Rab geranylgeranylation occurs preferentially via the pre-formed REP-RGGT complex and is regulated by geranylgeranyl pyrophosphate.
Prenylation (or geranylgeranylation) of Rab GTPases is catalysed by RGGT (Rab geranylgeranyl transferase) and requires REP (Rab escort protein). In the classical pathway, REP associates first with unprenylated Rab, which is then prenylated by RGGT. In the alternative pathway, REP associates first with RGGT; this complex then binds and prenylates Rab proteins. In the present paper we show that REP mutants defective in RGGT binding (REP1 F282L and REP1 F282L/V290F) are unable to compete with wild-type REP in the prenylation reaction in vitro. When over-expressed in cells, REP wild-type and mutants are unable to form stable cytosolic complexes with endogenous unprenylated Rabs. These results suggest that the alternative pathway may predominate in vivo. We also extend previous suggestions that GGPP (geranylgeranyl pyrophosphate) acts as an allosteric regulator of the prenylation reaction. We observed that REP-RGGT complexes are formed in vivo and are unstable in the absence of intracellular GGPP. RGGT increases the ability of REP to extract endogenous prenylated Rabs from membranes in vitro by stabilizing a soluble REP-RGGT-Rab-GG (geranylgeranylated Rab) complex. This effect is regulated by GGPP, which promotes the dissociation of RGGT and REP-Rab-GG to allow delivery of prenylated Rabs to membranes.
Molecular Medicine Section, National Heart and Lung Institute, Imperial College London, London SW7 2AZ, UK.
Rudi A
RA
Baron
Miguel C
MC
Seabra
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D019883:Alkyl and Aryl Transferases / Q000378:metabolism*; D000494:Allosteric Regulation; D000818:Animals; D002478:Cells, Cultured; D006801:Humans; D007700:Kinetics; D009097:Multienzyme Complexes / Q000187:drug effects / Q000378:metabolism; D009154:Mutation; D011106:Polyisoprenyl Phosphates / Q000494:pharmacology*; D054876:Prenylation / Q000187:drug effects*; D011994:Recombinant Proteins / Q000378:metabolism; D018411:Spodoptera; D020826:rab3A GTP-Binding Protein / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; C081787:CHM protein, human; D009097:Multienzyme Complexes; D011106:Polyisoprenyl Phosphates; D011994:Recombinant Proteins; D019883:Alkyl and Aryl Transferases; C075607:Rab geranylgeranyltransferase; D020826:rab3A GTP-Binding Protein; C002963:geranylgeranyl pyrophosphate