Rho is a negative regulator of human monocyte spreading.
Monocyte spreading is a sequential series of events including cell flattening, formation of new actin structures and focal adhesions, as well as development of cytoplasmic projections. To investigate the involvement of the GTP-binding protein Rho in spreading, we treated human blood monocytes or PMA-stimulated U937 or THP1 cells with Clostridium botulinum C3-transferase (C3), which ADP-ribosylates and inactivates Rho in intact cells. The C3 treatment caused 1) a four- to fivefold increase in the number of THP1 cells that spread on fibronectin within 24 h of PMA stimulation, 2) a greater area covered by the spread cells, and 3) accelerated and enhanced development of macrophage-like filopodial and pseudopodial projections. Similar results were obtained with PMA-stimulated U937 cells and human blood monocytes. Furthermore, cell staining revealed disorganization of subcortical actin in C3-treated THP1 cells, whereas circular actin formations at the substrate-attached part of the cells and vinculin-containing focal complexes/adhesions were unaffected. Finally, we found a decrease in membrane-associated RhoA in normal spreading THP1 cells, which suggests endogenous inactivation of Rho and might provide an explanation for the acceleration of spreading caused by the C3-transferase. In conclusion, these results indicate that active Rho is an important, negative regulator of human monocyte spreading by maintaining cell tension and cortical actin organization.
Aepfelbacher
M
M
Institute for Prevention of Cardiovascular Diseases, University of Munich, Germany.
Essler
M
M
Huber
E
E
Czech
A
A
Weber
P C
PC
D036002:ADP Ribose Transferases / Q000494:pharmacology; D000199:Actins / Q000378:metabolism; D001905:Botulinum Toxins; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D003014:Clostridium botulinum / Q000201:enzymology; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D006801:Humans; D009000:Monocytes / Q000187:drug effects / Q000502:physiology*; D016596:Vinculin / Q000378:metabolism; D020742:rhoA GTP-Binding Protein
D000199:Actins; D016596:Vinculin; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D001905:Botulinum Toxins; D019204:GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein