Abstract
IQ motif-containing GTPase-activating proteins (IQGAPs) modulate a wide range of cellular processes by acting as scaffolds and driving protein components into distinct signaling networks. Their functional states have been proposed to be controlled by members of the RHO family of GTPases, among other regulators. In this study, we show that IQGAP1 and IQGAP2 can associate with CDC42 and RAC1-like proteins but not with RIF, RHOD, or RHO-like proteins, including RHOA. This seems to be based on the distribution of charged surface residues, which varies significantly among RHO GTPases despite their high sequence homology. Although effector proteins bind first to the highly flexible switch regions of RHO GTPases, additional contacts outside are required for effector activation. Sequence alignment and structural, mutational, and competitive biochemical analyses revealed that RHO GTPases possess paralog-specific residues outside the two highly conserved switch regions that essentially determine the selectivity of RHO GTPase binding to IQGAPs. Amino acid substitution of these specific residues in RHOA to the corresponding residues in RAC1 resulted in RHOA association with IQGAP1. Thus, electrostatics most likely plays a decisive role in these interactions.
Highlights
IQ motif-containing GTPase-activating proteins (IQGAPs) belong to the class of multidomain scaffold proteins that play central roles in the assembly of protein complexes and signaling networks [1,2,3,4,5,6,7]
The following questions were addressed in this study: To what extent do IQGAP paralogs differ in their RHO GTPase-binding characteristics and specificity, and how does IQGAP1 distinguish different RHO GTPases? To this end, we investigated the interactions of IQGAPs with 14 RHO GTPases using the C794 and C795 segments of IQGAP1 and IQGAP2, respectively, which exhibit a sequence identity of 72%
Figure S1G,H), suggesting that IQGAP1 and IQGAP2 do not interact with these unconventional members of the RHO family
Summary
IQ motif-containing GTPase-activating proteins (IQGAPs) belong to the class of multidomain scaffold proteins that play central roles in the assembly of protein complexes and signaling networks [1,2,3,4,5,6,7]. The ubiquitously expressed IQGAP1 is the best-characterized paralog. Multiple domains enable IQGAPs to interact with a large number of proteins and to modulate the spatiotemporal distributions of distinct signal-transducing protein complexes, including B/CRAFMEK1/2-ERK1/2 [9,10,11], FGFR1-CDC42-NWASP-ARP2/3-actin [12,13,14], TIAM1-RAC1PAK6 [15,16], and CDC42/RAC1/CLIP170 [17,18]. IQGAP paralogs share similar domain organization and high sequence homology (Figure 1A). The polyproline-binding region (WW) binds ERK1/2 [9]
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