Degradation of regulator of calcineurin 1 (RCAN1) is mediated by both chaperone‐mediated autophagy and ubiquitin proteasome pathways

Liu, Heng; Wang, Pin; Song, Weihong; Sun, Xiulian

doi:10.1096/fj.09-134296

Cited by 115 publications

(112 citation statements)

References 74 publications

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“…Indeed, several proteins were shown to be degraded both by the proteasome and the lysosome. These include HTT/huntingtin, 46 RCAN1/regulator of calcineurin 1, 47 EGFR/epidermal growth factor receptor and ERBB2, 48 SNCA 49 and NFKBIE/IKBE/nuclear factor of κ light polypeptide gene enhancer in B-cells inhibitor, ε/IκB. 50 By analogy, it is conceivable that HIF1A may also be a "dual-pathway" substrate, being degraded both by the UPS and CMA.…”

Section: Discussionmentioning

confidence: 99%

STUB1/CHIP is required for HIF1A degradation by chaperone-mediated autophagy

Fofo²,

et al. 2013

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The transcription factor hiF1 is mostly regulated by the oxygen-dependent proteasomal degradation of the labile subunit hiF1A. recent data showed degradation of hiF1A in the lysosome through chaperone-mediated autophagy (cMA). however the molecular mechanism involved has not been elucidated. This study shows that the KFerQ-like motif, that has been identified in all cMA substrates, is required to mediate the interaction between hiF1A and the chaperone hSpA8. Moreover, mutations in the KFerQ-like motif of hiF1A preclude the interaction with the cMA receptor LAMp2A, thus inhibiting its lysosomal degradation. importantly, we show for the first time that the ubiquitin ligase STUB1 is required for degradation of hiF1A in the lysosome by cMA. indeed, mutations in STUB1 that inhibit either the ubiquitin ligase activity or its ability to bind to hSpA8, both prevent degradation of hiF1A by cMA. Moreover, we show that hiF1A binds to and is translocated into intact lysosomes isolated from rat livers. This new pathway for degradation of hiF1A does not depend on the presence of oxygen and is activated in response to nutrient deprivation such that the levels of hiF1A bound to cMA positive lysosomes significantly increase in starved animal livers and the binding of hiF1A to LAMp2A increases in response to serum deprivation. Moreover, excessive degradation of hiF1A by cMA compromises cells' ability to respond to and survive under hypoxia, suggesting that this pathway might be of pathophysiological importance in conditions that combine hypoxia with starvation. leucinylleucinylleucinal/proteasome inhibitor; MTT, 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide; RCC, renal cell carcinoma; Serpinb, serpin peptidase inhibitor, clade B/Ovalbumin; SLC2A1, solute carrier family 2 (facilitated glucose transporter)/glucose transporter; STUB1, STIP1 homology and U-box containing protein 1; TPR, tetratricopeptide repeat; UPS, ubiquitin-proteasome system; VEGFA, vascular endothelial growth factor A; u-box, modified RING finger domain lacking the metal-chelating residues

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Section: Discussionmentioning

confidence: 99%

STUB1/CHIP is required for HIF1A degradation by chaperone-mediated autophagy

Fofo²,

et al. 2013

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“…Besides tau, a connection between CMA and a second Alzheimer's disease-related protein, the regulator of calcineurin 1 (RCAN1) has also been established. CMA degrades RCAN1, a protein whose high expression in Alzheimer's disease brains has been linked to neuronal demise [63]. The possible contribution of the blockage of CMA by pathogenic tau to the elevation of RCAN1 levels in the affected neurons awaits evaluation.…”

Section: Alzheimer's Diseasementioning

confidence: 99%

Chaperone-mediated autophagy: roles in disease and aging

2013

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This review focuses on chaperone-mediated autophagy (CMA), one of the proteolytic systems that contributes to degradation of intracellular proteins in lysosomes. CMA substrate proteins are selectively targeted to lysosomes and translocated into the lysosomal lumen through the coordinated action of chaperones located at both sides of the membrane and a dedicated protein translocation complex. The selectivity of CMA permits timed degradation of specific proteins with regulatory purposes supporting a modulatory role for CMA in enzymatic metabolic processes and subsets of the cellular transcriptional program. In addition, CMA contributes to cellular quality control through the removal of damaged or malfunctioning proteins. Here, we describe recent advances in the understanding of the molecular dynamics, regulation and physiology of CMA, and discuss the evidence in support of the contribution of CMA dysfunction to severe human disorders such as neurodegeneration and cancer.

