Cdc42 and the actin-related protein/neural Wiskott-Aldrich syndrome protein network mediate cellular invasion by Cryptosporidium parvum.
Cryptosporidium parvum invasion of epithelial cells involves host cell membrane alterations which require a remodeling of the host cell actin cytoskeleton. In addition, an actin plaque, possibly associated with the dense-band region, forms within the host cytoplasm at the host-parasite interface. Here we show that Cdc42 and RhoA, but not Rac1, members of the Rho family of GTPases, are recruited to the host-parasite interface in an in vitro model of human biliary cryptosporidiosis. Interestingly, activation of Cdc42, but not RhoA, was detected in the infected cells. Neural Wiskott-Aldrich syndrome protein (N-WASP) and p34-Arc, actin-regulating downstream effectors of Cdc42, were also recruited to the host-parasite interface. Whereas cellular expression of a constitutively active mutant of Cdc42 promoted C. parvum invasion, overexpression of a dominant negative mutant of Cdc42, or depletion of Cdc42 mRNA by short interfering RNA-mediated gene silencing, inhibited C. parvum invasion. Expression of the WA fragment of N-WASP to block associated actin polymerization also inhibited C. parvum invasion. Moreover, inhibition of host cell Cdc42 activation by dominant negative mutation inhibited C. parvum-associated actin remodeling, membrane protrusion, and dense-band formation. In contrast, treatment of cells with a Rho inhibitor, exoenzyme C3, or cellular overexpression of dominant negative mutants of RhoA and Rac1 had no effect on C. parvum invasion. These data suggest that C. parvum invasion of target epithelia results from the organism's ability to activate a host cell Cdc42 GTPase signaling pathway to induce host cell actin remodeling at the attachment site.
Chen
Xian-Ming
XM
The Center for Basic Research in Digestive Diseases, Division of Gastroenterology and Hepatology, Mayo Medical School, Clinic and Foundation, Rochester, Minnesota 55905, USA.
Huang
Bing Q
BQ
Splinter
Patrick L
PL
Orth
James D
JD
Billadeau
Daniel D
DD
McNiven
Mark A
MA
LaRusso
Nicholas F
NF
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001660:Biliary Tract Diseases / Q000378:metabolism / Q000469:parasitology / Q000473:pathology; D002460:Cell Line; D003457:Cryptosporidiosis / Q000209:etiology / Q000378:metabolism / Q000469:parasitology; D016785:Cryptosporidium parvum / Q000472:pathogenicity*; D004847:Epithelial Cells / Q000378:metabolism / Q000469:parasitology; D006790:Host-Parasite Interactions; D006801:Humans; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D014923:Wiskott-Aldrich Syndrome / Q000235:genetics / Q000378:metabolism; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D020741:rho GTP-Binding Proteins / Q000378:metabolism
D000199:Actins; D009419:Nerve Tissue Proteins; C495771:WASL protein, human; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D020764:cdc42 GTP-Binding Protein; D020741:rho GTP-Binding Proteins