scholarly journals Role of Ran-regulated nuclear-cytoplasmic trafficking of pVHL in the regulation of microtubular stability-mediated HIF-1α in hypoxic cardiomyocytes

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Xupin Jiang ◽  
Dongxia Zhang ◽  
Hengshu Zhang ◽  
Yuesheng Huang ◽  
Miao Teng
Keyword(s):  
Nuclear Export ◽  
Hypoxia Inducible Factor ◽  
Microtubule Network ◽  
Nuclear Compartment ◽  
Von Hippel Lindau ◽  
Microtubule Stabilization ◽  
Microtubule Disruption ◽  
Underlying Mechanisms ◽  
Induced Changes

Abstract Our previous study suggested that microtubule network alteration affects the process of glycolysis in cardiomyocytes (CMs) via the regulation of hypoxia-inducible factor (HIF)-1α during the early stages of hypoxia. However, little is known regarding the underlying mechanisms of microtubule network alteration-induced changes of HIF-1α. The von Hippel–Lindau tumor suppressor protein (pVHL) has been shown to mediate the ubiquitination of HIF-1α in the nuclear compartment prior to HIF-1α exportation to the cytoplasm and pVHL dynamic nuclear-cytoplasmic trafficking is indicated to be involved in the process of HIF-1α degradation. In this study, by administering different microtubule-stabilizing and -depolymerizing interventions, we demonstrated that microtubule stabilization promoted pVHL nuclear export and drove the translocation of pVHL to the cytoplasm, while microtubule disruption prevented pVHL nuclear export in hypoxic CMs. Moreover, the ratio between nuclear and cytoplasmic pVHL was associated with HIF-1α regulation. Importantly, microtubule network alteration also affected the subcellular localization of Ran, which was involved in the regulation of pVHL nuclear-cytoplasmic trafficking. The above results suggest that the subcellular translocation of pVHL plays an important role in microtubular structure alteration-induced HIF-1α regulation. Interestingly, Ran is involved in the process of pVHL nuclear-cytoplasmic trafficking following microtubule network alteration in hypoxic CMs.

scholarly journals Oxygen-Dependent Ubiquitination and Degradation of Hypoxia-Inducible Factor Requires Nuclear-Cytoplasmic Trafficking of the von Hippel-Lindau Tumor Suppressor Protein

2002 ◽  
Vol 22 (15) ◽  
pp. 5319-5336 ◽  
Author(s):  
Isabelle Groulx ◽  
Stephen Lee
Keyword(s):  
Tumor Suppressor ◽  
Rna Polymerase Ii ◽  
Nuclear Export ◽  
Hypoxia Inducible Factor ◽  
Tumor Suppressor Protein ◽  
Nuclear Compartment ◽  
Von Hippel Lindau ◽  
Substrate Protein ◽  
Subsequent Degradation ◽  
Substrate Proteins

ABSTRACT It is becoming increasingly evident that the degradation of nuclear proteins requires nuclear-cytoplasmic trafficking of both the substrate proteins, as well as the E3 ubiquitin-ligases. Here, we show that nuclear-cytoplasmic trafficking of the von Hippel-Lindau tumor suppressor protein (VHL) is required for oxygen-dependent ubiquitination and degradation of the alpha subunits of hypoxia-inducible factor (HIF-α). VHL engages in a constitutive transcription-sensitive nuclear-cytoplasmic shuttle unaffected by oxygen tension or levels of nuclear substrate HIF-α. Ubiquitinated forms of HIF-α, as well as VHL/ubiquitinated HIF-α complexes, are found solely in the nuclear compartment of normoxic or reoxygenated VHL-competent cells. HIF-α localizes exclusively in the nucleus of hypoxic cells but is exported to the cytoplasm upon reoxygenation. Oxygen-dependent nuclear ubiquitination and nuclear export of HIF-α can be prevented by treatment with an HIF-specific prolyl hydroxylase inhibitor. Treatment with inhibitors of RNA polymerase II activity, which interfere with the ability of VHL to engage in nuclear export, also prevents cytoplasmic accumulation of HIF-α in reoxygenated cells. This caused a marked increase in the HIF-α half-life without affecting its nuclear ubiquitination. We present a model by which VHL-mediated ubiquitination of HIF-α and its subsequent degradation are dependent upon dynamic nuclear-cytoplasmic trafficking of both the E3 ubiquitin-ligase and the nuclear substrate protein.

