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Citron-N is a neuronal Rho-associated protein involved in Golgi organization through actin cytoskeleton regulation

Nature Cell Biology volume 5, pages 1071–1078 (2003)Cite this article

Abstract

The actin cytoskeleton is best known for its role during cellular morphogenesis. However, other evidence suggests that actin is also crucial for the organization and dynamics of membrane organelles such as endosomes and the Golgi complex1,2,3. As in morphogenesis, the Rho family of small GTPases are key mediators of organelle actin-driven events4,5, although it is unclear how these ubiquitously distributed proteins are activated to regulate actin dynamics in an organelle-specific manner. Here we show that the brain-specific Rho-binding protein Citron-N6 is enriched at, and associates with, the Golgi apparatus of hippocampal neurons in culture. Suppression of the whole protein or expression of a mutant form lacking the Rho-binding activity results in dispersion of the Golgi apparatus. In contrast, high intracellular levels induce localized accumulation of RhoA and filamentous actin, protecting the Golgi from the rupture normally produced by actin depolymerization. Biochemical and functional analyses indicate that Citron-N controls actin locally by assembling together the Rho effector ROCK-II and the actin-binding, neuron-specific, protein Profilin-IIa (PIIa). Together with recent data on endosomal dynamics5, our results highlight the importance of organelle-specific Rho modulators for actin-dependent organelle organization and dynamics.

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Figure 1: CIT-N is a neuronal and glial protein associated with the Golgi.
Figure 2: CIT-N is required for maintaining Golgi architecture.
Figure 3: Overexpression of CIT-N prevents cytochalasin D-induced Golgi fragmentation.
Figure 4: Characterization of proteins interacting with the C-terminal domain of CIT-N.
Figure 5: Functional interaction between CIT-N, Rho kinase and PIIa.

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References

  1. Allan, V.J. & Schroer, T.A. Membrane motors. Curr. Opin. Cell Biol. 11, 476–482 (1999).

    Article  CAS  PubMed  Google Scholar 

  2. Allan, V.J., Thompson, H.M. & McNiven, M.A. Motoring around the Golgi. Nature Cell Biol. 4, E236–E242 (2002).

    Article  CAS  PubMed  Google Scholar 

  3. di Campli, A. et al. Morphological changes in the Golgi complex correlate with actin cytoskeleton rearrangements. Cell Motil. Cytoskeleton 43, 334–348 (1999).

    Article  CAS  PubMed  Google Scholar 

  4. Etienne-Manneville, S. & Hall, A. Rho GTPases in cell biology. Nature 420, 629–635 (2002).

    Article  CAS  PubMed  Google Scholar 

  5. Gasman, S., Kalaidzidis, Y. & Zerial, M. RhoD regulates endosome dynamics through Diaphanous-related Formin and Src tyrosine kinase. Nature Cell Biol. 5, 195–204 (2003).

    Article  CAS  PubMed  Google Scholar 

  6. Madaule, P. et al. A novel partner for the GTP-bound forms of rho and rac. FEBS Lett. 377, 243–248 (1995).

    Article  CAS  PubMed  Google Scholar 

  7. Donaldson, J.G. & Lippincott-Schwartz, J. Sorting and signaling at the Golgi complex. Cell 101, 693–696 (2000).

    Article  CAS  PubMed  Google Scholar 

  8. Mellman, I. & Warren, G. The road taken: past and future foundations of membrane traffic. Cell 100, 99–112 (2000).

    Article  CAS  PubMed  Google Scholar 

  9. Thyberg, J. & Moskalewski, S. Role of microtubules in the organization of the Golgi complex. Exp. Cell Res. 246, 263–279 (1999).

    Article  CAS  PubMed  Google Scholar 

  10. Holleran, E.A. & Holzbaur, E.L. Speculating about spectrin: new insights into the Golgi-associated cytoskeleton. Trends Cell Biol. 8, 26–29 (1998).

    Article  CAS  PubMed  Google Scholar 

  11. Stow, J.L. & Heimann, K. Vesicle budding on Golgi membranes: regulation by G proteins and myosin motors. Biochim. Biophys. Acta 1404, 161–171 (1998).

    Article  CAS  PubMed  Google Scholar 

  12. Valderrama, F. et al. Actin microfilaments are essential for the cytological positioning and morphology of the Golgi complex. Eur. J. Cell Biol. 76, 9–17 (1998).

    Article  CAS  PubMed  Google Scholar 

  13. Erickson, J.W., Zhang, C., Kahn, R.A., Evans, T. & Cerione, R.A. Mammalian Cdc42 is a brefeldin A-sensitive component of the Golgi apparatus. J. Biol. Chem. 0271, 26850–26854 (1996).

    Article  CAS  Google Scholar 

  14. Miura, K. et al. ARAP1: a point of convergence for Arf and Rho signaling. Mol. Cell 9, 109–119 (2002).

    Article  CAS  PubMed  Google Scholar 

  15. Furuyashiki, T. et al. Citron, a Rho-target, interacts with PSD-95/SAP-90 at glutamatergic synapses in the thalamus. J. Neurosci. 19, 109–118 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Zhang, W., Vazquez, L., Apperson, M. & Kennedy, M.B. Citron binds to PSD-95 at glutamatergic synapses on inhibitory neurons in the hippocampus. J. Neurosci. 19, 96–108 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Di Cunto, F. et al. Defective neurogenesis in citron kinase knockout mice by altered cytokinesis and massive apoptosis. Neuron 28, 115–127 (2000).

