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HIV-1 Tat C phosphorylates VE-cadherin complex and increases human brain microvascular endothelial cell permeability.

Mishra R, Singh SK - BMC Neurosci (2014)

Bottom Line: We exposed hBMVECs to recombinant HIV-1 clade C Tat protein to study the effect of HIV-1 Tat C on permeability of hBMVECs.Redox-sensitive kinase; PYK2 activation led to increased tyrosine phosphorylation of VE-cadherin and β-catenin, leading to disruption of junctional assembly.Unrestricted phosphorylation of junctional proteins in hBMVECs, in response to HIV-1 Tat C protein; leads to the disruption of junctional complexes and increased endothelial permeability.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Neurovirology and Inflammation Biology, CSIR-Centre for Cellular and Molecular Biology (CCMB), Uppal Road, Hyderabad 500007, India. sunitsingh2000@gmail.com.

ABSTRACT

Background: Human brain microvascular endothelial cells (hBMVECs) are integral part of the blood brain barrier. Post-translational modifications of adherens junction proteins regulate the permeability of human brain microvascular endothelial cells. Pro-inflammatory signals can induce tyrosine phosphorylation of adherens junction proteins. The primary objective of this work is to provide a molecular model; how the HIV-1 Tat protein can compromise the BBB integrity and eventually lead to neurological consequences. We exposed hBMVECs to recombinant HIV-1 clade C Tat protein to study the effect of HIV-1 Tat C on permeability of hBMVECs. Trans-endothelial electrical resistance and fluorescent dye migration assay have been used to check the permeability of hBMVECs. DCFDA staining has been used for intracellular reactive oxygen species (ROS) detection. Western blotting has been used to study the expression levels and co-immunoprecipitation has been used to study the interactions among adherens junction proteins.

Results: HIV-1 Tat C protein induced NOX2 and NOX4 expression level and increased intracellular ROS level. Redox-sensitive kinase; PYK2 activation led to increased tyrosine phosphorylation of VE-cadherin and β-catenin, leading to disruption of junctional assembly. The dissociation of tyrosine phosphatases VE-PTP and SHP2 from cadherin complex resulted into increased tyrosine phosphorylation of VE-cadherin and β-catenin in HIV-1 Tat C treated hBMVECs.

Conclusion: Unrestricted phosphorylation of junctional proteins in hBMVECs, in response to HIV-1 Tat C protein; leads to the disruption of junctional complexes and increased endothelial permeability.

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Related in: MedlinePlus

HIV-1 Tat C mediated tyrosine phosphorylation of VE-cadherin complex and increase in permeability of hBMVECs. HIV-1 Tat C treatment induces the intracellular ROS generation by activating NADPH oxidases (NOX2/NOX4) proteins in hBMVECs. Elevated intracellular levels of ROS activate the levels of redox sensitive kinase PYK2, which led to increased tyrosine phosphorylation of β-catenin and VE-cadherin. HIV-1 Tat C treatment also downregulates major phosphatases VE-PTP and SHP2 and leads to their dissociation from VE-cadherin complex. These shift in PTK/ PTP ratio in hBMVECs after HIV-1 Tat C exposure leads to destabilized junctional assembly and thereby enhanced endothelial permeability.
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Figure 8: HIV-1 Tat C mediated tyrosine phosphorylation of VE-cadherin complex and increase in permeability of hBMVECs. HIV-1 Tat C treatment induces the intracellular ROS generation by activating NADPH oxidases (NOX2/NOX4) proteins in hBMVECs. Elevated intracellular levels of ROS activate the levels of redox sensitive kinase PYK2, which led to increased tyrosine phosphorylation of β-catenin and VE-cadherin. HIV-1 Tat C treatment also downregulates major phosphatases VE-PTP and SHP2 and leads to their dissociation from VE-cadherin complex. These shift in PTK/ PTP ratio in hBMVECs after HIV-1 Tat C exposure leads to destabilized junctional assembly and thereby enhanced endothelial permeability.

Mentions: HIV-1 Tat C mediated activation of NADPH oxidases (NOX2/ NOX4) led to the increased generation of ROS in hBMVECs and activation of PYK2 kinase. Upregulation of PYK2 activity resulted into the increased tyrosine phosphorylated forms of β-catenin and VE-cadherin. The dissociation of VE-PTP and SHP2 from VE-cadherin complex increased the tyrosine phosphorylation of VE-cadherin/β-catenin, which led to disrupted physical association of AJPs within VE-cadherin complex at endothelial junctions. These conformational alterations increased the permeability in hBMVECs. ROS scavenging as well as inhibition of the PYK2 activity showed a significant potential in combating the HIV-1 Tat C mediated disruption of endothelial permeability (Figure 8). This study suggests the involvement of post-translational modifications of AJPs in the disruption of barrier integrity in hBMVECs by HIV-1 Tat C protein. This finding will be helpful in understanding the effect of viral proteins in a bystander fashion on human brain microvascular endothelial cells.


