flag-Rab2A and constitutive activated MEK1 (AcMEK1).(E) The consensus Erk docking motifs were found in Rab2A and several other Erk binding partners. + and φ represent basic and hydrophobic amino acids, respectively. X represents any amino acids.(F) Mutations in the Erk docking motif in Rab2A impaired its binding to Erk1/2.(G) Rab2A and MKP3 competed to bind Erk1/2. Lysates of 293T cells transfected with decreasing doses of myc-MKP3 and a constant dose of Flag-Rab2A were immunoprecipitated with M2 (Flag) antibody, followed by western blot for Erk1/2 and Flag-Rab2A.(H) Rab2A competed with MKP3 and kept Erk1/2 in the phosphorylated status. 293T cells were transfected to express Rab2A, MKP3, and AcMEK1, which induced p-Erk1/2 in serum-starved cells and was largely reversed by Myc-MKP3 expression, whereas Flag-Rab2A expression dose-dependently restored Erk1/2 phosphorylation.See also Figures S4 and S5.
Mentions: To elucidate how Rab2A overexpression or its Q58H mutation activates Erk1/2, we first examined whether Rab2A co-localized with Erk1/2. Overexpressing WT Rab2A not only activated Erk1/2 but also surprisingly colocalized with activated Erk1/2 at the perinuclear region at 5 min (Figure 4A) and 1 hr (Figure S4A) after EGF stimulation. Overexpressing Rab2A Q58H at levels similar to the endogenous level also activated and colocalized with Erk1/2 (Figures 4A and S4A). Importantly, Rab2A and its Q58H mutant colocalized with Erk1/2 at the ERGIC, as assayed by ERGIC53 staining (Figure 4B). To examine whether Rab2A’s vesicular trafficking function is associated with Erk activation, we used brefeldin A (BFA) to block the trafficking from the ERGIC to ER. As shown previously (Hauri et al., 2000), BFA treatment damaged the ERGIC structure (Figure S4B) but did not obviously affect Erk phosphorylation (Figure S4C), suggesting that Erk1/2 activation is likely to be independent of Rab2A’s trafficking function.

Bottom Line: Mechanistically, Rab2A directly interacts with and prevents dephosphorylation/inactivation of Erk1/2 by the MKP3 phosphatase, resulting in Zeb1 upregulation and β-catenin nuclear translocation.Finally, Rab2A overexpression correlates with poor clinical outcome in breast cancer patients.Thus, Pin1/Rab2A/Erk drives BCSC expansion and tumorigenicity, suggesting potential drug targets.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine and Cancer Research Institute, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA; Breast Tumor Center, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120, China.

Show MeSH
Related in: MedlinePlus