scholarly journals Cancer-Causing Mutations in a Novel Transcription-Dependent Nuclear Export Motif of VHL Abrogate Oxygen-Dependent Degradation of Hypoxia-Inducible Factor

2007 ◽  
Vol 28 (1) ◽  
pp. 302-314 ◽  
Author(s):  
Mireille Khacho ◽  
Karim Mekhail ◽  
Karine Pilon-Larose ◽  
Josianne Payette ◽  
Stephen Lee
Keyword(s):  
Nuclear Export ◽  
Hypoxia Inducible Factor ◽  
Subcellular Trafficking ◽  
E3 Ligases ◽  
Von Hippel Lindau ◽  
Export Dynamics ◽  
Naturally Occurring ◽  
Key Residues ◽  
Vhl Tumor Suppressor Protein

ABSTRACT It is thought that degradation of nuclear proteins by the ubiquitylation system requires nuclear-cytoplasmic trafficking of E3 ubiquitin ligases. The von Hippel-Lindau (VHL) tumor suppressor protein is the substrate recognition component of a Cullin-2-containing E3 ubiquitin ligase that recruits hypoxia-inducible factor (HIF) for oxygen-dependent degradation. We demonstrated that VHL engages in nuclear-cytoplasmic trafficking that requires ongoing transcription to promote efficient HIF degradation. Here, we report the identification of a discreet motif, DXGX2DX2L, that directs transcription-dependent nuclear export of VHL and which is targeted by naturally occurring mutations associated with renal carcinoma and polycythemia in humans. The DXGX2DX2L motif is also found in other proteins, including poly(A)-binding protein 1, to direct its transcription-dependent nuclear export. We define DXGX2DX2L as TD-NEM (transcription-dependent nuclear export motif), since inhibition of transcription by actinomycin D or 5,6-dichlorobenzimidazole abrogates its nuclear export activity. Disease-causing mutations of key residues of TD-NEM restrain the ability of VHL to efficiently mediate oxygen-dependent degradation of HIF by altering its nuclear export dynamics without affecting interaction with its substrate. These results identify a novel nuclear export motif, further highlight the role of nuclear-cytoplasmic shuttling of E3 ligases in degradation of nuclear substrates, and provide evidence that disease-causing mutations can target subcellular trafficking.

scholarly journals Hypoxia-inducible factor (HIF): fuel for cancer progression

2021 ◽  
Vol 14 ◽  
Author(s):  
Saurabh Satija ◽  
Harpreet Kaur ◽  
Murtaza M. Tambuwala ◽  
Prabal Sharma ◽  
Manish Vyas ◽  
...  
Keyword(s):  
Biological Sciences ◽  
Tumor Microenvironment ◽  
Tumor Cells ◽  
Cancer Progression ◽  
Oxygen Deficiency ◽  
Hypoxia Inducible Factor ◽  
Transcription Factor Activation ◽  
Underlying Mechanisms ◽  
Adaptive Reactions

Hypoxia is an integral part of tumor microenvironment, caused primarily due to rapidly multiplying tumor cells and a lack of proper blood supply. Among the major hypoxic pathways, HIF-1 transcription factor activation is one of the widely investigated pathways in the hypoxic tumor microenvironment (TME). HIF-1 is known to activate several adaptive reactions in response to oxygen deficiency in tumor cells. HIF-1 has two subunits, HIF-1β (constitutive) and HIF-1α (inducible). The HIF-1α expression is largely regulated via various cytokines (through PI3K-ACT-mTOR signals), which involves the cascading of several growth factors and oncogenic cascades. These events lead to the loss of cellular tumor suppressant activity through changes in the level of oxygen via oxygen-dependent and oxygen-independent pathways. The significant and crucial role of HIF in cancer progression and its underlying mechanisms have gained much attention lately among the translational researchers in the fields of cancer and biological sciences, which have enabled them to correlate these mchanisms with various other disease modalities. In the present review, we have summarized the key findings related to the role of HIF in the progression of tumors.

