Ras GTPase-activating-like protein IQGAP1 (IQGAP1) also known as p195 is a ubiquitously expressed protein that in humans is encoded by the IQGAP1 gene. [5] [6] [7] IQGAP1 is a scaffold protein involved in regulating various cellular processes ranging from organization of the actin cytoskeleton, transcription, and cellular adhesion to regulating the cell cycle.
IQGAP1 was discovered in 1994. [5] Its name stems from the fact that its RasGAP-related domain (GRD) has sequence homology to the Sar1 GTPase. [8] It was hypothesized that IQGAP1 would act as a GTPase activating protein (GAP) protein, promoting the switch of ras GTPases from the active GTP to GDP-bound forms. However, despite the homology of IQGAP’s GAP domain to sar1 and the fact that IQGAP1 binds Rho GTPases Rac1 and Cdc42, IQGAP does not actually have GAP function. Instead, it binds the active (GTP-bound) forms of RAC1 and CDC42 with higher affinity than GDP-bound forms, and stabilizes the active form in vivo. [9]
IQGAP1 is now recognized as a protein scaffold that integrates signals regulating cell adhesion, actin cytoskeleton, the cell cycle, [9] and other cellular functions. IQGAP is particularly interesting as a therapeutic target since it acts as a node for so many signaling pathways implicated in cancer progression.
Analysis of IQGAP1 expression in human tissues has indicated that the scaffold is more or less ubiquitously expressed. [10] It is usually found in the nucleus, plasma membrane, and cytoplasm. In other words it is found throughout the cell as well as throughout tissue types. Expression analysis has also indicated that IQGAP1 is overexpressed in many cancers, and in more aggressive colorectal and ovarian cancers, IQGAP1 is localized at the invasive front of the neoplasm, indicating a role in mobilization of the cells. [8] Importantly, approximately 10% of genes that show increased expression in metastatic cells are IQGAP1 binding partners. [8]
IQGAP1 is a 190 kDa protein with 5 domains. [9] A protein domain is a subsection of a protein that shows up multiple times in biology and can exist independently of the surrounding protein. It is very similar to subsections of other proteins, and could be cut out of the current protein, exist and function by itself, or be pasted in to a new protein strand and still function properly. Since this area of the protein is conserved in amino acid sequence and structure, it can be characterized by function or binding partner. IQGAP1 has 5 well-known domains separated by other amino acids.
Starting at the N-terminus (or front of the protein), IQGAP1 contains a calponin homology domain (CHD), which mediates actin-binding [11] and binds calponin.
The WW, or poly-proline protein-protein domain, so named because of two functionally conserved tryptophans, W, is a protein-protein interaction domain that associates with proline-rich regions of other proteins. [12] [13]
The WW domain is followed by 4 IQ motifs which form an IQ domain. This domain binds calmodulin, [14] a protein known as a calcium sensor that can bind and regulate many target proteins. [15]
A GRD (rasGAP-related domain) follows the IQ domain. This domain is highly similar to the functional subunit of Ras GTPase-activating proteins (GAPs) and was thus thought to have GAP function. IQGAP1 does bind Rho GTPases CDC42 and RAC1, however, IQGAP1 is unusual in that it actually has no GAP function, and instead stabilizes the GTP-bound proteins in their active state. [16]
Finally, IQGAP1 has a RasGAP_c carboxy terminal sequence important for binding Beta-catenin and E-cadherin. [9]
Homologues of IQGAP1 are known in species as divergent as yeast, worms, and humans (as well as other mammals), though the domains are not always highly conserved. [9]
IQGAP1 is the most well studied member of the IQGAP family of scaffold proteins. The two other members of the family include IQGAP2 and IQGAP3 which have far more restricted expression patterns in comparison with IQGAP1. IQGAP2 is found in the liver, stomach, and platelets and is 62% identical to IQGAP1, [9] but appears to have a drastically divergent function in terms of pathology. [17]
In the brain, IQGAP3 appears to play an important role in neuronal morphogenesis. [18]
This gene encodes a member of the IQGAP family. The protein contains four IQ domains, one calponin homology domain, one Ras-GAP domain and one WW domain. It interacts with components of the cytoskeleton [19] such as the formin Dia1 (mDia1), [20] with cell adhesion molecules (CAMs), and with several signaling molecules to regulate cell morphology and motility. For example, IQGAP1 expression is necessary for neuronal process outgrowth on the cell adhesion molecule PTPmu (PTPRM). [21] Expression of the protein is upregulated by gene amplification in two gastric cancer cell lines [7] and its over-expression and distinct membrane localisation is also observed in a range of tumours. [22]
IQGAP1 is a node intersected by many signaling pathways. As such it has many binding partners, many of which have essential roles in control of the cell cycle and actin cytoskeleton.
