Unregulated actin polymerization by WASp causes defects of mitosis and cytokinesis in X-linked neutropenia.
Specific mutations in the human gene encoding the Wiskott-Aldrich syndrome protein (WASp) that compromise normal auto-inhibition of WASp result in unregulated activation of the actin-related protein 2/3 complex and increased actin polymerizing activity. These activating mutations are associated with an X-linked form of neutropenia with an intrinsic failure of myelopoiesis and an increase in the incidence of cytogenetic abnormalities. To study the underlying mechanisms, active mutant WASp(I294T) was expressed by gene transfer. This caused enhanced and delocalized actin polymerization throughout the cell, decreased proliferation, and increased apoptosis. Cells became binucleated, suggesting a failure of cytokinesis, and micronuclei were formed, indicative of genomic instability. Live cell imaging demonstrated a delay in mitosis from prometaphase to anaphase and confirmed that multinucleation was a result of aborted cytokinesis. During mitosis, filamentous actin was abnormally localized around the spindle and chromosomes throughout their alignment and separation, and it accumulated within the cleavage furrow around the spindle midzone. These findings reveal a novel mechanism for inhibition of myelopoiesis through defective mitosis and cytokinesis due to hyperactivation and mislocalization of actin polymerization.
Moulding
Dale A
DA
Wolfson Centre for Gene Therapy of Childhood Disease, UCL Institute of Child Health, University College London, London, UK.
Blundell
Michael P
MP
Spiller
David G
DG
White
Michael R H
MR
Cory
Giles O
GO
Calle
Yolanda
Y
Kempski
Helena
H
Sinclair
Jo
J
Ancliff
Phil J
PJ
Kinnon
Christine
C
Jones
Gareth E
GE
Thrasher
Adrian J
AJ
D000199:Actins / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002869:Chromosome Aberrations; D002877:Chromosomes, Human; D048749:Cytokinesis / Q000187:drug effects; D004247:DNA; D047630:Depsipeptides / Q000494:pharmacology; D040181:Genetic Diseases, X-Linked / Q000378:metabolism* / Q000473:pathology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D008938:Mitosis / Q000187:drug effects; D050505:Mutant Proteins / Q000378:metabolism; D009503:Neutropenia / Q000378:metabolism* / Q000473:pathology*; D011123:Polyploidy; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D019076:Transgenes; D051316:Wiskott-Aldrich Syndrome Protein / Q000378:metabolism*
D000199:Actins; D047630:Depsipeptides; D050505:Mutant Proteins; D011993:Recombinant Fusion Proteins; D051316:Wiskott-Aldrich Syndrome Protein; C485184:enhanced green fluorescent protein; C057531:jasplakinolide; D049452:Green Fluorescent Proteins; D004247:DNA