Centrosome cohesion is regulated by a balance of kinase and phosphatase activities

2001 ◽  
Vol 114 (20) ◽  
pp. 3749-3757 ◽  
Author(s):  
Patrick Meraldi ◽  
Erich A. Nigg
Keyword(s):  
Protein Phosphatase ◽  
Kinase Activity ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Microtubule Depolymerization ◽  
Drug Induced ◽  
Microtubule Network ◽  
Similar Phenotype ◽  
Underlying Mechanisms

Centrosome cohesion and separation are regulated throughout the cell cycle, but the underlying mechanisms are not well understood. Since overexpression of a protein kinase, Nek2, is able to trigger centrosome splitting (the separation of parental centrioles), we have surveyed a panel of centrosome-associated kinases for their ability to induce a similar phenotype. Cdk2, in association with either cyclin A or E, was as effective as Nek2, but several other kinases tested did not significantly interfere with centrosome cohesion. Centrosome splitting could also be triggered by inhibition of phosphatases, and protein phosphatase 1α (PP1α) was identified as a likely physiological antagonist of Nek2. Furthermore, we have revisited the role of the microtubule network in the control of centrosome cohesion. We could confirm that microtubule depolymerization by nocodazole causes centrosome splitting. Surprisingly, however, this drug-induced splitting also required kinase activity and could specifically be suppressed by a dominant-negative mutant of Nek2. These studies highlight the importance of protein phosphorylation in the control of centrosome cohesion, and they point to Nek2 and PP1α as critical regulators of centrosome structure.

scholarly journals Long-Noncoding RNA (lncRNA) in the Regulation of Hypoxia-Inducible Factor (HIF) in Cancer

2020 ◽  
Vol 6 (3) ◽  
pp. 27 ◽  
Author(s):  
Dominik A. Barth ◽  
Felix Prinz ◽  
Julia Teppan ◽  
Katharina Jonas ◽  
Christiane Klec ◽  
...  
Keyword(s):  
Noncoding Rna ◽  
Molecular Mechanisms ◽  
Hypoxia Inducible Factor ◽  
Protein Coding ◽  
Rna Molecules ◽  
Von Hippel Lindau ◽  
Hypoxic Conditions ◽  
Main Regulator ◽  
Upstream Regulators

Hypoxia is dangerous for oxygen-dependent cells, therefore, physiological adaption to cellular hypoxic conditions is essential. The transcription factor hypoxia-inducible factor (HIF) is the main regulator of hypoxic metabolic adaption reducing oxygen consumption and is regulated by gradual von Hippel-Lindau (VHL)-dependent proteasomal degradation. Beyond physiology, hypoxia is frequently encountered within solid tumors and first drugs are in clinical trials to tackle this pathway in cancer. Besides hypoxia, cancer cells may promote HIF expression under normoxic conditions by altering various upstream regulators, cumulating in HIF upregulation and enhanced glycolysis and angiogenesis, altogether promoting tumor proliferation and progression. Therefore, understanding the underlying molecular mechanisms is crucial to discover potential future therapeutic targets to evolve cancer therapy. Long non-coding RNAs (lncRNA) are a class of non-protein coding RNA molecules with a length of over 200 nucleotides. They participate in cancer development and progression and might act as either oncogenic or tumor suppressive factors. Additionally, a growing body of evidence supports the role of lncRNAs in the hypoxic and normoxic regulation of HIF and its subunits HIF-1α and HIF-2α in cancer. This review provides a comprehensive update and overview of lncRNAs as regulators of HIFs expression and activation and discusses and highlights potential involved pathways.

