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Cdk5 is involved in neuregulin-induced AChR expression at the neuromuscular junction

Nature Neuroscience volume 4, pages 374–381 (2001)Cite this article

Abstract

Here we describe an important involvement of Cdk5/p35 in regulating the gene expression of acetylcholine receptor (AChR) at the neuromuscular synapse. Cdk5 and p35 were prominently expressed in embryonic muscle, and concentrated at the neuromuscular junction in adulthood. Neuregulin increased the p35-associated Cdk5 kinase activity in the membrane fraction of cultured C2C12 myotubes. Co-immunoprecipitation studies revealed the association between Cdk5, p35 and ErbB receptors in muscle and cultured myotubes. Inhibition of Cdk5 activity not only blocked the NRG-induced AChR transcription, but also attenuated ErbB activation in cultured myotubes. In light of our finding that overexpression of p35 alone led to an increase in AChR promoter activity in muscle, Cdk5 activation is sufficient to mediate the up-regulation of AChR gene expression. Taken together, these results reveal the unexpected involvement of Cdk5/p35 in neuregulin signaling at the neuromuscular synapse.

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Figure 1: Cdk5 and p35 were prominently expressed in muscle during embryonic development.
Figure 2: Localization of Cdk5 and p35 in muscle during development and after nerve injury.
Figure 3: Regulation of expression of Cdk5 and p35 in NRG-treated myotubes.
Figure 4: NRG increased Cdk5 kinase activity associated with membrane fractions of C2C12 myotubes.
Figure 5: Association of Cdk5 and p35 with ErbB receptors and serine/threonine phosphorylation of ErbB3.
Figure 6: Inhibition of Cdk5 activity attenuated the NRG-induced AChR transcription and ErbB3 phosphorylation in cultured myotubes.
Figure 7: Overexpression of p35 increased the expression of AChRɛ transgene both in vitro and in vivo.

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References

  1. Lew, J., Winkfein, R. J., Paudel, H. K. & Wang, J. H. Brain proline-directed protein kinase is a neurofilament kinase which displays high sequence homology to p34cdc2. J. Biol. Chem. 267, 25922–25926 (1992).

    CAS  Google Scholar 

  2. Lew, J. et al. A brain-specific activator of cyclin-dependent kinase 5. Nature 371, 423–426 (1994).

    Article  CAS  Google Scholar 

  3. Tsai, L. H., Delalle, I., Caviness, V. S. J., Chae, T. & Harlow, E. p35 is a neural-specific regulatory subunit of cyclin-dependent kinase 5. Nature 371, 419–423 (1994).

    Article  CAS  Google Scholar 

  4. Tang, D. et al. An isoform of the neuronal cyclin-dependent kinase 5 (Cdk5) activator. J. Biol. Chem. 270, 26897–26903 (1995).

    Article  CAS  Google Scholar 

  5. Ohshima, T. et al. Targeted disruption of the cyclin-dependent kinase 5 gene results in abnormal corticogenesis, neuronal pathology and perinatal death. Proc. Natl. Acad. Sci. USA 93, 11173–11178 (1996).

    Article  CAS  Google Scholar 

  6. Chae, T. et al. Mice lacking p35, a neuronal specific activator of Cdk5, display cortical lamination defects, seizures and adult lethality. Neuron 18, 29–42 (1997).

    Article  CAS  Google Scholar 

  7. Kwon, Y. T. & Tsai, L. H. The role of the p35/cdk5 kinase in cortical development. Results Probl. Cell Differ. 30, 241–253 (2000).

    Article  CAS  Google Scholar 

  8. Paudel, H. K., Lew, J., Ali, Z. & Wang, J. H. Brain proline directed protein kinase phosphorylates tau on sites that are abnormally phosphorylated in tau associated with Alzheimer's paired helical filaments. J. Biol. Chem. 268, 23512–13518 (1993).

    CAS  Google Scholar 

  9. Tomizawa, K. et al. Localization and developmental changes in the neuron-specific cyclin-dependent kinase 5 activator (p35nck5a) in the rat brain. Neuroscience 74, 519–529 (1996).

    Article  CAS  Google Scholar 

  10. Lazaro, J.-B. et al. Cyclin dependent kinase 5, Cdk5, is a positive regulator of myogenesis in mouse C2 cells. J. Cell Sci. 110, 1251–1260 (1997).

    CAS  PubMed  Google Scholar 

  11. Philpott, A., Porro, E. B., Kirschner, M. W. & Tsai, L. H. The role of cyclin-dependent kinase 5 and a novel regulatory subunit in regulating muscle differentiation and patterning. Genes Dev . 11, 1409–1421 (1997).

