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The P-type ATPase transporter ATP7A promotes angiogenesis by limiting autophagic degradation of VEGFR2.

ABSTRACT: VEGFR2 (KDR/Flk1) signaling in endothelial cells (ECs) plays a central role in angiogenesis. The P-type ATPase transporter ATP7A regulates copper homeostasis, and its role in VEGFR2 signaling and angiogenesis is entirely unknown. Here, we describe the unexpected crosstalk between the Copper transporter ATP7A, autophagy, and VEGFR2 degradation. The functional significance of this Copper transporter was demonstrated by the finding that inducible EC-specific ATP7A deficient mice or ATP7A-dysfunctional ATP7Amut mice showed impaired post-ischemic neovascularization. In ECs, loss of ATP7A inhibited VEGF-induced VEGFR2 signaling and angiogenic responses, in part by promoting ligand-induced VEGFR2 protein degradation. Mechanistically, VEGF stimulated ATP7A translocation from the trans-Golgi network to the plasma membrane where it bound to VEGFR2, which prevented autophagy-mediated lysosomal VEGFR2 degradation by inhibiting autophagic cargo/adapter p62/SQSTM1 binding to ubiquitinated VEGFR2. Enhanced autophagy flux due to ATP7A dysfunction in vivo was confirmed by autophagy reporter CAG-ATP7Amut -RFP-EGFP-LC3 transgenic mice. In summary, our study uncovers a novel function of ATP7A to limit autophagy-mediated degradation of VEGFR2, thereby promoting VEGFR2 signaling and angiogenesis, which restores perfusion recovery and neovascularization. Thus, endothelial ATP7A is identified as a potential therapeutic target for treatment of ischemic cardiovascular diseases.

SUBMITTER: Ash D

PROVIDER: S-EPMC8149886 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:OBJECTIVE:Copper (Cu) is essential micronutrient, and its dysregulation is implicated in aortic aneurysm (AA) development. The Cu exporter ATP7A (copper-transporting P-type ATPase/Menkes ATPase) delivers Cu via the Cu chaperone Atox1 (antioxidant 1) to secretory Cu enzymes, such as lysyl oxidase, and excludes excess Cu. Lysyl oxidase is shown to protect against AA formation. However, the role and mechanism of ATP7A in AA pathogenesis remain unknown. Approach and Results: Here, we show that Cu chelator markedly inhibited Ang II (angiotensin II)-induced abdominal AA (AAA) in which ATP7A expression was markedly downregulated. Transgenic ATP7A overexpression prevented Ang II-induced AAA formation. Conversely, Cu transport dysfunctional ATP7Amut/+/ApoE-/- mice exhibited robust AAA formation and dissection, excess aortic Cu accumulation as assessed by X-ray fluorescence microscopy, and reduced lysyl oxidase activity. In contrast, AAA formation was not observed in Atox1-/-/ApoE-/- mice, suggesting that decreased lysyl oxidase activity, which depends on both ATP7A and Atox1, was not sufficient to develop AAA. Bone marrow transplantation suggested importance of ATP7A in vascular cells, not bone marrow cells, in AAA development. MicroRNA (miR) array identified miR-125b as a highly upregulated miR in AAA from ATP7Amut/+/ApoE-/- mice. Furthermore, miR-125b target genes (histone methyltransferase Suv39h1 and the NF-?B negative regulator TNFAIP3 [tumor necrosis factor alpha induced protein 3]) were downregulated, which resulted in increased proinflammatory cytokine expression, aortic macrophage recruitment, MMP (matrix metalloproteinase)-2/9 activity, elastin fragmentation, and vascular smooth muscle cell loss in ATP7Amut/+/ApoE-/- mice and reversed by locked nucleic acid-anti-miR-125b infusion. CONCLUSIONS:ATP7A downregulation/dysfunction promotes AAA formation via upregulating miR-125b, which augments proinflammatory signaling in a Cu-dependent manner. Thus, ATP7A is a potential therapeutic target for inflammatory vascular disease.

Project description:Background: Metastasis is the leading cause of death in colorectal cancer (CRC) patients. It is regulated mainly by tumor cell angiogenesis, and angiogenesis is caused by the binding of vascular endothelial growth factor (VEGF) to vascular endothelial growth factor receptor 2 (VEGFR2). Tumor necrosis factor-α-induced protein 8 (TNFAIP8, hereto after TIPE) plays an important role in tumorigenesis, development, and prognosis. However, the relationship between TIPE and VEGFR2 in CRC angiogenesis and the mechanism of action remain unknown. Method: In this study, we used quantitative real-time PCR, Western blotting and immunohistochemistry to detect TIPE and VEGFR2 expression in 55 specimens from CRC patients. We also used HCT116 CRC cells and human umbilical vein endothelial cells (HUVECs) for in vitro experiments by stably transducing shTIPE and shRNA control lentivirus into HCT116 cells, detecting VEGFR2 expression after TIPE knockdown and repurposing the culture supernatant as conditioned medium to stimulate angiogenesis of HUVECs. In vivo experiments with chicken chorioallantoic membranes (CAMs) and a nude mouse matrix subcutaneous tumor model were performed to validate the effects of TIPE on angiogenesis. Additionally, we analyzed the expression and phosphorylation levels of PDK1 and blocked PDK1 expression using inhibitors to determine whether TIPE-induced changes in VEGFR2-mediated angiogenesis acted via the PI3K-Akt pathway. Results: We found that TIPE and VEGFR2 are highly expressed in CRC and act as oncogenes. TIPE knockdown also downregulated VEGFR2 expression, which resulted in simultaneous inhibition of cell proliferation, cell migration and angiogenesis. Then, in vivo experiments further demonstrated that TIPE promotes angiogenesis in CRC. Finally, we found that TIPE promotes VEGFR2-mediated angiogenesis by upregulating PDK1 expression and phosphorylation and that blocking PDK1 expression can inhibit this process. Conclusion: TIPE promotes angiogenesis in CRC by regulating the expression of VEGFR2, which may be a target for antiangiogenic cancer therapy.

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The P-type ATPase transporter ATP7A promotes angiogenesis by limiting autophagic degradation of VEGFR2.

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