Chaperone-mediated Autophagy Targets Hypoxia-inducible Factor-1α (HIF-1α) for Lysosomal Degradation

Hubbi, Maimon E.; Hu, Hongxia; Kshitiz, .; Ahmed, Ishrat; Levchenko, Andre; Semenza, Gregg L.

doi:10.1074/jbc.m112.414771

Cited by 202 publications

(187 citation statements)

References 58 publications

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“…Mechanisms by which HIF-1α is regulated in an O 2 -independent manner have also been identified (16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26). In addition to proteasome-dependent pathways for HIF-1α degradation, we identified a pathway by which HIF-1α can be targeted for lysosomal degradation through chaperonemediated autophagy (27), which subsequently was confirmed by others (28)(29)(30).…”

supporting

confidence: 48%

“…Cells were plated on gelatin-coated glass-bottomed plates (Live Assay). Posttreatment, samples were washed with ice-cold PBS, fixed with 4% (wt/vol) paraformaldehyde for 20 min at room temperature, permeabilized with 0.05% Triton X-100 for 15 min, washed twice with PBS, and blocked with 10% (vol/vol) goat serum and 1% AlbuMAX (Invitrogen) for 1 h. Samples were incubated with rabbit polyclonal anti-MCM5 (Santa Cruz) and sheep polyclonal anti-BrdU (Abcam) primary antibodies for 1 h, washed, and incubated with Alexa Fluor-conjugated secondary antibodies (Invitrogen) for 1 h (27). Samples were washed and mounted on microscope slides with a drop of SlowFade (Invitrogen) and sealed with Vectashield (Vector Labs).…”

Section: Methodsmentioning

confidence: 99%

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Cyclin-dependent kinases regulate lysosomal degradation of hypoxia-inducible factor 1α to promote cell-cycle progression

Hubbi¹,

Gilkes

Hu³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that mediates adaptive responses to oxygen deprivation. In addition, the HIF-1α subunit has a nontranscriptional role as a negative regulator of DNA replication through effects on minichromosome maintenance helicase loading and activation. However, some cell types continue to replicate under hypoxic conditions. The mechanism by which these cells maintain proliferation in the presence of elevated HIF-1α levels is unclear. Here we report that HIF-1α physically and functionally interacts with cyclin-dependent kinase 1 (Cdk1) and Cdk2. Cdk1 activity blocks lysosomal degradation of HIF-1α and increases HIF-1α protein stability and transcriptional activity. By contrast, Cdk2 activity promotes lysosomal degradation of HIF-1α at the G1/S phase transition. Blocking lysosomal degradation by genetic or pharmacological means leads to HIF-1α-dependent cell-cycle arrest, demonstrating that lysosomal degradation of HIF-1α is an essential step for the maintenance of cell-cycle progression under hypoxic conditions. cell proliferation | chaperone mediated autophagy | lysosome

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supporting

confidence: 48%

Section: Methodsmentioning

confidence: 99%

Cyclin-dependent kinases regulate lysosomal degradation of hypoxia-inducible factor 1α to promote cell-cycle progression

Hubbi¹,

Gilkes

Hu³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…7D and E). 53,54 To further validate this finding in vivo, we used NP-specific Hif1a knockout mice generated by crossing Hif1a floxed mice with mice expressing Cre recombinase under control of the notochord specific Foxa2 promoter/enhancer (Fig. 7F).…”

Section: Hypoxic Induction Of Autophagy In Np Cells Is Independent Ofmentioning

confidence: 99%

Hypoxia promotes noncanonical autophagy in nucleus pulposus cells independent of MTOR and HIF1A signaling

et al. 2016

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Nucleus pulposus (NP) cells reside in the avascular and hypoxic microenvironment of intervertebral discs. Importantly, many activities related to survival and function of NP cells are controlled by the HIF-family of transcription factors. We hypothesize that NP cells adapt to their hypoxic niche through modulation of macroautophagy/autophagy. In various cell types, hypoxia induces autophagy in a HIF1A-dependent fashion; however, little is known about hypoxic regulation of autophagy in NP cells. Hypoxia increases the number of autophagosomes as seen by TEM analysis and LC3-positive puncta in NP cells. Hypoxic induction of autophagy was also demonstrated by a significantly higher number of autophagosomes and smaller change in autolysosomes in NP cells expressing tandem-mCherry-EGFP-LC3B. Increased LC3-II levels were not accompanied by a concomitant increase in BECN1 or the ATG12-ATG5 complex. In addition, ULK1 phosphorylation at Ser757 and Ser777 responsive to MTOR and AMPK, respectively, was not affected in hypoxia. Interestingly, when MTOR activity was inhibited by rapamycin or Torin1, LC3-II levels did not change, suggesting a novel MTOR-independent regulation. Noteworthy, while silencing of HIF1A affected hypoxic induction of BNIP3, it did not affect LC3-II levels, indicating hypoxia-induced autophagy is HIF1-independent. Importantly, there was no change in the number of LC3-positive autophagosomes in NP-specific Hif1a null mice. Finally, inhibition of autophagic flux did not affect the glycolytic metabolism of NP cells, suggesting a possible nonmetabolic role of autophagy. Taken together, our study for the first time shows that NP cells regulate autophagy in a noncanonical fashion independent of MTOR and HIF1A signaling.

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“…12 More recently, it was demonstrated that HIF1A is a substrate for chaperone-mediated autophagy (CMA), however the molecular mechanisms and players involved were not elucidated. 13 Nevertheless, degradation of typical proteasomal substrates by the lysosomes remains poorly understood. Although the pathways for lysosomal and proteasomal degradation of substrates are mechanistically different, they share common traits of specificity toward substrates.…”

Section: Introductionmentioning

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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Chaperone-mediated Autophagy Targets Hypoxia-inducible Factor-1α (HIF-1α) for Lysosomal Degradation

Cited by 202 publications

References 58 publications

Cyclin-dependent kinases regulate lysosomal degradation of hypoxia-inducible factor 1α to promote cell-cycle progression

Cyclin-dependent kinases regulate lysosomal degradation of hypoxia-inducible factor 1α to promote cell-cycle progression

Hypoxia promotes noncanonical autophagy in nucleus pulposus cells independent of MTOR and HIF1A signaling

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

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