IQGAP1 has been shown to interact with:
Protein binding does not by itself construct an interesting story. Far more important is the outcome of the binding event. Does binding change the target protein’s localization? Does it activate the target, or in some way change the target (or effector molecule’s) conformation? As a scaffolding protein, IQGAP1 binds and regulates many targets—its role is to integrate and mediate signaling from diverse pathways and insulate key pathway members from crosstalk.
Scaffolds organize signaling pathways—help regulate how various extracellular signals can be transduced by the same canonical pathway members into various cellular outputs. [33] Generally, scaffolds regulate output, localization, and selectivity of pathways. [34]
As a scaffold involved in different signaling pathways (actin cytoskeleton, cellular adhesion, cell cycle, transcription), IQGAP1 has a unique ability to potentially couple diverse cellular functions. For example IQGAP1 is associated with actin dynamics through direct binding of actin and indirect regulation via Cdc42/Rac1, but also modulates the MAPK pathway which is associated with cell cycle control. Thus IQGAP1 may couple MAPK signaling (decisions about cell fate) to the cytoskeleton or cellular adhesion (potentially acting out those decisions)—an important implication for cancer.
To simplify, due to its diverse range of binding partners, IQGAP1 may act as a link between logically related but molecularly distinct cellular functions. In the above example, actin cytoskeleton rearrangement is required for proliferation (cytokinesis during mitosis). IQGAP1 helps cells both listen to and act on signals, playing an integral role in connecting the dots between signals for proliferation and the actual cellular response.
The Ras→Raf→MEK→ERK MAPK signaling pathway plays an integral part in the processes of cell proliferation, differentiation, and apoptosis. This pathway is conserved across all eukaryotes.
Various extracellular signals induce the ERK MAPK pathway including EGF, IGF-1, PDGF, and NGF. [33] The various scaffolds of this pathway, including IQGAP1, are responsible for modulating the cellular response to the activity of this pathway. For instance, in a given cell line, activation by one extracellular signal may induce differentiation but not proliferation, while activation of the same ERK MAPK pathway by a different extracellular signal will induce proliferation but not differentiation. [33] IQGAP1 seems to be responsible for the specific output of the pathway upon activation by EGF.
IQGAP1 plays a significant role in the propagation of this MAPK signaling pathway. IQGAP directly binds b-RAF, [35] MEK1/2 and ERK1/2, and is in fact necessary for the phosphorylation (activation) of ERK upon stimulation by EGF. [36] [37]
Actin is a major building block of every eukaryotic cell’s cytoskeleton. Actin dynamics play a major role in cell motility (filaments are built at the leading edge of a moving cell and deconstructed at the receding edge). IQGAP1 binds actin and influences actin dynamics by localizing to the leading edge and recruiting actin polymerization machinery. [8] [9] [19]
IQGAP1 binds and is a target of the Rho GTPases CDC42 and RAC1 which are well known regulators of the actin cytoskeleton. [38] [39] Despite its name, IQGAP1 does not have GAP function, and instead stabilizes active Cdc42. This increase in a local pool of active Cdc42 stimulates actin filament formation and thus filopodia formation. [9]
IQGAP1 can crosslink actin, [40] and in many organisms, IQGAP1 is involved in cytokinesis. [41]
Cadherins are a family of adhesion proteins that localize to the cell surface where they anchor a cell to its neighbors by clasping on to the extracellular portion of the neighbor’s cadherins. Actin binds a-catenin which binds beta-catenin which in turn binds E-cadherin. E-cadherin juts into the extracellular space to grasp the extracellular domains of neighboring E-cadherins. IQGAP1 localizes to cell-cell contacts and binds actin, b-catenin, and E-cadherin, weakening these junctions and thus decreasing cell-cell adhesion. [9] [42] IQGAP weakens cell adhesion by displacing a-catenin from the complex. [43]
Active RAC1 binds IQGAP1 to crosslink actin filaments and prevents IQGAP1 from interacting with beta-catenin, stabilizing cell-cell contacts. [44] When IQGAP1 does not bind Rac1, however, it binds beta-catenin, displacing a-catenin from the cadherin-catenin cellular adhesion complex.
