Dystrophin Dp71f associates with the beta1-integrin adhesion complex to modulate PC12 cell adhesion.
Dystrophin Dp71 is the main product of the Duchenne muscular dystrophy gene in the brain; however, its function is unknown. To study the role of Dp71 in neuronal cells, we previously generated by antisense treatment PC12 neuronal cell clones with decreased Dp71 expression (antisense-Dp71 cells). PC12 cells express two different splicing isoforms of Dp71, a cytoplasmic variant called Dp71f and a nuclear isoform called Dp71d. We previously reported that antisense-Dp71 cells display deficient adhesion to substrate and reduced immunostaining of beta1-integrin in the cell area contacting the substrate. In this study, we isolated additional antisense-Dp71 clones to analyze in detail the potential involvement of Dp71f isoform with the beta1-integrin adhesion system of PC12 cells. Immunofluorescence analyses as well as immunoprecipitation assays demonstrated that the PC12 cell beta1-integrin adhesion complex is composed of beta1-integrin, talin, paxillin, alpha-actinin, FAK and actin. In addition, our results showed that Dp71f associates with most of the beta1-integrin complex components (beta1-integrin, FAK, alpha-actinin, talin and actin). In the antisense-Dp71 cells, the deficiency of Dp71 provokes a significant reduction of the beta1-integrin adhesion complex and, consequently, the deficient adhesion of these cells to laminin. In vitro binding experiments confirmed the interaction of Dp71f with FAK and beta1-integrin. Our data indicate that Dp71f is a structural component of the beta1-integrin adhesion complex of PC12 cells that modulates PC12 cell adhesion by conferring proper complex assembly and/or maintenance.
Cerna
Joel
J
Departmento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del I.P.N., Av. Instituto Politécnico Nacional 2508, Col. San Pedro Zacatenco, México, D. F. 07360, México.
Cerecedo
Doris
D
Ortega
Arturo
A
García-Sierra
Francisco
F
Centeno
Federico
F
Garrido
Efrain
E
Mornet
Dominique
D
Cisneros
Bulmaro
B
D000185:Actinin / Q000378:metabolism; D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D016189:Dystrophin / Q000235:genetics / Q000378:metabolism*; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D019012:Integrin beta1 / Q000378:metabolism; D007797:Laminin / Q000378:metabolism; D008954:Models, Biological; D009474:Neurons / Q000502:physiology*; D016716:PC12 Cells; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D016608:Talin / Q000378:metabolism
D016189:Dystrophin; D019012:Integrin beta1; D007797:Laminin; D020033:Protein Isoforms; D011994:Recombinant Proteins; D016608:Talin; D000185:Actinin; D005982:Glutathione Transferase; D051416:Focal Adhesion Protein-Tyrosine Kinases