Alternative titles; symbols
HGNC Approved Gene Symbol: CAPN2
Cytogenetic location: 1q41 Genomic coordinates (GRCh38): 1:223,701,597-223,776,018 (from NCBI)
The calpains, or calcium-activated neutral proteases (EC 3.4.22.17), are nonlysosomal intracellular cysteine proteases. The mammalian calpains include 2 ubiquitous isoforms, calpain I (mu-calpain) and calpain II (m-calpain), 2 stomach-specific proteins, and CAPN3 (114240), which is muscle-specific. Calpain I and calpain II are heterodimers with distinct large subunits, encoded by the CAPN1 (114220) and CAPN2 genes, respectively, associated with a common small subunit (CAPNS1; 114170).
By screening a human skeletal muscle cDNA library using the large subunits of rabbit and chicken CANP as probes, Imajoh et al. (1988) cloned CAPN2. The deduced protein contains 700 amino acids. N-terminal sequencing of CAPN2 purified from human liver indicated that the N-terminal methionine is removed, resulting in a mature 699-amino acid subunit with a calculated molecular mass of 79.9 kD. Comparison of the CAPN2 sequence with CAPN1 and the chicken CANP large subunit showed conservation of the 4-domain structure and of the active-site cysteine (cys140) in the protease domain (domain II) and the 4 putative Ca(2+)-binding sites in domain IV. Domain III, presumably a regulatory domain, is also highly conserved between the 3 proteins, but domain I is not.
By quantitative RT-PCR, Ueyama et al. (1998) found that expression of calpain-1 and calpain-2 mRNA was significantly increased in muscle biopsy samples derived from 5 men with progressive muscular dystrophy (e.g., DMD; 310200) and 2 men and 3 women with amyotrophic lateral sclerosis (ALS; 105400) compared with controls.
Morford et al. (2002) found that calpain-2, but not calpain-1, associated with membrane lipid rafts on human peripheral blood T cells and Jurkat T cells. Membrane raft-associated calpain activity in human T cells was enhanced with exogenous calcium. Calpain cleaved the cytoskeletal-associated protein talin (TLN1; 186745) during the first 30 minutes after cell stimulation. Morford et al. (2002) hypothesized that lipid raft-associated calpain-2 could function early in T-cell receptor signaling to facilitate immune synapse formation through cytoskeletal remodeling mechanisms.
Using confocal microscopy and isopycnic density centrifugation, Hood et al. (2003) found that calpain-1, calpain-2, the small regulatory calpain subunit, and calpastatin (CAST; 114090) associated with the endoplasmic reticulum and Golgi apparatus of human fibroblasts and glioblastoma cells. The association between these proteins and the endoplasmic reticulum and Golgi apparatus increased in the glioblastoma cell line following laminin (see LAMA1; 150320) stimulation. Calpain-2 also colocalized with inositol 4,5-bisphosphate and with membrane lipid rafts.
Adamec et al. (2002) investigated calpain-2 activation in a broad range of neurodegenerative diseases using immunofluorescence imaging. Activated calpain-2 was detected in all neurodegenerative diseases examined, including Alzheimer disease (AD; 104300), Down syndrome (190685), and Pick disease (172700), with the possible exception of frontotemporal dementia with inclusions (see 600274). Activated calpain-2 was detected in different cell types and colocalized with different pathologic structures. In neurons and glial cells, calpain-2 primarily colocalized with hyperphosphorylated tau protein (MAPT; 157140). In brains with AD neurofibrillary changes, colocalization of calpain-2 with phosphorylated tau was most extensive at transentorhinal stages of neurofibrillary degeneration rather than at later limbic stages. Aggregates of different proteins, such as huntingtin (HTT; 613004) and alpha-synuclein (SNCA; 163890), were negative for activated calpain-2, indicating that the presence of pathologic inclusions was not by itself the stimulus responsible for calpain-2 activation.
Using cell biologic, pharmacologic, and genetic methods, Chandramohanadas et al. (2009) found that the apicomplexan parasites Plasmodium falciparum and Toxoplasma gondii, the causative agents of malaria and toxoplasmosis, respectively, used host cell calpains to facilitate parasite egress. Immunodepletion and inhibition experiments showed that calpain-1 was required for escape of P. falciparum from human erythrocytes. Similarly, elimination of both calpain-1 and calpain-2 via small interfering RNA against the common regulatory subunit CAPNS1 in human osteosarcoma cells or deletion of Capns1 in mouse embryonic fibroblasts blocked egress of T. gondii. Chandramohanadas et al. (2009) concluded that P. falciparum and T. gondii both exploit host cell calpains to facilitate escape from intracellular parasitophorous vacuoles and/or the host plasma membrane, a process required for parasite proliferation.
Saraiva et al. (2013) demonstrated that human GAAP (TMBIM4; 616874) promoted cell adhesion and migration in a CAPN2-dependent manner. GAAP caused activation of store-operated Ca(2+) entry and thereby Ca(2+)-mediated stimulation of CAPN2 activity near the plasma membrane, leading to an increase in turnover of focal adhesions. Saraiva et al. (2013) concluded that GAAP is involved in coordinating cell migration via localized Ca(2+)-dependent activation of CAPN2.
