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Biochem J. Nov 1, 1996; 319(Pt 3): 683–690.
PMCID: PMC1217843

Non-erythroid alpha-spectrin breakdown by calpain and interleukin 1 beta-converting-enzyme-like protease(s) in apoptotic cells: contributory roles of both protease families in neuronal apoptosis.


The cytoskeletal protein non-erythroid alpha-spectrin is well documented as an endogenous calpain substrate, especially under pathophysiological conditions. In cell necrosis (e.g. maitotoxin-treated neuroblastoma SH-SY5Y cells), alpha-spectrin breakdown products (SBDPs) of 150 kDa and 145 kDa were produced by cellular calpains. In contrast, in neuronal cells undergoing apoptosis (cerebellar granule neurons subjected to low potassium and SH-SY5Y cells treated with staurosporine), an additional SBDP of 120 kDa was also observed. The formation of the 120 kDa SBDP was insensitive to calpain inhibitors but was completely blocked by an interleukin 1 beta-converting-enzyme (ICE)-like protease inhibitor, Z-Asp-CH2OC(O)-2,6-dichlorobenzene. Autolytic activation of both calpain and the ICE homologue CPP32 was also observed in apoptotic cells. alpha-Spectrin can also be cleaved in vitro by purified calpains to produce the SBDP doublet of 150/145 kDa and by ICE and ICE homologues [ICH-1, ICH-2 and CPP32(beta)] to produce a 150 kDa SBDP. In addition, CPP32 and ICE also produced a 120 kDa SBDP. Furthermore inhibition of either ICE-like protease(s) or calpain protects both granule neurons and SH-SY5Y cells against apoptosis. Our results suggest that both protease families participate in the expression of neuronal apoptosis.

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Selected References

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  • Cohen JJ. Apoptosis. Immunol Today. 1993 Mar;14(3):126–130. [PubMed]
  • Rubin LL, Gatchalian CL, Rimon G, Brooks SF. The molecular mechanisms of neuronal apoptosis. Curr Opin Neurobiol. 1994 Oct;4(5):696–702. [PubMed]
  • Martin SJ, Green DR, Cotter TG. Dicing with death: dissecting the components of the apoptosis machinery. Trends Biochem Sci. 1994 Jan;19(1):26–30. [PubMed]
  • Wang L, Miura M, Bergeron L, Zhu H, Yuan J. Ich-1, an Ice/ced-3-related gene, encodes both positive and negative regulators of programmed cell death. Cell. 1994 Sep 9;78(5):739–750. [PubMed]
  • Fernandes-Alnemri T, Litwack G, Alnemri ES. CPP32, a novel human apoptotic protein with homology to Caenorhabditis elegans cell death protein Ced-3 and mammalian interleukin-1 beta-converting enzyme. J Biol Chem. 1994 Dec 9;269(49):30761–30764. [PubMed]
  • Fernandes-Alnemri T, Takahashi A, Armstrong R, Krebs J, Fritz L, Tomaselli KJ, Wang L, Yu Z, Croce CM, Salveson G, et al. Mch3, a novel human apoptotic cysteine protease highly related to CPP32. Cancer Res. 1995 Dec 15;55(24):6045–6052. [PubMed]
  • Enari M, Hug H, Nagata S. Involvement of an ICE-like protease in Fas-mediated apoptosis. Nature. 1995 May 4;375(6526):78–81. [PubMed]
  • Kamens J, Paskind M, Hugunin M, Talanian RV, Allen H, Banach D, Bump N, Hackett M, Johnston CG, Li P, et al. Identification and characterization of ICH-2, a novel member of the interleukin-1 beta-converting enzyme family of cysteine proteases. J Biol Chem. 1995 Jun 23;270(25):15250–15256. [PubMed]