Phosphatidylinositol 3′-kinase signaling pathway is essential for Rac1-induced hypoxia-inducible factor-1α and vascular endothelial growth factor expression

2011 ◽  
Vol 300 (6) ◽  
pp. H2169-H2176 ◽  
Author(s):  
Yan Xue ◽  
Nan-Lin Li ◽  
Jing-Yue Yang ◽  
Yan Chen ◽  
Lu-Lu Yang ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Tumor Suppressor ◽  
Active Form ◽  
Hypoxia Inducible Factor ◽  
Pi3k Pathway ◽  
Rho Proteins ◽  
Phosphatidylinositol 3 Kinase ◽  
Von Hippel Lindau ◽  
Phosphatidylinositol 3

We have previously demonstrated the roles of RhoA, Rac1, and Cdc42 in hypoxia-driven angiogenesis. However, the role of oncogenes in hypoxia signaling is poorly understood. Given the importance of Rho proteins in the hypoxic response, we hypothesized that Rho family members could act as mediators of hypoxic signal transduction. We investigated the cross-talk between hypoxia and oncogene-driven signal transduction pathways and explored the role of Rac1 on hypoxia-induced hypoxia-inducible factor (HIF)-1α and VEGF expression. Since the phosphatidylinositol 3′-kinase (PI3K) pathway is involved in signal transduction of many oncogenes, we explored the role of PI3K on Rac1-mediated expression of HIF-1α and VEGF in hypoxia. We showed that LY-294002, a PI3K inhibitor, suppressed HIF-1α and VEGF induction under hypoxic conditions by up to 50%. Activation of Rac1 resulted in an upregulation of hypoxia-induced HIF-1α expression, which was blocked by LY-294002. These data suggested that Rac1 is an intermediate in the PI3K-mediated induction of HIF-1α. Interestingly, there was a significant downregulation of the tumor suppressor genes p53 and von Hippel-Lindau tumor suppressor (VHL) in cells expressing a constitutively active form of Rac1. Rac1-mediated inhibition of p53 and VHL could therefore be implicated in the upregulation of HIF-1α expression.

The role of iron and 2-oxoglutarate oxygenases in signalling

2003 ◽  
Vol 31 (3) ◽  
pp. 510-515 ◽  
Author(s):  
K.S. Hewitson ◽  
L.A. McNeill ◽  
J.M. Elkins ◽  
C.J. Schofield
Keyword(s):  
Transcriptional Activation ◽  
Hypoxia Inducible Factor ◽  
Transcriptional Response ◽  
Transactivation Domain ◽  
Von Hippel Lindau ◽  
Helix Loop Helix ◽  
Hypoxic Conditions ◽  
Tumour Suppressor Protein ◽  
Dependent Modification

Sensing of ambient dioxygen levels and appropriate feedback mechanisms are essential processes for all multicellular organisms. In animals, moderate hypoxia causes an increase in the transcription levels of specific genes, including those encoding vascular endothelial growth factor and erythropoietin. The hypoxic response is mediated by hypoxia-inducible factor (HIF), an αβ heterodimeric transcription factor in which both the HIF subunits are members of the basic helix–loop–helix PAS (PER-ARNT-SIM) domain family. Under hypoxic conditions, levels of HIFα rise, allowing dimerization with HIFβ and initiating transcriptional activation. Two types of dioxygen-dependent modification to HIFα have been identified, both of which inhibit the transcriptional response. Firstly, HIFα undergoes trans-4-hydroxylation at two conserved proline residues that enable its recognition by the von Hippel-Lindau tumour-suppressor protein. Subsequent ubiquitinylation, mediated by an ubiquitin ligase complex, targets HIFα for degradation. Secondly, hydroxylation of an asparagine residue in the C-terminal transactivation domain of HIFα directly prevents its interaction with the co-activator p300. Hydroxylation of HIFα is catalysed by enzymes of the iron(II)- and 2-oxoglutarate-dependent dioxygenase family. In humans, three prolyl hydroxylase isoenzymes (PHD1–3) and an asparagine hydroxylase [factor inhibiting HIF (FIH)] have been identified. The role of 2-oxoglutarate oxygenases in the hypoxic and other signalling pathways is discussed.