Crystal Structure
Moldoveanu et al. (2008) reported the 3.0-angstrom crystal structure of calcium-bound m-calpain (calpain II) in complex with the first calpastatin (114090) repeat, both from rat, revealing the mechanism of exclusive specificity. The structure highlighted the complexity of calpain activation by calcium, illustrating key residues in a peripheral domain that serve to stabilize the protease core on calcium binding. Fully activated calpain binds 10 Ca(2+) atoms, resulting in several conformational changes allowing recognition by calpastatin. Calpain inhibition is mediated by the intimate contact with 3 critical regions of calpastatin. Two regions target the penta-EF-hand domains of calpain, and the third occupies the substrate-binding cleft, projecting a loop around the active site thiol to evade proteolysis.
Hanna et al. (2008) reported the 2.4-angstrom resolution crystal structure of the calcium-bound calpain II heterodimer bound by 1 of the 4 inhibitory domains of calpastatin. They observed that calpastatin inhibits calpain by occupying both sides of the active site cleft. Although the inhibitor passes through the active site cleft, it escapes cleavage in a novel manner by looping out and around the active site cysteine. The inhibitory domain of calpastatin recognizes multiple lower affinity sites present only in the calcium-bound form of the enzyme, resulting in an interaction that is tight, specific, and calcium-dependent. Hanna et al. (2008) concluded that this crystal structure, and that of the related complex described by Moldoveanu et al. (2008), also revealed the conformational changes that calpain undergoes on binding calcium, which include opening of the active site cleft and movement of the domains relative to each other to produce a more compact enzyme.
By a combination of spot blot hybridization with sorted chromosomes and Southern hybridization with human-mouse cell hybrid DNAs, Ohno et al. (1989, 1990) mapped the CANPL2 gene to chromosome 1.
Adamec, E., Mohan, P., Vonsattel, J. P., Nixon, R. A. Calpain activation in neurodegenerative diseases: confocal immunofluorescence study with antibodies specifically recognizing the active form of calpain 2. Acta Neuropath. 104: 92-104, 2002. [PubMed: 12070670] [Full Text: https://doi.org/10.1007/s00401-002-0528-6]
Chandramohanadas, R., Davis, P. H., Beiting, D. P., Harbut, M. B., Darling, C., Velmourougane, G., Lee, M. Y., Greer, P. A., Roos, D. S., Greenbaum, D. C. Apicomplexan parasites co-opt host calpains to facilitate their escape from infected cells. Science 324: 794-797, 2009. [PubMed: 19342550] [Full Text: https://doi.org/10.1126/science.1171085]
Hanna, R. A., Campbell, R. L., Davies, P. L. Calcium-bound structure of calpain and its mechanism of inhibition by calpastatin. Nature 456: 409-412, 2008. [PubMed: 19020623] [Full Text: https://doi.org/10.1038/nature07451]
Hood, J. L., Logan, B. B., Sinai, A. P., Brooks, W. H., Roszman, T. L. Association of the calpain/calpastatin network with subcellular organelles. Biochem. Biophys. Res. Commun. 310: 1200-1212, 2003. [PubMed: 14559243] [Full Text: https://doi.org/10.1016/j.bbrc.2003.09.142]
Imajoh, S., Aoki, K., Ohno, S., Emori, Y., Kawasaki, H., Sugihara, H., Suzuki, K. Molecular cloning of the cDNA for the large subunit of the high Ca(2+)-requiring form of human Ca(2+)-activated neutral protease. Biochemistry 27: 8122-8128, 1988. [PubMed: 2852952] [Full Text: https://doi.org/10.1021/bi00421a022]
Moldoveanu, T., Gehring, K., Green, D. R. Concerted multi-pronged attack by calpastatin to occlude the catalytic cleft of heterodimeric calpains. Nature 456: 404-408, 2008. [PubMed: 19020622] [Full Text: https://doi.org/10.1038/nature07353]
Morford, L. A., Forrest, K., Logan, B., Overstreet, L. K., Goebel, J., Brooks, W. H., Roszman, T. L. Calpain II colocalizes with detergent-insoluble rafts on human and Jurkat T-cells. Biochem. Biophys. Res. Commun. 295: 540-546, 2002. [PubMed: 12150984] [Full Text: https://doi.org/10.1016/s0006-291x(02)00676-9]
Ohno, S., Minoshima, S., Kudoh, J., Fukuyama, R., Ohmi-Imajoh, S., Suzuki, K., Shimizu, Y., Shimizu, N. Four genes for the calpain family locate on four distinct human chromosomes. Cytogenet. Cell Genet. 51: 1054-1055, 1989.
Ohno, S., Minoshima, S., Kudoh, J., Fukuyama, R., Shimizu, Y., Ohmi-Imajoh, S., Shimizu, N., Suzuki, K. Four genes for the calpain family locate on four distinct human chromosomes. Cytogenet. Cell Genet. 53: 225-229, 1990. [PubMed: 2209092] [Full Text: https://doi.org/10.1159/000132937]
Saraiva, N., Prole, D. L., Carrara, G., Johnson, B. F., Taylor, C. W., Parsons, M., Smith, G. L. hGAAP promotes cell adhesion and migration via the stimulation of store-operated Ca(2+) entry and calpain 2. J. Cell Biol. 202: 699-713, 2013. [PubMed: 23940116] [Full Text: https://doi.org/10.1083/jcb.201301016]
Ueyama, H., Kumamoto, T., Fujimoto, S., Murakami, T., Tsuda, T. Expression of three calpain isoform genes in human skeletal muscles. J. Neurol. Sci. 155: 163-169, 1998. [PubMed: 9562261] [Full Text: https://doi.org/10.1016/s0022-510x(97)00309-2]