ANKRD1

Structure

Human cardiac ankyrin repeat protein is a 36.2kDa protein composed of 319 amino acids., though in cardiomyocytes, CARP can exist as multiple alternatively spliced forms. CARP contains five tandem ankyrin repeats. Studies have shown that CARP can homodimerize. Studies have also shown that CARP is N-terminally, post-translationally cleaved by calpain-3 in skeletal muscle, suggesting alternate bioactive forms of CARP exist. CARP has been localized to nuclei and Z-discs in animal and human muscle cells, and at intercalated discs in human cardiac muscle cells.

Function

CARP was originally identified as a YB-1-associating, cardiac-restricted transcription co-repressor in the homeobox NKX2-5 pathway that is involved in cardiac ventricular chamber specification, maturation and morphogenesis, and whose mRNA levels are exquisitely sensitive to Doxorubicin, mediated through a hydrogen peroxide/ERK/p38MAP kinase-dependent as well as M-CAT cis-element-dependent mechanism. Subsequent studies showed that CARP expression in cardiomyocytes is regulated by alpha-adrenergic signaling, in part via the transcription factor GATA4. An additional study showed that beta-adrenergic signaling via protein kinase A and CaM kinase induces the expression of CARP, and that CARP may have a negative effect on contractile function. CARP has also been identified as a transcriptional co-activator of tumor suppressor protein p53 for stimulating gene expression in muscle; p53 was found to be an upstream effector of CARP via upregulation of the proximal ANKRD1 promoter. CARP has a relatively short half-life being longer in cardiomyocytes than endothelial cells; and CARP is degraded by the 26S proteasome via a PEST degron.

In animal models of disease and injury, CARP has been characterized to be a stress-inducible myofibrillar protein. CARP has been shown to play a role in skeletal muscle structure remodeling, and repair, being expressed in skeletal muscle near myotendinous junctions, and in vascular smooth muscle cells, as a downstream target of TGF-beta/Smad sigmaling in response to balloon injury and atherosclerotic plaques. Further studies have identified a role for CARP in initiation and regulation of arteriogenesis. Decreased expression of CARP in cardiac cells within the ischemic region was detected in a rat model of ischemic injury, and was thought to be linked to the induction of GADD153, an apoptosis-related gene. In cardiomyocytes treated with doxorubicin, a model of anthracycline-induced cardiomyopathy, CARP mRNA and protein levels were depleted, myofilament gene transcription was attenuated and sarcomeres showed significant disarray.

In a transgenic mouse model of cardiac-specific overexpression of CARP, mice exhibited normal physiology at baseline, but were protected against pathological cardiac hypertrophy induced via pressure-overload or isoproterenol, which could be attributed to the downregulation of the ERK1/2, MEK and TGFbeta-1 pathways. Another study demonstrated that adenoviral overexpression of CARP in cardiomyocytes enhances cardiac hypertrophy induced by Angiotensin II or pressure-overload and promotes cardiomyocyte apoptosis via p53 activation and mitochondrial dysfunction. However, transgenic knockout models of either CARP alone or CARP in combination with the other muscle ankyrin repeat proteins (MARPs), ANKRD2 and ANKRD23 demonstrated a lack of cardiac phenotype; mice displayed normal cardiac function at baseline and in response to pressure overload-induced cardiac hypertrophy, suggesting that these proteins are not essential.

Interactions between CARP and the sarcomeric proteins myopalladin and titin suggest that it may also be involved in the myofibrillar stretch-sensor system. Passive stretch in fetal cardiomyocytes induced differential CARP distribution at nuclei and I-band titin N2A regions. In a mouse model of muscular dystrophy with myositis (mdm) caused by a small deletion in titin, CARP mRNA expression was shown to be 30-fold elevated in skeletal muscle tissue.

Clinical significance

A wide spectrum of clinical features have been associated with ANKRD1/CARP. Mutations in ANKRD1 have been associated with dilated cardiomyopathy, two of which result in altered binding with TLN1 and FHL2. Mutations in ANKRD1 have also been associated with hypertrophic cardiomyopathy, and have shown to increase binding of CARP to Titin and MYPN. Examination of the functional effects of CARP hypertrophic cardiomyopathy mutations in engineered heart tissue demonstrated that Thr123Met was a gain-of-function mutation exhibiting augmented contractile properties; whereas Pro52Ala and Ile280Val were unstable and failed to incorporate into sarcomeres, an effect that was remedied upon proteasome inhibition via epoxomicin.