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“…It has been demonstrated that in HEK293 cells, during calcineurin activation, RCAN1 is rapidly degraded in the lysosome, (26) and an alkalinization of lysosomal pH by chloroquine increases RCAN1 protein levels and decreases NFAT-calcineurin activity. (26) To confirm whether a similar mechanism occurs in osteoclasts, we treated þ/þ osteoclasts with lysosomal inhibitors (5 mM ammonium chloride or 10 mM chloroquine), or proteasomal inhibitor (1 mM MG-132; used as a positive control to show inhibition of protein degradation), incubated for 2 hours at 378C 10% CO 2 (extracellular acidosis) to activate NFATc1 signaling, (15) and assessed RCAN1 degradation by immunoblotting. Results demonstrated that þ/þ osteoclasts exposed to chloroquine and MG-132 had significantly higher RCAN1 protein levels compared with control (Fig.…”

Section: Rcan1 Is the Factor Responsible For Nfatc1 Inhibitionmentioning

confidence: 99%

“…(35,36) Moreover, during osteoclastogenesis, Rcan1 expression levels dramatically increase, (24) and Rcan1 is believed to be one of the genes associated with the craniofacial and skeletal defects present in Down syndrome. (22,25) Coincidentally, it has been demonstrated that in HEK293 cells, during calcineurin activation, RCAN1 is rapidly degraded in the lysosome, (26) and an alkalinization of lysosomal pH by chloroquine increases RCAN1 protein levels and decreases NFAT-calcineurin activity. (26) We found that bone marrow-derived osteoclasts express RCAN1, and gene and protein levels were, somewhat unexpectedly given the reduction in NFATc1 expression, similar in þ/þ and þ/R740S cells; however, relative to NFATc1 protein (Fig.…”

Section: Journal Of Bone and Mineral Researchmentioning

confidence: 99%

The R740S mutation in the V-ATPase a3 subunit increases lysosomal pH, impairs NFATc1 translocation, and decreases in vitro osteoclastogenesis

Voronov

Ochotny

Jaumouillé

et al. 2012

Journal of Bone and Mineral Research

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Vacuolar Hþ -ATPase (V-ATPase), a multisubunit enzyme located at the ruffled border and in lysosomes of osteoclasts, is necessary for bone resorption. We previously showed that heterozygous mice with an R740S mutation in the a3 subunit of V-ATPase (þ/R740S) have mild osteopetrosis resulting from an $90% reduction in proton translocation across osteoclast membranes. Here we show that lysosomal pH is also higher in þ/R740S compared with wild-type (þ/þ) osteoclasts. Both osteoclast number and size were decreased in cultures of þ/R740S compared with þ/þ bone marrow cells, with concomitant decreased expression of key osteoclast markers (TRAP, cathepsin K, OSCAR, DC-STAMP, and NFATc1), suggesting that low lysosomal pH plays an important role in osteoclastogenesis. To elucidate the molecular mechanism of this inhibition, NFATc1 activation was assessed. NFATc1 nuclear translocation was significantly reduced in þ/R740S compared with þ/þ cells; however, this was not because of impaired enzymatic activity of calcineurin, the phosphatase responsible for NFATc1 dephosphorylation. Protein and RNA expression levels of regulator of calcineurin 1 (RCAN1), an endogenous inhibitor of NFATc1 activation and a protein degraded in lysosomes, were not significantly different between þ/R740S and þ/þ osteoclasts, but the RCAN1/NFATc1 ratio was significantly higher in þ/R740S versus þ/þ cells. The lysosomal inhibitor chloroquine significantly increased RCAN1 accumulation in þ/þ cells, consistent with the hypothesis that higher lysosomal pH impairs RCAN1 degradation, leading to a higher RCAN1/NFATc1 ratio and consequently NFATc1 inhibition. Our data indicate that increased lysosomal pH in osteoclasts leads to decreased NFATc1 signaling and nuclear translocation, resulting in a cell autonomous impairment of osteoclastogenesis in vitro. ß

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Degradation of regulator of calcineurin 1 (RCAN1) is mediated by both chaperone‐mediated autophagy and ubiquitin proteasome pathways

Cited by 115 publications

References 74 publications

STUB1/CHIP is required for HIF1A degradation by chaperone-mediated autophagy

STUB1/CHIP is required for HIF1A degradation by chaperone-mediated autophagy

Chaperone-mediated autophagy: roles in disease and aging

The R740S mutation in the V-ATPase a3 subunit increases lysosomal pH, impairs NFATc1 translocation, and decreases in vitro osteoclastogenesis

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