Leukotriene D(4) induces stress-fibre formation in intestinal epithelial cells via activation of RhoA and PKCdelta.
The intestinal epithelial barrier, which is regulated by the actin cytoskeleton, exhibits permeability changes during inflammation. Here we show that activation of the CysLT(1) receptor by the inflammatory mediator leukotriene D(4) (LTD(4)) causes a rapid increase in stress-fibre formation in intestinal epithelial cells. This effect was mimicked by cytotoxic necrotising factor-1 (CNF-1)-induced activation of RhoA, overexpression of constitutively active RhoA (L63-RhoA) and phorbol-ester-induced activation of protein kinase C (PKC). In accordance, inhibition of RhoA, by C3 exoenzyme or by dominant-negative RhoA (N19-RhoA), as well as GF109203X-induced inhibition of PKC, suppressed the LTD(4)-induced stress-fibre formation. Introduction of the dominant-negative regulatory domain of PKCdelta, but not the corresponding structures from PKCalpha, betaII or epsilon, blocked the LTD(4)-induced stress-fibre formation. Evaluating the relationship between PKCdelta and RhoA in LTD(4)-induced stress-fibre formation, we found that C3 exoenzyme inhibited the rapid LTD(4)-elicited translocation of PKCdelta to the plasma membrane. Furthermore, CNF-1-induced stress-fibre formation was blocked by GF109203X and by overexpression of the regulatory domain of PKC-delta, whereas PKC-induced stress-fibre production was not affected by N19-RhoA. We conclude that PKC-delta is located downstream of RhoA and that active RhoA and PKCdelta are both necessary for LTD(4)-induced stress-fibre formation.
Massoumi
Ramin
R
Experimental Pathology, Department of Laboratory Medicine, Lund University, Malmö University Hospital, SE-205 02 Malmö, Sweden.
Larsson
Christer
C
Sjölander
Anita
A
D036002:ADP Ribose Transferases / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D001427:Bacterial Toxins / Q000378:metabolism; D001905:Botulinum Toxins / Q000378:metabolism; D002458:Cell Fractionation; D002460:Cell Line; D003599:Cytoskeleton / Q000378:metabolism; D003603:Cytotoxins / Q000378:metabolism; D004789:Enzyme Activation; D004847:Epithelial Cells / Q000378:metabolism*; D029968:Escherichia coli Proteins; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology / Q000378:metabolism*; D017998:Leukotriene D4 / Q000378:metabolism*; D008565:Membrane Proteins; D008856:Microscopy, Fluorescence; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D051745:Protein Kinase C-delta; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D018077:Receptors, Leukotriene / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D022502:Stress Fibers / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
D000199:Actins; D001427:Bacterial Toxins; D003603:Cytotoxins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D018077:Receptors, Leukotriene; D011993:Recombinant Fusion Proteins; C060296:cytotoxic necrotizing factor type 1; D017998:Leukotriene D4; C412375:cysteinyl leukotriene receptor 2; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; C497258:PRKCD protein, human; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; D001905:Botulinum Toxins; D020742:rhoA GTP-Binding Protein; C083169:leukotriene D4 receptor; D013755:Tetradecanoylphorbol Acetate