Artículo
TRPC6 Binds to and activates calpain, independent of its channel activity, and regulates podocyte cytoskeleton, cell adhesion, and motility
Farmer, Louise K.; Rollason, Ruth; Whitcomb, Daniel J.; Ni, Lan; Goodliff, Alexander; Lay, Abigail C.; Birnbaumer, Lutz
; Heesom, Kate J.; Xu, Shang Zhong; Saleem, Moin A.; Welsh, Gavin I.
Fecha de publicación:
10/2019
Editorial:
Synthesis-Stuttgart
Revista:
Journal of the American Society of Nephrology
ISSN:
1046-6673
Idioma:
Inglés
Tipo de recurso:
Artículo publicado
Clasificación temática:
Resumen
Background Mutations in the transient receptor potential channel 6 (TRPC6) gene are associated with an inherited form of FSGS. Despite widespread expression, patients with TRPC6 mutations do not present with any other pathologic phenotype, suggesting that this protein has a unique yet unidentified rolewithin the target cell for FSGS, the kidney podocyte. Methods We generated a stable TRPC6 knockout podocyte cell line from TRPC6 knockout mice. These cells were engineered to express wild-type TRPC6, a dominant negative TRPC6mutation, or either of two disease-causing mutations of TRPC6, G109S or K874*. We extensively characterized these cells using motility, detachment, and calpain activity assays; immunofluorescence; confocal or total internal reflection fluorescence microscopy; and western blotting. Results Compared with wild-type cells, TRPC62/2 podocytes are less motile and more adhesive, with an altered actin cytoskeleton.We found that TRPC6 binds to ERK1/2 and the actin regulatory proteins, caldesmon (a calmodulin- A nd actin-binding protein) and calpain 1 and 2 (calcium-dependent cysteine proteases that control the podocyte cytoskeleton, cell adhesion, andmotility via cleavage of paxillin, focal adhesion kinase, and talin). Knockdown or expression of the truncated K874* mutation (but not expression of the gain-of-function G019S mutation or dominant negative mutant of TRPC6) results in the mislocalization of calpain 1 and 2 and significant downregulation of calpain activity; this leads to altered podocyte cytoskeleton,motility, and adhesion-characteristics of TRPC6 2/2 podocytes. Conclusions Our data demonstrate that independent of TRPC6 channel activity, the physical interaction between TRPC6 and calpain in the podocyte is important for cell motility and detachment and demonstrates a scaffolding role of the TRPC6 protein in disease.
Palabras clave:
focal segmental glomerulosclerosis
,
glomerulus
,
podocyte
,
renal cell biology
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Colecciones
Articulos(BIOMED)
Articulos de INSTITUTO DE INVESTIGACIONES BIOMEDICAS
Articulos de INSTITUTO DE INVESTIGACIONES BIOMEDICAS
Citación
Farmer, Louise K.; Rollason, Ruth; Whitcomb, Daniel J.; Ni, Lan; Goodliff, Alexander; et al.; TRPC6 Binds to and activates calpain, independent of its channel activity, and regulates podocyte cytoskeleton, cell adhesion, and motility; Synthesis-Stuttgart; Journal of the American Society of Nephrology; 30; 10; 10-2019; 1910-1924
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