    Article  CAS  Google Scholar 

  12. Fischbach, G. D. & Rosen, K. M. ARIA: a neuromuscular neuregulin. Annu. Rev. Neurosci. 20, 429–458 (1997).

    Article  CAS  Google Scholar 

  13. Carraway, K. L. III, Burden, S. J. Neuregulins and their receptors. Curr. Opin. Neurobiol. 5, 606–612 (1995).

    Article  CAS  Google Scholar 

  14. Corfas, G. & Fischbach, G. D. The number of Na+ channels in cultured chick muscle is increased by ARIA, an acetylcholine receptor-inducing activity. J. Neurosci. 13, 2118–2125 (1993).

    Article  CAS  Google Scholar 

  15. Chu, G. C., Moscoso, L. M., Sliwkowski, M. X. & Merlie, J. P. Regulation of the acetylcholine receptor epsilon subunit gene by recombinant ARIA: an in vitro model for transynaptic gene regulation. Neuron 14, 329–339 (1995).

    Article  CAS  Google Scholar 

  16. Gramolini, A. O. et al. Induction of utrophin gene expression by heregulin in skeletal muscle cells: Role of the N-box motif and GA binding protein. Proc. Natl. Acad. Sci. USA 96, 3223–3227 (1999).

    Article  CAS  Google Scholar 

  17. Fu, A. K. Y., Cheung, W. M. W., Ip, F. C. F. & Ip, N. Y. Identification of genes induced by neuregulin in cultured myotubes. Mol. Cell Neurosci. 14, 241–253 (1999).

    Article  CAS  Google Scholar 

  18. Loeb, J. A. & Fischbach, G. D. ARIA can be released from extracellular matrix though cleavage of a heparin-binding domain. J. Cell Biol. 130, 127–135 (1995).

    Article  CAS  Google Scholar 

  19. Moscoso, L. M. et al. Synapse-associated expression of an acetylcholine receptor-inducing protein, ARIA/heregulin, and its putative receptors, erbB2 and erbB3, in developing mammalian muscle. Dev. Biol. 172, 158–169 (1995).

    Article  CAS  Google Scholar 

  20. Si, J., Luo, Z. & Mei, L. Induction of acetylcholine receptor gene expression by ARIA requires activation of mitogen-activated protein kinase. J. Biol. Chem. 271, 19752–19759 (1996).

    Article  CAS  Google Scholar 

  21. Si, J., Wang, Q. & Mei, L. Essential roles of c-JUN and c-JUN N-terminal kinase (JNK) in neuregulin-increased expression of the acetylcholine receptor epsilon-subunit. J. Neurosci. 19, 8498–8508 (1999).

    Article  CAS  Google Scholar 

  22. Tansey, M. G., Chu, G. C. & Merlie, J. P. ARIA/HRG regulates AChRɛ subunit gene expression at the neuromuscular synapse via activation of phosphatidylinositol 3-kinase and Ras/MAPK pathway. J. Cell Biol. 134, 465–476 (1996).

    Article  CAS  Google Scholar 

  23. Meijer, L. et al. Biochemical and cellular effects of roscovitine, a potent and selective inhibitor of the cyclin-dependent kinases cdc2, cdk2 and cdk5. Eur. J. Biochem 243, 527–536 (1997).

    Article  CAS  Google Scholar 

  24. Si, J., Miller, D. S. & Mei, L. Identification of an element required for acetylcholine receptor-inducing activity (ARIA)-induced expression of the acetylcholine receptor ɛ subunit gene. J. Biol. Chem. 272, 10367–10371 (1997).

    Article  CAS  Google Scholar 

  25. Zukerberg, L. R. et al. Cables links Cdk5 and c-abl and facilitates Cdk5 tyrosine phosphorylation, kinase upregulation and neurite outgrowth. Neuron 26, 633–646 (2000).

    Article  CAS  Google Scholar 

  26. Minden, A., Lin, A., Claret, F. X., Abo, A. & Karin, M. Selective activation of the JNK signaling cascade and c-Jun transcriptional activity by the small GTPases Rac and Cdc42Hs. Cell 81, 1147–1157 (1995).

    Article  CAS  Google Scholar 

  27. Kiyokawa, N. et al. Involvement of cdc2-mediated phosphorylation in the cell cycle-dependent regulation of p185neu. Oncogene 15, 2633–2641 (1997).

    Article  CAS  Google Scholar 

  28. Feinmesser, R. L., Wicks, S. J., Taverner, C. J. & Chantry, A. Ca2+/Calmodulin-dependent kinase II phosphorylates the epidermal growth factor receptor on multiple sites in the cytoplasmic tail and serine 744 within the kinase domain to regulate signal generation. J. Biol. Chem. 274, 16168–16173 (1999).

    Article  CAS  Google Scholar 

  29. Lew, J. & Wang, J. H. Neuronal Cdc2-like kinase. TIBS 20, 33–37 (1995).

    CAS  PubMed  Google Scholar 

  30. Ozaki, M., Sasner, M., Yano, R., Lu, H. S. & Buonanno, A. Neuregulin-β induces expression of an NMDA-receptor subunit. Nature 390, 691–694 (1997).

