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Inhibition of calpain increases LIS1 expression and partially rescues in vivo phenotypes in a mouse model of lissencephaly

Nature Medicine volume 15, pages 1202–1207 (2009)Cite this article

Abstract

Lissencephaly is a devastating neurological disorder caused by defective neuronal migration. LIS1 (official symbol PAFAH1B1, for platelet-activating factor acetylhydrolase, isoform 1b, subunit 1) was identified as the gene mutated in individuals with lissencephaly, and it was found to regulate cytoplasmic dynein function and localization. Here we show that inhibition or knockdown of calpains protects LIS1 from proteolysis, resulting in the augmentation of LIS1 amounts in Lis1+/− mouse embryonic fibroblast cells and rescue of the aberrant distribution of cytoplasmic dynein, mitochondria and β-COP–positive vesicles. We also show that calpain inhibitors improve neuronal migration of Lis1+/− cerebellar granular neurons. Intraperitoneal injection of the calpain inhibitor ALLN to pregnant Lis1+/− dams rescued apoptotic neuronal cell death and neuronal migration defects in Lis1+/− offspring. Furthermore, in utero knockdown of calpain by short hairpin RNA rescued defective cortical layering in Lis1+/− mice. Thus, calpain inhibition is a potential therapeutic intervention for lissencephaly.

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Figure 1: Calpain inhibitors restore LIS1 expression in Lis1+/− MEF cells or DRG neurons.
Figure 2: Rescue of neuronal migration by administration of calpain inhibitors.
Figure 3: Knockdown of calpain by siRNA.
Figure 4: Rescue of defective corticogenesis in Lis1+/− mice by intraperitoneal injection of ALLN.
Figure 5: Rescue of impaired behavior in Lis1+/− mice by intraperitoneal injection of ALLN.

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Acknowledgements

We thank S. Kawashima (Tokyo Metropolitan Institute of Medical Science) for providing us with a calpain-specific antibody (1D10A7)36. We thank Y. Funae, H. Iwao, T. Yamauch, M. Muramatsu and Y. Nagai for generous support and encouragement. We also thank Y. Kira, Y. Yabunaka and R. Zako for technical support, H. Nishimura and K. Fujimoto for mouse breeding, T. Bando for in utero injection and K. Nakanishi for behavior study. This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science, Sports and Culture of Japan from the Ministry of Education, Science, Sports and Culture of Japan to M.S. and S.H. This work was also supported by The Sagawa Foundation for Promotion of Cancer Research, The Cell Science Research Foundation, The Japan Spina Bifida & Hydrocephalus Research Foundation, Takeda Science Foundation, The Hoh-ansha Foundation and Knowledge Cluster Initiative (Stage-2) Research Foundation to Shinji Hirotsune and by US National Institutes of Health grants NS41030 and HD47380 to A.W.-B. This work was also supported by a Grant-in-Aid for Scientific Research on Priority Areas -Integrative Brain Research (Shien)- from MEXT and a Grant-in-Aid from Neuroinformatics Japan Center to T.M.

Author information

Author notes

  1. Masami Yamada and Yuko Yoshida: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Genetic Disease Research, Osaka City University Graduate School of Medicine, Asahi-machi, Osaka, Japan

    Masami Yamada, Yuko Yoshida, Daisuke Mori, Takako Takitoh & Shinji Hirotsune

  2. Institute for Integrated Cell-Material Sciences, Kyoto University Yoshida-honmachi, Sakyo-ku, Kyoto, Japan

    Mineko Kengaku & Hiroki Umeshima

  3. Division of Systems Medical Science, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi, Japan

    Keizo Takao & Tsuyoshi Miyakawa

  4. Genetic Engineering and Functional Genomics Group, Frontier Technology Center, Kyoto University Faculty of Medicine, Yoshida-Konoe-cho, Sakyo-ku, Kyoto, Japan

    Keizo Takao & Tsuyoshi Miyakawa

  5. Japan Science and Technology Agency, Institute for Bioinformatics Research and Development and Core Research of Evolutional Science & Technology, Kawaguchi, Saitama, Japan

    Keizo Takao & Tsuyoshi Miyakawa

  6. Division of Cell Biology and Neuroscience, Department of Morphological and Physiological Sciences, Faculty of Medical Sciences, University of Fukui, Fukui, Japan

    Makoto Sato

  7. Research and Education Program for Life Science, Faculty of Medical Sciences, University of Fukui, Fukui, Japan

    Makoto Sato

  8. Department of Enzymatic Regulation for Cell Functions (Calpain Project), The Tokyo Metropolitan Institute of Medical Science (Rinshoken), Tokyo Metropolitan Organization for Medical Research, Setagaya-ku, Tokyo, Japan

    Hiroyuki Sorimachi

  9. Department of Pediatrics and Institute for Human Genetics, University of California, San Francisco School of Medicine, San Francisco, California, USA

    Anthony Wynshaw-Boris

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  1. Masami Yamada
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Contributions

M.Y. conducted most of the experiments and wrote the manuscript. Y.Y. conducted most of the experiments. D.M. contributed in vivo analysis of mutant mice. T.T., M.K. and H.U. contributed to reaggregation assay of granular neurons. K.T. and T.M. contributed to behavior analysis. M.S. contributed to in utero injection. H.S. contributed to experiments regarding calpain inhibition. A.W.-B. wrote the manuscript and provided valuable suggestions. S.H. conducted the most of experiments and wrote the manuscript.

Corresponding author

Correspondence to Shinji Hirotsune.

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Yamada, M., Yoshida, Y., Mori, D. et al. Inhibition of calpain increases LIS1 expression and partially rescues in vivo phenotypes in a mouse model of lissencephaly. Nat Med 15, 1202–1207 (2009). https://doi.org/10.1038/nm.2023

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