scholarly journals Activation of the Hypoxia-Inducible Factor Pathway Expands Early Thymic Progenitors

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2896-2896
Author(s):  
Anita Hollenbeck ◽  
Stefanie Weber ◽  
Kathrin Händschke ◽  
Mandy Necke ◽  
Bertram Opalka ◽  
...  
Keyword(s):  
Cellular Response ◽  
Target Genes ◽  
Hypoxia Inducible Factor ◽  
Cell Receptor ◽  
Normal Numbers ◽  
Cellular Compartment ◽  
Vhl Gene ◽  
Von Hippel Lindau

Abstract Early thymic progenitors enter the thymus and are exposed to regional hypoxia while they develop in a step-wise manner to mature functional T-cells. Therefore, hypoxia might represent an important component of the highly specialized thymic microenvironment. On the molecular level the hypoxia-inducible factor pathway controls the cellular response to hypoxia. In this pathway, the von-Hippel-Lindau protein (pVHL) continuously mediates the destruction of the transcription factor hypoxia-inducible factor-1α (HIF-1α) under normoxic conditions. Under hypoxia HIF-1α degradation is inhibited leading to the activation of HIF-1α target genes. Others used lck-Cre transgene-mediated conditional in vivo deletion of the Vhl gene to study the role of the oxygen-sensing pathway in developing thymocytes and found normal numbers of early double-negative (DN; CD4-CD8-) thymocytes (Biju et al., Mol Cell Biol, 2004). However, lck-Cre deletion initiates at the DN3 (CD25+CD44-) stage leaving the Vhl locus of very early DN1 (CD25-CD44+), DN2 (CD25+CD44+) and DN3 thymocytes unaltered. Therefore, we here used the ubiquitous hematopoietic deleter strain vav-Cre to investigate the role of pVHL in very early thymocytes (vav-Cre;VhlloxP;loxP mice). Using a PCR-based strategy we confirmed complete deletion of the Vhl gene in this model. We observed unaltered DN1 and DN2 progenitor numbers, however in contrast to the published lck-cre-mediated system we consistently observed an up to twofold expansion of the DN3 cellular compartment. As the hypoxia-inducible factor pathway was shown to modulate NOTCH1 signaling we studied Notch1 expression on Vhl-deficient thymocytes. Strikingly, Notch1 expression was significantly increased on expanded Vhl null DN3 thymocytes. At the DN3 developmental stage selection of cells with an accurately re-arranged T-cell receptor β-locus occurs. Thus, we analyzed pre- and post-β-selection DN3 cells by CD28 staining. Interestingly, we found both pre- and post-β-selection DN3 subpopulations expanded. In order to investigate whether the progenitor expansion is mediated by the lack of HIF-1α inhibition in the Vhl-deficient context we studied DN3 thymocytes in a conditional hematopoietic HIF-1α gain-of-function model (vav-Cre;HIF1dPA). Overexpression of HIF-1α, which is insensitive to pVHL-mediated degradation in vav-Cre;HIF1dPAmice, also resulted in expanded DN3 thymocytes. In summary, we describe novel conditional models to genetically alter the hypoxia-inducible factor pathway within very early thymic progenitors. Genetic Vhl loss led to an expansion of DN3 thymocytes. This DN3 expansion is most likely due to the absence of HIF-1α-inhibition, because HIF-1α overexpression phenocopied the Vhl-deficient DN3 thymocyte expansion. Disclosures Dührsen: Celgene: Honoraria, Research Funding.

scholarly journals Calpain Mediates a von Hippel-Lindau Protein–independent Destruction of Hypoxia-inducible Factor-1α

2006 ◽  
Vol 17 (4) ◽  
pp. 1549-1558 ◽  
Author(s):  
Jie Zhou ◽  
Roman Köhl ◽  
Barbara Herr ◽  
Ronald Frank ◽  
Bernhard Brüne
Keyword(s):  
Renal Carcinoma ◽  
Hypoxia Inducible Factor ◽  
Active Role ◽  
Carcinoma Cells ◽  
No Donor ◽  
Human Embryonic Kidney Cells ◽  
Von Hippel Lindau ◽  
Hypoxia Inducible Factor 1 ◽  
Calpain Inhibitors