  • Alnemri ES, Fernandes-Alnemri T, Litwack G. Cloning and expression of four novel isoforms of human interleukin-1 beta converting enzyme with different apoptotic activities. J Biol Chem. 1995 Mar 3;270(9):4312–4317. [PubMed]
  • Munday NA, Vaillancourt JP, Ali A, Casano FJ, Miller DK, Molineaux SM, Yamin TT, Yu VL, Nicholson DW. Molecular cloning and pro-apoptotic activity of ICErelII and ICErelIII, members of the ICE/CED-3 family of cysteine proteases. J Biol Chem. 1995 Jun 30;270(26):15870–15876. [PubMed]
  • Kumar S, Kinoshita M, Noda M, Copeland NG, Jenkins NA. Induction of apoptosis by the mouse Nedd2 gene, which encodes a protein similar to the product of the Caenorhabditis elegans cell death gene ced-3 and the mammalian IL-1 beta-converting enzyme. Genes Dev. 1994 Jul 15;8(14):1613–1626. [PubMed]
  • Miura M, Zhu H, Rotello R, Hartwieg EA, Yuan J. Induction of apoptosis in fibroblasts by IL-1 beta-converting enzyme, a mammalian homolog of the C. elegans cell death gene ced-3. Cell. 1993 Nov 19;75(4):653–660. [PubMed]
  • Nicholson DW, Ali A, Thornberry NA, Vaillancourt JP, Ding CK, Gallant M, Gareau Y, Griffin PR, Labelle M, Lazebnik YA, et al. Identification and inhibition of the ICE/CED-3 protease necessary for mammalian apoptosis. Nature. 1995 Jul 6;376(6535):37–43. [PubMed]
  • Chow SC, Weis M, Kass GE, Holmström TH, Eriksson JE, Orrenius S. Involvement of multiple proteases during Fas-mediated apoptosis in T lymphocytes. FEBS Lett. 1995 May 8;364(2):134–138. [PubMed]
  • Schlegel J, Peters I, Orrenius S. Isolation and partial characterization of a protease involved in Fas-induced apoptosis. FEBS Lett. 1995 May 8;364(2):139–142. [PubMed]
  • Siman R, Noszek JC. Excitatory amino acids activate calpain I and induce structural protein breakdown in vivo. Neuron. 1988 Jun;1(4):279–287. [PubMed]
  • Azuma M, David LL, Shearer TR. Cysteine protease inhibitor E64 reduces the rate of formation of selenite cataract in the whole animal. Curr Eye Res. 1991 Jul;10(7):657–666. [PubMed]
  • Wang KK, Yuen PW. Calpain inhibition: an overview of its therapeutic potential. Trends Pharmacol Sci. 1994 Nov;15(11):412–419. [PubMed]
  • Sarin A, Adams DH, Henkart PA. Protease inhibitors selectively block T cell receptor-triggered programmed cell death in a murine T cell hybridoma and activated peripheral T cells. J Exp Med. 1993 Nov 1;178(5):1693–1700. [PMC free article] [PubMed]
  • Squìer MK, Miller AC, Malkinson AM, Cohen JJ. Calpain activation in apoptosis. J Cell Physiol. 1994 May;159(2):229–237. [PubMed]
  • Sarin A, Clerici M, Blatt SP, Hendrix CW, Shearer GM, Henkart PA. Inhibition of activation-induced programmed cell death and restoration of defective immune responses of HIV+ donors by cysteine protease inhibitors. J Immunol. 1994 Jul 15;153(2):862–872. [PubMed]
  • Lazebnik YA, Kaufmann SH, Desnoyers S, Poirier GG, Earnshaw WC. Cleavage of poly(ADP-ribose) polymerase by a proteinase with properties like ICE. Nature. 1994 Sep 22;371(6495):346–347. [PubMed]
