The phosphorylation process is an important mechanism of cell signalling and regulation. It has been implicated recently in defence strategies against a variety of pathogens that alter host signalling pathways in order to facilitate their invasion and survival within host cells. In this study, the involvement of protein kinases (PKs) has been investigated in attachment and invasion by the pathogenic fungus Fonsecaea pedrosoi within epithelial cells and macrophages. The use of the PK inhibitors staurosporine, genistein and calphostin C prior to infection provided significant information about the role played by PKs in the F. pedrosoi–host cell interaction. All three PK inhibitors could reduce cell invasion by F. pedrosoi significantly. Pre-treatment of macrophages, epithelial cells or conidia with PK inhibitors decreased fungus invasion, and this effect could be overcome by okadaic acid, a phosphatase inhibitor. Immunofluorescence assays showed that tyrosine residues were phosphorylated in the first step of the interaction, while serine residues were phosphorylated in the subsequent step of entry of the parasite into the host cell. These results suggest that both host-cell and conidium PK activities are important in the interaction process, playing a significant role in cell invasion.
AkiyamaT,
IshidaJ,
NakagawaS,
OgawaraH,
WatanabeS,
ItohN,
ShibuyaM,
FukamiY.1987; Genistein, a specific inhibitor of tyrosine-specific protein kinases. J Biol Chem 262:5592–5595
AllenL. A. H,
AderemA.1995; A role for MARCKS, the α isozyme of protein kinase C and myosin I in zymosan phagocytosis by macrophages. J Exp Med 182:829–840[CrossRef]
AllenL. A. H,
AderemA.1996; Molecular definition of distinct cytoskeletal structures involved in complement- and Fc receptor-mediated phagocytosis in macrophages. J Exp Med 184:627–637[CrossRef]
ClercP,
SansonettiP. J.1987; Entry of Shigella flexneri into HeLa cells: evidence for directed phagocytosis involving actin polymerization and myosin accumulation. Infect Immun 55:2681–2688
FarbiarzS. R,
De CarvalhoT. U,
AlvianoC,
De SouzaW.1990; Fine structure and cytochemistry of the interaction between Fonsecaea pedrosoi and mouse resident macrophages. J Med Vet Mycol 28:373–383[CrossRef]
FarbiarzS. R,
De CarvalhoT. U,
AlvianoC,
De SouzaW.1992; Inhibitory effect of melanin on the interaction of Fonsecaea pedrosoi with mammalian cells in vitro. J Med Vet Mycol 30:265–273[CrossRef]
FinlayB. B,
FalkowS.1988; Comparison of the invasion strategies used by Salmonella cholerae-suis , Shigella flexneri and Yersinia enterocolitica to enter cultured animal cells: endosome acidification is not required for bacterial invasion or intracellular replication. Biochimie 70:1089–1099[CrossRef]
FinlayB. B,
FryJ,
RockE. P,
FalkowS.1989; Passage of Salmonella through polarized epithelial cells: role of the host and bacterium. J Cell Sci Suppl 11:99–107
JarvisW. D,
TurnerA. J,
PovirkL. F,
TraylorR. S,
GrantS.1994; Induction of apoptotic DNA fragmentation and cell death in HL-60 human promyelocytic leukemia cells by pharmacological inhibitors of protein kinase C. Cancer Res 54:1707–1714
JouaultT,
FradinC,
TrinelP. A,
BernigaudA,
PoulainD.1998; Early signal transduction induced by Candida albicans in macrophages through shedding of a glycolipid. J Infect Dis 178:792–802[CrossRef]
KáposztaR,
MaródiL,
HollinsheadM,
GordonS,
Da SilvaR. P.1999; Rapid recruitment of late endosomes and lysosomes in mouse macrophages ingesting Candida albicans . J Cell Sci 112:3237–3248
LimongiC. L,
RozentalS,
AlvianoC. S,
De SouzaW.1997; The influence of surface carbohydrates on the interaction of Fonsecaea pedrosoi with Chinese hamster ovary glycosylation mutant cells. Mycopathologia 138:127–135[CrossRef]
LimongiC. L,
AlvianoC. S,
De SouzaW,
RozentalS.2001; Isolation and partial characterization of an adhesin from Fonsecaea pedrosoi . Med Mycol 39:429–437[CrossRef]
OhmoriH,
ToyamaS,
ToyamaS.1992; Direct proof that the primary site of action of cytochalasin on cell motility processes is actin. J Cell Biol 116:933–941
OliveiraL. G,
ResendeM. A,
LopesC. F,
CisalpinoE. O.1973; Isolamento e identificação dos agentes da cromomicose em Belo Horizonte. Rev Soc Bras Med Trop 7: 1 (in Portuguese [CrossRef]
ShahanT. A,
SorensonW. G,
SimpsonJ,
KefalidesN. A,
LewisD. M.2000; Tyrosine kinase activation in response to fungal spores is primarily dependent on endogenous reactive oxygen production in macrophages. J Biol Chem 275:10175–10181[CrossRef]
van der GeerP,
HunterT,
LindbergR. A.1994; Receptor protein-tyrosine kinases and their signal transduction pathways. Annu Rev Cell Biol 10:251–337[CrossRef]
VieiraM. C. F,
De CarvalhoT. U,
De SouzaW.1994; Effect of protein kinase inhibitors on the invasion process of macrophages by Trypanosoma cruzi . Biochem Biophys Res Commun 203:967–971[CrossRef]