HIV-1 Tat C phosphorylates VE-cadherin complex and increases human brain microvascular endothelial cell permeability.

Mishra R, Singh SK - BMC Neurosci (2014)

HIV-1 Tat C mediated tyrosine phosphorylation of VE-cadherin complex and increase in permeability of hBMVECs. HIV-1 Tat C treatment induces the intracellular ROS generation by activating NADPH oxidases (NOX2/NOX4) proteins in hBMVECs. Elevated intracellular levels of ROS activate the levels of redox sensitive kinase PYK2, which led to increased tyrosine phosphorylation of β-catenin and VE-cadherin. HIV-1 Tat C treatment also downregulates major phosphatases VE-PTP and SHP2 and leads to their dissociation from VE-cadherin complex. These shift in PTK/ PTP ratio in hBMVECs after HIV-1 Tat C exposure leads to destabilized junctional assembly and thereby enhanced endothelial permeability.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4230799&req=5

Figure 8: HIV-1 Tat C mediated tyrosine phosphorylation of VE-cadherin complex and increase in permeability of hBMVECs. HIV-1 Tat C treatment induces the intracellular ROS generation by activating NADPH oxidases (NOX2/NOX4) proteins in hBMVECs. Elevated intracellular levels of ROS activate the levels of redox sensitive kinase PYK2, which led to increased tyrosine phosphorylation of β-catenin and VE-cadherin. HIV-1 Tat C treatment also downregulates major phosphatases VE-PTP and SHP2 and leads to their dissociation from VE-cadherin complex. These shift in PTK/ PTP ratio in hBMVECs after HIV-1 Tat C exposure leads to destabilized junctional assembly and thereby enhanced endothelial permeability.
Mentions: HIV-1 Tat C mediated activation of NADPH oxidases (NOX2/ NOX4) led to the increased generation of ROS in hBMVECs and activation of PYK2 kinase. Upregulation of PYK2 activity resulted into the increased tyrosine phosphorylated forms of β-catenin and VE-cadherin. The dissociation of VE-PTP and SHP2 from VE-cadherin complex increased the tyrosine phosphorylation of VE-cadherin/β-catenin, which led to disrupted physical association of AJPs within VE-cadherin complex at endothelial junctions. These conformational alterations increased the permeability in hBMVECs. ROS scavenging as well as inhibition of the PYK2 activity showed a significant potential in combating the HIV-1 Tat C mediated disruption of endothelial permeability (Figure 8). This study suggests the involvement of post-translational modifications of AJPs in the disruption of barrier integrity in hBMVECs by HIV-1 Tat C protein. This finding will be helpful in understanding the effect of viral proteins in a bystander fashion on human brain microvascular endothelial cells.

Bottom Line: We exposed hBMVECs to recombinant HIV-1 clade C Tat protein to study the effect of HIV-1 Tat C on permeability of hBMVECs.Redox-sensitive kinase; PYK2 activation led to increased tyrosine phosphorylation of VE-cadherin and β-catenin, leading to disruption of junctional assembly.Unrestricted phosphorylation of junctional proteins in hBMVECs, in response to HIV-1 Tat C protein; leads to the disruption of junctional complexes and increased endothelial permeability.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Neurovirology and Inflammation Biology, CSIR-Centre for Cellular and Molecular Biology (CCMB), Uppal Road, Hyderabad 500007, India. sunitsingh2000@gmail.com.

ABSTRACT

Background: Human brain microvascular endothelial cells (hBMVECs) are integral part of the blood brain barrier. Post-translational modifications of adherens junction proteins regulate the permeability of human brain microvascular endothelial cells. Pro-inflammatory signals can induce tyrosine phosphorylation of adherens junction proteins. The primary objective of this work is to provide a molecular model; how the HIV-1 Tat protein can compromise the BBB integrity and eventually lead to neurological consequences. We exposed hBMVECs to recombinant HIV-1 clade C Tat protein to study the effect of HIV-1 Tat C on permeability of hBMVECs. Trans-endothelial electrical resistance and fluorescent dye migration assay have been used to check the permeability of hBMVECs. DCFDA staining has been used for intracellular reactive oxygen species (ROS) detection. Western blotting has been used to study the expression levels and co-immunoprecipitation has been used to study the interactions among adherens junction proteins.

Results: HIV-1 Tat C protein induced NOX2 and NOX4 expression level and increased intracellular ROS level. Redox-sensitive kinase; PYK2 activation led to increased tyrosine phosphorylation of VE-cadherin and β-catenin, leading to disruption of junctional assembly. The dissociation of tyrosine phosphatases VE-PTP and SHP2 from cadherin complex resulted into increased tyrosine phosphorylation of VE-cadherin and β-catenin in HIV-1 Tat C treated hBMVECs.

Conclusion: Unrestricted phosphorylation of junctional proteins in hBMVECs, in response to HIV-1 Tat C protein; leads to the disruption of junctional complexes and increased endothelial permeability.

Show MeSH
Related in: MedlinePlus