  • Casciola-Rosen LA, Miller DK, Anhalt GJ, Rosen A. Specific cleavage of the 70-kDa protein component of the U1 small nuclear ribonucleoprotein is a characteristic biochemical feature of apoptotic cell death. J Biol Chem. 1994 Dec 9;269(49):30757–30760. [PubMed]
  • Tewari M, Beidler DR, Dixit VM. CrmA-inhibitable cleavage of the 70-kDa protein component of the U1 small nuclear ribonucleoprotein during Fas- and tumor necrosis factor-induced apoptosis. J Biol Chem. 1995 Aug 11;270(32):18738–18741. [PubMed]
  • Tewari M, Quan LT, O'Rourke K, Desnoyers S, Zeng Z, Beidler DR, Poirier GG, Salvesen GS, Dixit VM. Yama/CPP32 beta, a mammalian homolog of CED-3, is a CrmA-inhibitable protease that cleaves the death substrate poly(ADP-ribose) polymerase. Cell. 1995 Jun 2;81(5):801–809. [PubMed]
  • Gu Y, Sarnecki C, Aldape RA, Livingston DJ, Su MS. Cleavage of poly(ADP-ribose) polymerase by interleukin-1 beta converting enzyme and its homologs TX and Nedd-2. J Biol Chem. 1995 Aug 11;270(32):18715–18718. [PubMed]
  • Martin SJ, O'Brien GA, Nishioka WK, McGahon AJ, Mahboubi A, Saido TC, Green DR. Proteolysis of fodrin (non-erythroid spectrin) during apoptosis. J Biol Chem. 1995 Mar 24;270(12):6425–6428. [PubMed]
  • Linnik MD, Zobrist RH, Hatfield MD. Evidence supporting a role for programmed cell death in focal cerebral ischemia in rats. Stroke. 1993 Dec;24(12):2002–2009. [PubMed]
  • MacManus JP, Buchan AM, Hill IE, Rasquinha I, Preston E. Global ischemia can cause DNA fragmentation indicative of apoptosis in rat brain. Neurosci Lett. 1993 Dec 24;164(1-2):89–92. [PubMed]
  • Dragunow M, Faull RL, Lawlor P, Beilharz EJ, Singleton K, Walker EB, Mee E. In situ evidence for DNA fragmentation in Huntington's disease striatum and Alzheimer's disease temporal lobes. Neuroreport. 1995 May 9;6(7):1053–1057. [PubMed]
  • Franklin JL, Johnson EM., Jr Suppression of programmed neuronal death by sustained elevation of cytoplasmic calcium. Trends Neurosci. 1992 Dec;15(12):501–508. [PubMed]
  • Wang KK, Nath R, Posner A, Raser KJ, Buroker-Kilgore M, Hajimohammadreza I, Probert A W, Jr, Marcoux FW, Ye Q, Takano E, et al. An alpha-mercaptoacrylic acid derivative is a selective nonpeptide cell-permeable calpain inhibitor and is neuroprotective. Proc Natl Acad Sci U S A. 1996 Jun 25;93(13):6687–6692. [PMC free article] [PubMed]
  • Dolle RE, Hoyer D, Prasad CV, Schmidt SJ, Helaszek CT, Miller RE, Ator MA. P1 aspartate-based peptide alpha-((2,6-dichlorobenzoyl)oxy)methyl ketones as potent time-dependent inhibitors of interleukin-1 beta-converting enzyme. J Med Chem. 1994 Mar 4;37(5):563–564. [PubMed]
  • Bertrand R, Solary E, O'Connor P, Kohn KW, Pommier Y. Induction of a common pathway of apoptosis by staurosporine. Exp Cell Res. 1994 Apr;211(2):314–321. [PubMed]
  • Mesner PW, Winters TR, Green SH. Nerve growth factor withdrawal-induced cell death in neuronal PC12 cells resembles that in sympathetic neurons. J Cell Biol. 1992 Dec;119(6):1669–1680. [PMC free article] [PubMed]
  • Bump NJ, Hackett M, Hugunin M, Seshagiri S, Brady K, Chen P, Ferenz C, Franklin S, Ghayur T, Li P, et al. Inhibition of ICE family proteases by baculovirus antiapoptotic protein p35. Science. 1995 Sep 29;269(5232):1885–1888. [PubMed]