    Article  CAS  Google Scholar 

  31. Rieff, H. I. et al. Neuregulin induces GABA(A) receptor subunit expression and neurite outgrowth in cerebellar granule cells. J. Neurosci. 19, 10757–10766 (1999).

    Article  CAS  Google Scholar 

  32. Yang, X., Kuo, Y., Devay, P., Yu, C. & Role, L. A cysteine-rich isoform of neuregulin controls the level of expression of neuronal nicotinic receptor channels during synaptogenesis. Neuron 20, 255–270 (1998).

    Article  CAS  Google Scholar 

  33. Sheng, M. & Wyszynski, M. Ion channel targeting in neurons. Bioessays 19, 847–853 (1997).

    Article  CAS  Google Scholar 

  34. Garcia, R. A. G., Vasudevan, K. & Buonanno, A. The neuregulin receptor ErbB-4 interacts with PDZ-containing proteins at neuronal synapses. Proc. Natl. Acad. Sci. USA 97, 3596–3601 (2000).

    Article  CAS  Google Scholar 

  35. Huang, Y. Z. et al. Regulation of neuregulin signaling by PSD-95 interacting with ErbB4 at CNS synapses. Neuron 26, 443–455 (2000).

    Article  CAS  Google Scholar 

  36. Fu, A. K. Y. et al. Xenopus muscle-specific kinase: molecular cloning and prominent expression in neural tissues during early embryonic development. Eur. J. Neurosci. 11, 373–382 (1999).

    Article  CAS  Google Scholar 

  37. Yang, J. F., Ng, Y. P., Pun, S., Ip, N. Y. & Tsim, K. W. K. The EGF-like domain of chick acetylcholine receptor-inducing activity (ARIA) contains its full biological activity. FEBS Lett. 403, 163–167 (1997).

    Article  CAS  Google Scholar 

  38. Ip, F. C. F., Fu, A. K. Y., Tsim, K. W. K. & Ip, N.Y. Cloning of the α component of the chick ciliary neurotrophic factor receptor: developmental expression and down-regulation in denervated skeletal muscle. J. Neurochem. 65, 2393–2400 (1995).

    Article  CAS  Google Scholar 

  39. Ching, Y. P., Qi, Z. & Wang, J. H. Cloning of three novel neuronal Cdk5 activator binding proteins. Gene 242, 285–294 (2000).

    Article  CAS  Google Scholar 

  40. Ip, F. C. F. et al. Cloning and characterization of muscle specific kinase (MuSK) in chicken. Mol. Cell. Neurosci. 16, 661–673 (2000).

    Article  CAS  Google Scholar 

  41. Pigino, G., Paglini, G., Ulloa, L., Avila, J. & Caceres, A. Analysis of the expression, distribution and function of cyclin dependent kinase 5 (cdk5) in developing cerebellar macroneurons. J. Cell Sci. 110, 257–270 (1997).

    CAS  PubMed  Google Scholar 

  42. Ojala, J., Choudhury, M. & Bag, J. Inhibition of troponin C production without affecting other muscle protein synthesis by the antisense oligodeoxynucleotide. Antisense Nucleic Acid Drug Dev. 7, 31–38 (1997).

    Article  CAS  Google Scholar 

  43. Nikolic, M., Dudek, H., Kwon, Y. T., Ramos, Y. F. & Tsai, L. H. The Cdk5/p35 kinase is essential for neurite outgrowth during neuronal differentiation. Genes Dev. 10, 816–925 (1996).

    Article  CAS  Google Scholar 

  44. Si, J. & Mei, L. ERK MAP kinase activation is required for acetylcholine receptor inducing activity-induced increase in all five acetylcholine receptor subunit mRNAs as well as synapse-specific expression of acetylcholine receptor ɛ-transgene. Mol. Brain Res. 67, 18–27 (1999).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank Z. Wu for the myogenin cDNA plasmid, L. Mei for the ErbB2 and ErbB3 expression constructs and Y.P. Ching for advice. We thank P. Xie, R. Choi and K.C. Lok for technical assistance. This study was supported in part by grants from the Research Grants Council of Hong Kong to N.Y.I. (HKUST 2/99C, 6107/98M and 568/95M) and the Hong Kong Jockey Club. N.Y.I. was a recipient of the Croucher Foundation Senior Research Fellowship.

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  1. Amy K. Y. Fu and Wing-Yu Fu: The first two authors contributed equally to this work

Authors and Affiliations

  1. Department of Biochemistry, Biotechnology Research Institute, Molecular Neuroscience Center, Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China

    Amy K. Y. Fu, Wing-Yu Fu, Janet Cheung, Fanny C. F. Ip, Jerry H. Wang & Nancy Y. Ip

  2. Department of Biology, Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China

    Karl W. K. Tsim

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Fu, A., Fu, WY., Cheung, J. et al. Cdk5 is involved in neuregulin-induced AChR expression at the neuromuscular junction. Nat Neurosci 4, 374–381 (2001). https://doi.org/10.1038/86019

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