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VRK2 anchors KSR1-MEK1 to endoplasmic reticulum forming a macromolecular complex that compartmentalizes MAPK signaling

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Abstract

The spatial and temporal regulation of intracellular signaling is determined by the spatial and temporal organization of complexes assembled on scaffold proteins, which can be modulated by their interactions with additional proteins as well as subcellular localization. The scaffold KSR1 protein interacts with MAPK forming a complex that conveys a differential signaling in response to growth factors. The aim of this work is to determine the unknown mechanism by which VRK2A downregulates MAPK signaling. We have characterized the multiprotein complex formed by KSR1 and the Ser-Thr kinase VRK2A. VRK2A is a protein bound to the endoplasmic reticulum (ER) and retains a fraction of KSR1 complexes on the surface of this organelle. Both proteins, VRK2A and KSR1, directly interact by their respective C-terminal regions. In addition, MEK1 is also incorporated in the basal complex. MEK1 independently interacts with the CA5 region of KSR1 and with the N-terminus of VRK2A. Thus, VRK2A can form a high molecular size (600–1,000 kDa) stable complex with both MEK1 and KSR1. Knockdown of VRK2A resulted in disassembly of these high molecular size complexes. Overexpression of VRK2A increased the amount of KSR1 in the particulate fraction and prevented the incorporation of ERK1/2 into the complex after stimulation with EGF. Neither VRK2A nor KSR1 interact with the VHR, MKP1, MKP2, or MKP3 phosphatases. The KSR1 complex assembled and retained by VRK2A in the ER can have a modulatory effect on the signal mediated by MAPK, thus locally affecting the magnitude of its responses, and can explain differential responses depending on cell type.

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Abbreviations

ERK:

Extracellular signal regulated kinase (MAPK3)

JIP:

c-Jun interacting protein

JNK:

c-Jun N-terminal Kinase

KSR1:

Kinase suppressor of RAS 1

MAPK:

Mitogen-activated protein kinase

MEK1:

MAPK/ERK kinase 1 (MAP2K1)

VRK:

Vaccinia-related kinase

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Acknowledgments

I.F.F. has a JAE-CSIC predoctoral fellowship. This work was funded by grants from Ministerio de Ciencia e Innovación (SAF2010-14935 and CSD2007-0017), Junta de Castilla y León (Consejería de Educación CSI-006A11-2) and Kutxa-Fundación Inbiomed to P.A.L., and grant SAF2010-20203 from Ministerio de Ciencia e Innovación to J.L.

Conflict of interest

The authors declare they have no competing interests.

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Authors and Affiliations

  1. Experimental Therapeutics and Translational Oncology Program, Instituto de Biología Molecular y Celular del Cáncer, CSIC-Universidad de Salamanca, Campus Miguel de Unamuno, 37007, Salamanca, Spain

    Isabel F. Fernández, Sandra Blanco & Pedro A. Lazo

  2. Departamento de Biología Molecular y Bioquímica, Facultad de Ciencias, Universidad de Málaga, Malaga, Spain

    Luis G. Pérez-Rivas, Adrián A. Castillo-Dominguez & José Lozano

  3. Laboratorio de Oncología Molecular, Fundación IMABIS, Hospital Clínico Universitario Virgen de la Victoria, Malaga, Spain

    Luis G. Pérez-Rivas, Adrián A. Castillo-Dominguez & José Lozano

  4. Instituto de Investigación Biomédica de Salamanca (IBSAL), Hospital Universitario de Salamanca, Campus Miguel de Unamuno, 37007, Salamanca, Spain

    Pedro A. Lazo

Authors
  1. Isabel F. Fernández
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  2. Luis G. Pérez-Rivas
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  3. Sandra Blanco
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  4. Adrián A. Castillo-Dominguez
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  5. José Lozano
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  6. Pedro A. Lazo
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Corresponding author

Correspondence to Pedro A. Lazo.

Additional information

The protein interactions from this publication have been submitted to the EMBL-EBI IMEx (http://www.imexconsortium.org/) consortium through IntAct (pmid 22121220) and assigned the identifier IM-17102 for the VRK2–MEK1 interactions and the identifier IM-17103 for the VRK2–KSR1 interactions.

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Fernández, I.F., Pérez-Rivas, L.G., Blanco, S. et al. VRK2 anchors KSR1-MEK1 to endoplasmic reticulum forming a macromolecular complex that compartmentalizes MAPK signaling. Cell. Mol. Life Sci. 69, 3881–3893 (2012). https://doi.org/10.1007/s00018-012-1056-8

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