A missense mutation in ANKRD1 was shown to be associated with the congenital heart defect, Anomalous pulmonary venous connection. CARP has been found as a sensitive and specific biomarker for the differential diagnosis of rhabdomyosarcoma. ANKRD1 mRNA levels correlate with patient platinum sensitivity, thus ANKRD1 associates with platinum-based chemotherapy treatment outcome in ovarian adenocarcinoma patients.

CARP and mRNA expression has been shown to be upregulated in left ventricles of heart failure patients. Studies in patients with amyotrophic lateral sclerosis, spinal muscular atrophy, and congenital myopathy, also found altered expression of CARP in skeletal muscle fibers. Another study in congenital muscular dystrophy and Duchenne muscular dystrophy patients showed elevated expression of CARP. CARP expression is also elevated in patients with lupus nephritis, and associates with proteinuria severity, suggesting that it may have biomarker potential.

Interactions

ANKRD1 has been shown to interact with:

• Titin,
• MYPN,
• YBX1,
• CASQ2,
• DES,
• TP53,
• TLN1, and
• FHL2.

References

References

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12. (Sep 1997). "A novel cardiac-restricted target for doxorubicin. CARP, a nuclear modulator of gene expression in cardiac progenitor cells and cardiomyocytes". The Journal of Biological Chemistry.
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31. (2012). "Cardiac ankyrin repeat protein attenuates cardiac hypertrophy by inhibition of ERK1/2 and TGF-β signaling pathways". PLOS ONE.
32. (2014). "Cytosolic CARP promotes angiotensin II- or pressure overload-induced cardiomyocyte hypertrophy through calcineurin accumulation". PLOS ONE.
33. (May 2015). "Overexpression of ankyrin repeat domain 1 enhances cardiomyocyte apoptosis by promoting p53 activation and mitochondrial dysfunction in rodents". Clinical Science.
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38. (Sep 2009). "Mutations in the ANKRD1 gene encoding CARP are responsible for human dilated cardiomyopathy". European Heart Journal.
39. (Jul 2009). "Cardiac ankyrin repeat protein gene (ANKRD1) mutations in hypertrophic cardiomyopathy". Journal of the American College of Cardiology.
40. (May 2013). "Impact of ANKRD1 mutations associated with hypertrophic cardiomyopathy on contraction parameters of engineered heart tissue". Basic Research in Cardiology.
41. (Apr 2008). "Transcriptional deregulation and a missense mutation define ANKRD1 as a candidate gene for total anomalous pulmonary venous return". Human Mutation.
42. (Nov 2008). "Expression of cardiac ankyrin repeat protein, CARP, in malignant tumors: diagnostic use of CARP protein immunostaining in rhabdomyosarcoma". Human Pathology.
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46. (Jul 2004). "Altered titin expression, myocardial stiffness, and left ventricular function in patients with dilated cardiomyopathy". Circulation.
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48. (Apr 2003). "Altered expression of cardiac ankyrin repeat protein and its homologue, ankyrin repeat protein with PEST and proline-rich region, in atrophic muscles in amyotrophic lateral sclerosis". Pathobiology.
49. (Oct 2003). "Cardiac ankyrin repeat protein is preferentially induced in atrophic myofibers of congenital myopathy and spinal muscular atrophy". Pathology International.
50. (May 2003). "Cardiac-restricted ankyrin-repeated protein is differentially induced in duchenne and congenital muscular dystrophy". Laboratory Investigation.
51. (Mar 2007). "Upregulated expression of cardiac ankyrin-repeated protein in renal podocytes is associated with proteinuria severity in lupus nephritis". Human Pathology.
52. (November 2003). "The muscle ankyrin repeat proteins: CARP, ankrd2/Arpp and DARP as a family of titin filament-based stress response molecules". J. Mol. Biol..
53. (April 2001). "Myopalladin, a novel 145-kilodalton sarcomeric protein with multiple roles in Z-disc and I-band protein assemblies". J. Cell Biol..
54. (Feb 2005). "ANKRD1 specifically binds CASQ2 in heart extracts and both proteins are co-enriched in piglet cardiac Purkinje cells". Journal of Molecular and Cellular Cardiology.