HGNC Approved Gene Symbol: CAST
SNOMEDCT: 1237509001;
Cytogenetic location: 5q15 Genomic coordinates (GRCh38): 5:95,961,429-96,774,683 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 5q15 | Peeling skin with leukonychia, acral punctate keratoses, cheilitis, and knuckle pads | 616295 | Autosomal recessive | 3 |
Calpastatin is an endogenous inhibitor of calpains (see 114170) consisting of 4 inhibitory repeats, each of which neutralizes an activated calpain. Unlike proteinases, it is an intrinsically unstructured protein, adopting a defined structure only upon binding to active calpain (Moldoveanu et al., 2008; Hanna et al., 2008).
In erythrocytes of patients with essential hypertension (see 145500), the level of calpastatin activity is significantly lower than in the red cells of normotensive subjects. Pontremoli et al. (1988) demonstrated by Western blot analysis that the decreased inhibitor activity is the result of a decrease in the amount of the inhibitor protein. Calpastatin isolated and purified from erythrocytes of normotensive and hypertensive patients had identical specific activities. Pontremoli et al. (1988) also presented evidence indicating that the decreased level of calpastatin cannot be ascribed to accelerated decay during the red cell life span.
Mimori et al. (1995) demonstrated that anti-calpastatin autoantibodies are present in as many as 57% of rheumatoid arthritis patients and concluded that they may participate in pathogenic mechanisms of this and other rheumatic diseases which showed a lower frequency.
Lin et al. (2015) performed siRNA-mediated knockdown of CAST in the HaCaT immortalized keratinocyte cell line. Immunocytochemical analysis before and after mechanical stress revealed breakage of intercellular connections in CAST-knockdown cell monolayers, independent of whether they had been subjected to mechanical stress. In contrast, control cells showed stretched keratin filaments after mechanical stress but no disruption in intercellular adhesion prior to stress. Lin et al. (2015) concluded that calpastatin plays a role in keratinocyte adhesion.
Crystal Structure
Moldoveanu et al. (2008) reported the 3.0-angstrom crystal structure of calcium-bound m-calpain (CAPN2; 114230) in complex with the first calpastatin 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-2 (m-calpain) 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.
Using a cDNA probe encoding the 5-prime terminal region of CAST for spot-blot analysis of sorted chromosomes and chromosomal in situ hybridization, Inazawa et al. (1990) assigned the CAST gene to 5q14-q22. Inazawa et al. (1991) mapped CAST to 5q15-q21 by 2 methods of in situ hybridization and confirmed the results by spot-blot analysis of sorted chromosomes.
In 4 affected individuals from 3 unrelated families with peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads (PLACK; 616295), Lin et al. (2015) identified homozygosity for 3 different truncating mutations in the CAST gene (114090.0001-114090.0003) that segregated with disease in each family. Analysis of patient skin demonstrated impaired intercellular adhesion as well as keratinocyte apoptosis; in vitro knockdown studies confirmed a role for calpastatin in keratinocyte adhesion.
Reduced sarcolemmal integrity in dystrophin (300377)-deficient muscles of mdx mice and Duchenne muscular dystrophy (DMD; 310200) patients had been reported to result in altered calcium homeostasis. Previous studies had shown a correlative relationship between calpain activity in dystrophic muscle and muscle necrosis, but had not tested whether calpain activation precedes cell death or is a consequence of it. Spencer and Mellgren (2002) generated mdx mice that overexpressed a calpastatin transgene in muscle. Normal-appearing transgenic mice overexpressing calpastatin were crossed with mdx mice. Two lines of mice were examined, with different levels of calpastatin overexpression. Both lines of transgenic/mdx mice showed reductions in muscle necrosis at 4 weeks of age. The extent of improvement correlated with the level of calpastatin protein expression. Membrane damage, as assessed by procion orange and creatine kinase assays, was unchanged, supporting the idea that calpains act downstream of the primary muscle defect. The authors hypothesized that calpains may play an active role in necrotic processes in dystrophic muscle, and that inhibition of calpains might provide a therapeutic option for treatment of DMD.
