rac regulates its effector phospholipase Cgamma2 through interaction with a split pleckstrin homology domain.
Several isoforms of phospholipase C (PLC) are regulated through interactions with Ras superfamily GTPases, including Rac proteins. Interestingly, of two closely related PLCgamma isoforms, only PLCgamma(2) has previously been shown to be activated by Rac. Here, we explore the molecular basis of this interaction as well as the structural properties of PLCgamma(2) required for activation. Based on reconstitution experiments with isolated PLCgamma variants and Rac2, we show that an unusual pleckstrin homology (PH) domain, designated as the split PH domain (spPH), is both necessary and sufficient to effect activation of PLCgamma(2) by Rac2. We also demonstrate that Rac2 directly binds to PLCgamma(2) as well as to the isolated spPH of this isoform. Furthermore, through the use of NMR spectroscopy and mutational analysis, we determine the structure of spPH, define the structural features of spPH required for Rac interaction, and identify critical amino acid residues at the interaction interface. We further discuss parallels and differences between PLCgamma(1) and PLCgamma(2) and the implications of our findings for their respective signaling roles.
Walliser
Claudia
C
Institute of Pharmacology and Toxicology, University of Ulm Medical Center, 89070 Ulm, Germany.
Retlich
Michael
M
Harris
Richard
R
Everett
Katy L
KL
Josephs
Michelle B
MB
Vatter
Petra
P
Esposito
Diego
D
Driscoll
Paul C
PC
Katan
Matilda
M
Gierschik
Peter
P
Bunney
Tom D
TD
D000818:Animals; D001798:Blood Proteins / Q000737:chemistry*; D019556:COS Cells; D002522:Chlorocebus aethiops; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D008954:Models, Biological; D008958:Models, Molecular; D008968:Molecular Conformation; D051966:Phospholipase C gamma / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry*; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D015398:Signal Transduction
D001798:Blood Proteins; D010750:Phosphoproteins; D020033:Protein Isoforms; C039077:platelet protein P47; D051057:Proto-Oncogene Proteins c-akt; D051966:Phospholipase C gamma