Papilloma protein E6 abrogates shear stress-dependent survival in human endothelial cells: evidence for specialized functions of paxillin.
BACKGROUND
To investigate how endothelial cells transduce intracellular signals in response to laminar shear stress (SS), we made use of the papilloma virus oncoprotein E6 which interacts with and induces degradation of numerous cellular proteins including p53 and members of the PDZ-domain family. E6 also recognizes paxillin (PXN), a fundamental component of focal adhesions, interfering with its association to focal adhesion kinase (FAK).


RESULTS
Human umbilical vein endothelial cells, expressing E6 or its mutated variant DeltaE6(105-110) (DeltaE6) which does not inactivate p53, were cultured under static conditions or exposed to a laminar SS of 12 dyn/cm(2) for 16h. In response to SS, cells expressing E6 or DeltaE6 failed to synthesise nitric oxide and directionally remodel their cytoskeleton, as indicated by morphology and phalloidin staining of actin microfilaments. Under these conditions, PXN association with FAK, its localization to the plasma membrane, and its phosphorylation on tyrosine-31, which partially encompasses the PXN/FAK docking site, were severely compromised. These alterations were paralleled by the impairment of important SS-dependent endothelial functions, including nitric oxide production and survival upon serum deprivation. The direct targeting of PXN expression by RNA interference partially reproduced the E6 phenotype, impairing flow-dependent cell orientation and survival but not nitric oxide production.


CONCLUSIONS
These results provide evidence that papilloma virus E6 protein interferes with the function of the SS-mechanosensor and suggests a potential a role for PXN in this process.
Mattiussi
Stefania
S
Laboratorio di Patologia Vascolare, Istituto Dermopatico dell' Immacolata, Roma, Italy.
Matsumoto
Kazue
K
Illi
Barbara
B
Martelli
Fabio
F
Capogrossi
Maurizio C
MC
Gaetano
Carlo
C
D015153:Blotting, Western / Q000379:methods; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism* / Q000473:pathology / Q000821:virology; D004730:Endothelium, Vascular / Q000378:metabolism* / Q000473:pathology / Q000821:virology; D005455:Fluorescent Antibody Technique; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000494:pharmacology*; D030361:Papillomavirus Infections / Q000378:metabolism*; D051419:Paxillin / Q000502:physiology*; D010766:Phosphorylation; D034622:RNA Interference; D013314:Stress, Mechanical; D014161:Transduction, Genetic; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
C118860:E6 protein, Human papillomavirus type 6; D009856:Oncogene Proteins, Viral; D051419:Paxillin; D016159:Tumor Suppressor Protein p53; D009569:Nitric Oxide; C496314:NOS3 protein, human; D052250:Nitric Oxide Synthase Type III; D051416:Focal Adhesion Protein-Tyrosine Kinases