In a 28-year-old Chinese woman (individual 1) with peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads (PLACK; 616295), Lin et al. (2015) identified homozygosity for a 1-bp duplication, c.607dup (c.607dup, NM_001042440.3), in the CAST gene, causing a frameshift predicted to result in a premature termination codon (Ile203AsnfsTer8). Her unaffected parents, sibs, and daughters were heterozygous for the duplication, which was not found in 200 ethnically matched controls. Immunohistochemical analysis of patient skin showed absent calpastatin staining in contrast to control skin, which showed calpastatin throughout the epidermis and localized in the cytoplasm. Transmission electron microscopy revealed widening of intercellular spaces with chromatin condensation and margination in the upper stratum spinosum in lesional skin, suggesting impaired intercellular adhesion as well as keratinocyte apoptosis. A significant increase in apoptotic keratinocytes was also observed in TUNEL assays of lesional skin compared to normal control skin.
In a Nepalese woman (individual 2) with peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads (PLACK; 616295), Lin et al. (2015) identified homozygosity for a c.424A-T transversion (c.424A-T, NM_001042440.3) in the CAST gene, resulting in a lys142-to-ter (K142X) substitution. Her unaffected parents were heterozygous for the mutation. Immunofluorescence analysis of patient skin showed reduced calpastatin labeling compared to control skin.
In 2 brothers, 54 and 58 years of age (individuals 3 and 4, respectively), with peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads (PLACK; 616295), originally reported by Haber and Rose (1986), Lin et al. (2015) identified homozygosity for a 1-bp deletion, c.1750delG (c.1750delG, NM_001042440.3) in the CAST gene, causing a frameshift predicted to result in a premature termination codon (Val584TrpfsTer37). The mutation segregated with disease in the family.
Haber, R. M., Rose, T. H. Autosomal recessive pachyonychia congenita. Arch. Derm. 122: 919-923, 1986. [PubMed: 3527073]
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]
Inazawa, J., Nakagawa, H., Misawa, S., Abe, T., Minoshima, S., Fukuyama, R., Maki, M., Murachi, T., Hatanaka, M., Shimizu, N. Assignment of the human calpastatin gene (CAST) to chromosome 5 at region q14-q22. Cytogenet. Cell Genet. 54: 156-158, 1990. [PubMed: 2265559] [Full Text: https://doi.org/10.1159/000132982]
Inazawa, J., Nakagawa, H., Misawa, S., Abe, T., Minoshima, S., Fukuyama, R., Maki, M., Murachi, T., Hatanaka, M., Shimizu, N. Assignment of the human calpastatin gene (CAST) to chromosome 5 at region q15-q21. (Abstract) Cytogenet. Cell Genet. 58: 1898 only, 1991.
Lin, Z., Zhao, J., Nitoiu, D., Scott, C. A., Plagnol, V., Smith, F. J. D., Wilson, N. J., Cole, C., Schwartz, M. E., McLean, W. H. I., Wang, H., Feng, C., and 10 others. Loss-of-function mutations in CAST cause peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads. Am. J. Hum. Genet. 96: 440-447, 2015. [PubMed: 25683118] [Full Text: https://doi.org/10.1016/j.ajhg.2014.12.026]
Mellgren, R. L. Enzyme knocked for a loop. Nature 456: 337-338, 2008. [PubMed: 19020611] [Full Text: https://doi.org/10.1038/456337a]
Mimori, T., Suganuma, K., Tanami, Y., Nojima, T., Matsumura, M., Fujii, T., Yoshizawa, T., Suzuki, K., Akizuki, M. Autoantibodies to calpastatin (an endogenous inhibitor for calcium-dependent neutral protease, calpain) in systemic rheumatic diseases. Proc. Nat. Acad. Sci. 92: 7267-7271, 1995. [PubMed: 7638179] [Full Text: https://doi.org/10.1073/pnas.92.16.7267]
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]
Pontremoli, S., Salamino, F., Sparatore, B., De Tullio, R., Pontremoli, R., Melloni, E. Characterization of the calpastatin defect in erythrocytes from patients with essential hypertension. Biochem. Biophys. Res. Commun. 157: 867-874, 1988. [PubMed: 2849943] [Full Text: https://doi.org/10.1016/s0006-291x(88)80955-0]
Spencer, M. J., Mellgren, R. L. Overexpression of a calpastatin transgene in mdx muscle reduces dystrophic pathology. Hum. Molec. Genet. 11: 2645-2655, 2002. [PubMed: 12354790] [Full Text: https://doi.org/10.1093/hmg/11.21.2645]