  • Wang KK, Roufogalis BD, Villalobo A. Further characterization of calpain-mediated proteolysis of the human erythrocyte plasma membrane Ca2+-ATPase. Arch Biochem Biophys. 1988 Nov 15;267(1):317–327. [PubMed]
  • Koh JY, Choi DW. Quantitative determination of glutamate mediated cortical neuronal injury in cell culture by lactate dehydrogenase efflux assay. J Neurosci Methods. 1987 May;20(1):83–90. [PubMed]
  • Wang KK, Posner A, Hajimohammadreza I. Total protein extraction from cultured cells for use in electrophoresis and western blotting. Biotechniques. 1996 Apr;20(4):662–668. [PubMed]
  • Vasilakos JP, Ghayur T, Carroll RT, Giegel DA, Saunders JM, Quintal L, Keane KM, Shivers BD. IL-1 beta converting enzyme (ICE) is not required for apoptosis induced by lymphokine deprivation in an IL-2-dependent T cell line. J Immunol. 1995 Oct 1;155(7):3433–3442. [PubMed]
  • Saido TC, Yokota M, Nagao S, Yamaura I, Tani E, Tsuchiya T, Suzuki K, Kawashima S. Spatial resolution of fodrin proteolysis in postischemic brain. J Biol Chem. 1993 Nov 25;268(33):25239–25243. [PubMed]
  • Saido TC, Shibata M, Takenawa T, Murofushi H, Suzuki K. Positive regulation of mu-calpain action by polyphosphoinositides. J Biol Chem. 1992 Dec 5;267(34):24585–24590. [PubMed]
  • D'Mello SR, Galli C, Ciotti T, Calissano P. Induction of apoptosis in cerebellar granule neurons by low potassium: inhibition of death by insulin-like growth factor I and cAMP. Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):10989–10993. [PMC free article] [PubMed]
  • Harris AS, Croall DE, Morrow JS. The calmodulin-binding site in alpha-fodrin is near the calcium-dependent protease-I cleavage site. J Biol Chem. 1988 Oct 25;263(30):15754–15761. [PubMed]
  • Saito K, Elce JS, Hamos JE, Nixon RA. Widespread activation of calcium-activated neutral proteinase (calpain) in the brain in Alzheimer disease: a potential molecular basis for neuronal degeneration. Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):2628–2632. [PMC free article] [PubMed]
  • Crawford C, Willis AC, Gagnon J. The effects of autolysis on the structure of chicken calpain II. Biochem J. 1987 Dec 1;248(2):579–588. [PMC free article] [PubMed]
  • Mellgren RL, Lane RD. Myocardial calpain 2 is inhibited by monoclonal antibodies specific for the small, noncatalytic subunit. Biochim Biophys Acta. 1988 May 18;954(2):154–160. [PubMed]
  • Mashima T, Naito M, Kataoka S, Kawai H, Tsuruo T. Aspartate-based inhibitor of interleukin-1 beta-converting enzyme prevents antitumor agent-induced apoptosis in human myeloid leukemia U937 cells. Biochem Biophys Res Commun. 1995 Apr 26;209(3):907–915. [PubMed]
  • Wang KK, Villalobo A, Roufogalis BD. Calmodulin-binding proteins as calpain substrates. Biochem J. 1989 Sep 15;262(3):693–706. [PMC free article] [PubMed]
  • Morrow JS. The spectrin membrane skeleton: emerging concepts. Curr Opin Cell Biol. 1989 Feb;1(1):23–29. [PubMed]
  • Dhermy D. The spectrin super-family. Biol Cell. 1991;71(3):249–254. [PubMed]
  • Feinstein E, Kimchi A, Wallach D, Boldin M, Varfolomeev E. The death domain: a module shared by proteins with diverse cellular functions. Trends Biochem Sci. 1995 Sep;20(9):342–344. [PubMed]

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