Diverse MAP kinase pathways respond to different external cues and exert distinct biological functions. However, there seems to be potential for extensive overlap with regard not only to the activating stimuli but also to downstream targets. The mechanism by which the specificity and fidelity of distinct biological responses are maintained is largely unknown. Recent reports indicate that specificity in MAP kinase signaling in Saccharomyces cerevisiae is attained predominantly by scaffolding proteins (38). Similarly, a number of scaffolding proteins that modulate signaling from Raf to MEK and ERK have been identified in mammalian cells (for reviews, see references 23 and 36). These include kinase suppressor of Ras (KSR) (37), connector enhancer of KSR (19), Sur8 (28), MEK partner 1 (MP1) (42), MAP kinase organizer 1 (48), Sef (45), and Raf kinase inhibitor protein (51).IQGAPs are multidomain molecules that contain several protein-interacting motifs (for reviews, see references 4 and 5). The name is derived from the presence of an IQ domain (with four tandem IQ motifs) and a region with significant sequence similarity to the catalytic domain of Ras-GTPase-activating proteins (GAPs). Other motifs include a calponin homology domain (CHD), a coiled-coil region and a WW domain. The multiple conserved modules in IQGAP1 suggest that it participates in protein-protein interactions and may be a component of several signaling pathways. This hypothesis has been validated by both our group and others who documented that IQGAP1 binds activated Cdc42 and Rac1 (but not RhoA or Ras) (11, 16, 20), actin (10, 17, 32), calmodulin (17, 20, 32), E-cadherin (25, 29), -catenin (3, 25), S100B (33), nectin (21), CLIP-170 (12), ERK2 (40), and adenomatous polyposis coli (49). The diversity of IQGAP1 targets has led to the hypothesis that IQGAP1 functions as a scaffolding protein that can assemble multiprotein complexes within various subcellular domains (4, 17). In support of this concept, published evidence reveals that IQGAP1 is capable of binding several proteins simultaneously. For example, complexes of IQGAP1 with actin and Cdc42 have been isolated in vitro (10). Moreover, complexes of IQGAP1 with Cdc42 and calmodulin (17), Rac1 and calmodulin (41), or Rac1/Cdc42 and CLIP-170 (12) have been reported. These findings strongly suggest that IQGAP1 functions as a scaffold within cells.We previously demonstrated that IQGAP1 binds directly to ERK2 and regulates its activity (40). Moreover, altering intracellular IQGAP1 levels inhibited epidermal growth factor (EGF)-stimulated activation of ERK. These findings led to the question of whether IQGAP1 may modulate MEK, the molecule immediately upstream of ERK in the Ras-Raf-MEK-ERK pathway. Here, we present evidence that IQGAP1 and MEK associate both in vitro and in intact cells. Analogous to our observations with ERK, the ability of EGF to stimulate phosphorylation of MEK was abrogated in cells in which endogenous IQGAP1 was specifically knocked down by small interfering RNA (siRNA). Both overe...
IQGAP1 binds several proteins including actin, calmodulin, E-cadherin, -catenin, Cdc42, Rac1, and CLIP-170. The interaction with these targets enables IQGAP1 to participate in many cellular functions varying from regulation of the cytoskeleton to gene transcription. Here we show that extracellular signal-regulated kinase (ERK) 2 binds to IQGAP1. In vitro analysis with purified proteins demonstrated a direct interaction between ERK2 and IQGAP1. Moreover, binding occurred in cells as endogenous ERK2 co-immunoprecipitated with IQGAP1 from human breast epithelial cell lysates. The association between ERK2 and IQGAP1 was independent of epidermal growth factor. The in vivo interaction has functional significance. Manipulation of intracellular IQGAP1 levels significantly reduced growth factorstimulated ERK1 and ERK2 activity. Similarly, stimulation of ERK1 and ERK2 activity by insulin-like growth factor I was reduced when IQGAP1 levels were changed. In contrast, overexpression of an IQGAP1 construct lacking the ERK2 binding region did not interfere with activation of ERK1 and ERK2 by epidermal growth factor. Our data disclose a previously unidentified communication between IQGAP1 and the ERK pathway and imply that IQGAP1 modulates the Ras/mitogen-activated protein kinase signaling cascade.Extracellular signals are converted into cellular responses via a complex network of intersecting signaling pathways (1). A prominent component of intracellular signaling is the family of mitogen-activated protein (MAP) 1 kinases. The archetypal MAP kinase pathway consists of a three-kinase cascade, a MAP kinase kinase kinase (MAPKKK or MEKK) that activates the downstream MAP/ERK kinase (MAPKK or MEK) by phosphorylation, which in turn elicits a phosphorylation-dependent increase in the activity of the MAP kinase (2, 3). The MAP kinase then induces phosphorylation of a variety of cytosolic or nuclear targets, transcription factors, transcriptional adaptor proteins, and other protein kinases. Of the various mammalian MAP kinase pathways, the Ras/Raf/MEK/ERK is the most studied. This cascade, triggered by a diverse range of stimuli acting through cell surface receptors, is under the control of the small G protein Ras (4). When GTP-bound, Ras binds to Raf and recruits it to the cell membrane where Raf is activated (5-7). Raf phosphorylates and activates the dual specificity protein kinases MEK1 and MEK2, which in turn stimulate ERK1 and ERK2 by catalyzing their phosphorylation (3). The Ras/Raf/MEK/ERK cascade is conserved in all eukaryotes, and ERK plays a vital role in several biological processes, particularly those involving cellular proliferation, differentiation, survival, and apoptosis (8).IQGAP1 is a scaffolding protein with multiple protein-interacting domains (for reviews, see Refs. 9 -11). These motifs include a calponin homology domain (CHD), four IQ motifs, and a RasGAP-related domain. Targets for IQGAP1 include calmodulin (12-14), Cdc42 (12, 13), Rac1 (13), actin (14, 15), -catenin (16, 17), E-cadherin (16, 18), S100B (19) ...
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