A partial down-regulation of WASP is sufficient to inhibit podosome formation in dendritic cells.
The Wiskott Aldrich syndrome protein (WASP) is a hematopoietic-specific cytoskeletal regulator that is necessary for induction of normal immunity. In the context of effective gene therapy for WAS, cellular models of human WASP deficiency are important for definition of the threshold of protein expression required for optimal activity. Using lentiviral vector-mediated RNA interference (RNAi), we were able to down-regulate the levels of human WASP in cell lines and primary cells. In dendritic cells (DC), RNAi-induced WASP deficiency did not impair phenotypic maturation but perturbed cytoskeletal organization. As a result, podosomes, which are actin-rich structures present in immature adherent DC, were formed less efficiently and motility was disturbed. Overall, treatment of cells with RNAi recapitulated the phenotype of cells derived from patients or animals with inactivating mutations of the WAS gene. Interestingly, reduction of the levels of WASP to about 60% of normal was sufficient to inhibit the formation of podosomes in DC, implying that this cell type requires near-normal levels of WASP to sustain physiological cytoskeleton-dependent activities.
Olivier
Aurelie
A
Unité mixte INSERM-Université Paris XI, U362, Institut Gustave Roussy, Villejuif F-94805 and CNRS, UMR 8115, Généthon, 91002 Evry, France.
Jeanson-Leh
Laurence
L
Bouma
Gerben
G
Compagno
Daniel
D
Blondeau
Johanna
J
Seye
Khalil
K
Charrier
Sabine
S
Burns
Siobhan
S
Thrasher
Adrian J
AJ
Danos
Olivier
O
Vainchenker
William
W
Galy
Anne
A
D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D003713:Dendritic Cells / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000627:therapeutic use; D006801:Humans; D019169:Jurkat Cells; D016086:Lentivirus / Q000235:genetics; D009154:Mutation; D034741:RNA, Small Interfering / Q000378:metabolism / Q000494:pharmacology*; D014161:Transduction, Genetic; D014923:Wiskott-Aldrich Syndrome / Q000235:genetics; D051316:Wiskott-Aldrich Syndrome Protein / Q000172:deficiency / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D034741:RNA, Small Interfering; D051316:Wiskott-Aldrich Syndrome Protein