Hypoxia-inducible factor 1 (HIF-1) is controlled through stability regulation of its alpha subunit, which is expressed under hypoxia but degraded under normoxia. Degradation of HIF-1α requires association of the von Hippel Lindau protein (pVHL) to provoke ubiquitination followed by proteasomal digestion. Besides hypoxia, nitric oxide (NO) stabilizes HIF-1α under normoxia but destabilizes the protein under hypoxia. To understand the role of NO under hypoxia we made use of pVHL-deficient renal carcinoma cells (RCC4) that show a high steady state HIF-1α expression under normoxia. Exposing RCC4 cells to hypoxia in combination with the NO donor DETA-NO (2,2′-(hydroxynitrosohydrazono) bis-ethanimine), but not hypoxia or DETA-NO alone, decreased HIF-1α protein and attenuated HIF-1 transactivation. Mechanistically, we noticed a role of calpain because calpain inhibitors reversed HIF-1α degradation. Furthermore, chelating intracellular calcium attenuated HIF-1α destruction by hypoxia/DETA-NO, whereas a calcium increase was sufficient to lower the amount of HIF-1α even under normoxia. An active role of calpain in lowering HIF-1α amount was also evident in pVHL-containing human embryonic kidney cells when the calcium pump inhibitor thapsigargin reduced HIF-1α that was stabilized by the prolyl hydroxylase inhibitor dimethyloxalylglycine (DMOG). We conclude that calcium contributes to HIF-1α destruction involving the calpain system.

Hypoxia-inducible factor prolyl 4-hydroxylases: common and specific roles

2013 ◽  
Vol 394 (4) ◽  
pp. 435-448 ◽  
Author(s):  
Johanna Myllyharju ◽  
Peppi Koivunen
Keyword(s):  
Target Genes ◽  
Hypoxia Inducible Factor ◽  
Drug Candidate ◽  
Hypoxia Response ◽  
Von Hippel Lindau ◽  
Ubiquitin Ligase Complex ◽  
Biological Studies ◽  
The Common

Abstract Hypoxia-inducible transcription factor (HIF), an αβ dimer, is the key inducer of hypoxia-responsive genes that operate both during normal development and pathological processes in association with decreased oxygen availability. The products of HIF target genes function in, e.g., hematopoiesis, angiogenesis, iron transport, glucose utilization, resistance to oxidative stress, cell proliferation, survival and apoptosis, extracellular matrix homeostasis, and tumorigenesis and metastasis. HIF is accumulated in hypoxia, whereas it is rapidly degraded in normoxic cells. The oxygen-sensing mechanism behind this phenomenon is provided by HIF prolyl 4-hydroxylases (HIF-P4Hs, commonly known as PHDs and EglNs) that require oxygen in their reaction. In normoxia, two prolines in the oxygen-dependent degradation domain of the HIFα subunit become hydroxylated by the HIF-P4Hs. The 4-hydroxyproline residues formed serve as recognition sites for the von Hippel-Lindau E3 ubiquitin ligase complex and result in subsequent ubiquitination and instant proteasomal degradation of HIFα in normoxia. The HIF-P4H reaction is inhibited in hypoxia. HIFα evades degradation and forms a functional dimer with HIFβ, leading to activation of the HIF target genes. The central role of HIF-P4Hs in the regulation of the hypoxia response pathway has provided an attractive possibility as a drug candidate for treatment of, e.g., severe anemias and ischemic conditions, and several companies are currently carrying out clinical studies on the use of HIF-P4H inhibitors to treat anemia in patients with a kidney disease. Therefore, it is important to understand the effects of individual HIF-P4H isoenzymes on the hypoxia response and potential other pathways in vivo. The common and specific functions of the HIF-P4H isoenzymes are discussed in this review on the basis of available data from cell biological studies and gene-modified animals.

Sign in / Sign up

Export Citation Format

Share Document