    Article  CAS  PubMed  Google Scholar 

  18. Krijnse-Locker, J., Parton, R.G., Fuller, S.D., Griffiths, G. & Dotti, C.G. The organization of the endoplasmic reticulum and the intermediate compartment in cultured rat hippocampal neurons. Mol. Biol. Cell 6, 1315–1332 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Robinson, M.S. & Kreis, T.E. Recruitment of coat proteins onto Golgi membranes in intact and permeabilized cells: effects of brefeldin A and G protein activators. Cell 69, 129–138 (1992).

    Article  CAS  PubMed  Google Scholar 

  20. Summerton, J. Morpholino antisense oligomers: the case for an RNase H-independent structural type. Biochim. Biophys. Acta 1489, 141–158 (1999).

    Article  CAS  PubMed  Google Scholar 

  21. Hall, A. Rho GTPases and the actin cytoskeleton. Science 279, 509–514 (1998).

    Article  CAS  PubMed  Google Scholar 

  22. Zmuda, J.F. & Rivas, R.J. Actin filament disruption blocks cerebellar granule neurons at the unipolar stage of differentiation in vitro. J. Neurobiol. 43, 313–328 (2000).

    Article  CAS  PubMed  Google Scholar 

  23. Nakagawa, O. et al. ROCK-I and ROCK-II, two isoforms of Rho-associated coiled-coil forming protein serine/threonine kinase in mice. FEBS Lett. 392, 189–193 (1996).

    Article  CAS  PubMed  Google Scholar 

  24. Gebbink, M.F. et al. Identification of a novel, putative Rho-specific GDP/GTP exchange factor and a RhoA-binding protein: control of neuronal morphology. J. Cell Biol. 137, 1603–1613 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Mizuno, K. et al. Identification of a human cDNA encoding a novel protein kinase with two repeats of the LIM/double zinc finger motif. Oncogene 9, 1605–1612 (1994).

    CAS  PubMed  Google Scholar 

  26. Witke, W. et al. In mouse brain profilin I and profilin II associate with regulators of the endocytic pathway and actin assembly. EMBO J. 17, 967–976 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Krebs, A., Rothkegel, M., Klar, M. & Jockusch, B.M. Characterization of functional domains of mDia1, a link between the small GTPase Rho and the actin cytoskeleton. J. Cell Sci. 114, 3663–3672 (2001).

    CAS  PubMed  Google Scholar 

  28. Maekawa, M. et al. Signaling from Rho to the actin cytoskeleton through protein kinases ROCK and LIM-kinase. Science 285, 895–898 (1999).

    Article  CAS  PubMed  Google Scholar 

  29. Uehata, M. et al. Calcium sensitization of smooth muscle mediated by a Rho-associated protein kinase in hypertension. Nature 389, 990–994 (1997).

    Article  CAS  PubMed  Google Scholar 

  30. Da Silva, J.S. et al. RhoA/ROCK regulation of neuritogenesis via Profilin IIa-mediated control of actin stability. J. Cell Biol. 162, 1267–1279 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Gavin, A.C. et al. Functional organization of the yeast proteome by systematic analysis of protein complexes. Nature 415, 141–147 (2002).

    Article  CAS  PubMed  Google Scholar 

  32. Pierce, J.P., Mayer, T. & McCarthy, J.B. Evidence for a satellite secretory pathway in neuronal dendritic spines. Curr. Biol. 11, 351–355 (2001).

    Article  CAS  PubMed  Google Scholar 

  33. Goslin, K. & Banker, G.A. Culturing nerve cells. 251–281 (MIT Press, Cambridge, MA, 1991).

  34. Griffiths, G.M. & Isaaz, S. Granzymes A and B are targeted to the lytic granules of lymphocytes by the mannose-6-phosphate receptor. J. Cell Biol. 120, 885–896 (1993).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We are grateful to B. Hellias and E. Cassin for the excellent preparation of the primary cultures, to L. Doglio for help with the microinjections and to C. Galvan for help with the cell fractionation experiments. We are also grateful to A. Habermann for excellent technical assistance with the immuno-electron microscopy and to W. Witke for the generous gift of anti-PIIa antibodies. J.S.S. is supported by an FCT/PRAXIS XXI scholarship (Portuguese Ministry of Science and Technology). The financial support of Telethon-Italy to C.G.D. (GP0245Y02) and F.D.C. (GGP02075) is gratefully acknowledged. In addition, C.G.D. and F.D.C. are supported by the EU DIADEM project (QLRT-2000-02392) and by the Jerome Lejeune foundation, respectively.

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Author notes

  1. Paola Camera and Jorge Santos Da Silva: These authors have contributed equally to this work.

Authors and Affiliations

  1. Dipartimento di Genetica, Biologia e Biochimica, Università Degli Studi di Torino Via Santena, 5 bis, Torino, Italy

    Paola Camera, Luciana Ferrara, Sara Imarisio, Lorenzo Silengo & Ferdinando Di Cunto

  2. Cavalieri Ottolenghi Scientific Institute Università Degli Studi di Torino A.O. San Luigi Gonzaga Regione Gonzole 10, Orbassano (Torino), 10024, Italy

    Jorge Santos Da Silva, Vanessa Schubert & Carlos G. Dotti

  3. Department of Cell Biology, EMBL, Postfach 102209, Heidelberg, D- 69012, Germany

    Gareth Griffiths

  4. Consiglio Nazionale delle Ricerche, Istituto Scienze Produzioni Alimentari, Bioindustry Park del Canavese, Colleretto Giacosa (Torino), Italy

    Maria Gabriella Giuffrida

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  1. Paola Camera
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Correspondence to Carlos G. Dotti or Ferdinando Di Cunto.

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Camera, P., Da Silva, J., Griffiths, G. et al. Citron-N is a neuronal Rho-associated protein involved in Golgi organization through actin cytoskeleton regulation. Nat Cell Biol 5, 1071–1078 (2003). https://doi.org/10.1038/ncb1064

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