Activation of phospholipase D1 by ADP-ribosylated RhoA.
Clostridium botulinum exoenzyme C3 exclusively ADP-ribosylates RhoA, B, and C to inactivate them, resulting in disaggregation of the actin filaments in intact cells. The ADP-ribose resides at Asn-41 in the effector binding region, leading to the notion that ADP-ribosylation inactivates Rho by blocking coupling of Rho to its downstream effectors. In a recombinant system, however, ADP-ribosylated Rho bound to effector proteins such as phospholipase D-1 (PLD1), Rho-kinase (ROK), and rhotekin. The ADP-ribose rather mediated binding of Rho-GDP to PLD1. ADP-ribosylation of Rho-GDP followed by GTP-gamma-S loading resulted in binding but not in PLD activation. On the other hand, ADP-ribosylation of Rho previously activated by binding to GTP-gamma-S resulted in full PLD activation. This finding indicates that ADP-ribosylation seems to prevent GTP-induced change to the active conformation of switch I, the prerequisite of Rho-PLD interaction. In contrast to recombinant systems, ADP-ribosylation in intact cells results in functional inactivation of Rho, indicating other mechanisms of inactivation than blocking effector coupling.
Genth
Harald
H
Institut für Toxikologie, Medizinische Hochschule Hannover, Carl-Neuberg-Strasse 1, Hannover D-30625, Germany. genth.harald@mh-hannover.de
Schmidt
Martina
M
Gerhard
Ralf
R
Aktories
Klaus
K
Just
Ingo
I
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism*; D015153:Blotting, Western; D004789:Enzyme Activation; D006801:Humans; D010739:Phospholipase D / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D000246:Adenosine Diphosphate Ribose; D010739:Phospholipase D; C106801:phospholipase D1; D020742:rhoA GTP-Binding Protein