IQGAP1 also affects transcription through the Wnt signaling pathway by its interaction with beta-catenin. [8] Beta-catenin is usually sequestered in a complex and excluded from the nucleus, but upon WNT activation this complex is broken and beta-catenin translocates to the nucleus where it activates transcriptional programs. IQGAP1 binds b-catenin and increases nuclear localization and expression of beta-catenin’s transcriptional targets.
IQGAP1 is associated with cytoskeletal dynamics, transcription, cell adhesion, cell cycle, and morphology, all of which are disrupted in cancer. As a modulatory protein intersecting all of these pathways, IQGAP1 can couple many of them, and is also responsible for their proper propagation. Since cancer is a disease characterized by the perturbation of many of these cellular processes, IQGAP1 is a logical oncogene candidate and therapeutic target.
Expression analysis has implicated IQGAP1 in colorectal, squamous cell, breast, gastric, liver, lung, and ovarian cancers, [45] and in some of these cancers, higher IQGAP1 expression levels indicate a poor prognosis. [46]
In order for a cancer to metastasize, cells must gain migratory abilities and invade other tissues. Through Rac1/CDC42, IQGAP1 regulates cellular adhesion and actin dynamics.
In normal cells IQGAP1 localizes to areas of high actin turnover. This characteristic is echoed in invasive tissues, where IQGAP1 localizes to the leading edge of migrating cells. [8] Over-expression of IQGAP1 was associated with increased migration and invasion in a human breast epithelial cancer cell line (MCF-7 cells). [8] [47] IQGAP1 may also be involved in the deregulation of proliferation and differentiation through its modulation of the ERK MAPK pathway.
IQGAP1 may be necessary for tumorigenesis. IQGAP1 knockdown in MCF-7 cancer cells reduced the malignant phenotype (serum-dependent proliferation and anchorage independent growth). 100% of mice injected with MCF-7 cells overexpressing IQGAP1 developed tumors and these tumors were highly invasive. Control MCF-7 cells formed tumors in 60% of the mice, and MCF-7 cells with stable knockdown of IQGAP1 only formed tumors 20% of the time. [47] The mechanism for how IQGAP1 may modulate tumorigenesis/invasion through its various binding partners is of great interest.
IQGAP1 null mice appear significantly normal, with the only life history abnormality being an increase in gastric hyperplasia. [48] Thus, IQGAP1 may be an effective therapeutic target, if its knockdown has little effect in homeostatic tissue but its expression is important in cancer.
Catenins are a family of proteins found in complexes with cadherin cell adhesion molecules of animal cells. The first two catenins that were identified became known as α-catenin and β-catenin. α-Catenin can bind to β-catenin and can also bind filamentous actin (F-actin). β-Catenin binds directly to the cytoplasmic tail of classical cadherins. Additional catenins such as γ-catenin and δ-catenin have been identified. The name "catenin" was originally selected because it was suspected that catenins might link cadherins to the cytoskeleton.
Cortactin is a monomeric protein located in the cytoplasm of cells that can be activated by external stimuli to promote polymerization and rearrangement of the actin cytoskeleton, especially the actin cortex around the cellular periphery. It is present in all cell types. When activated, it will recruit Arp2/3 complex proteins to existing actin microfilaments, facilitating and stabilizing nucleation sites for actin branching. Cortactin is important in promoting lamellipodia formation, invadopodia formation, cell migration, and endocytosis.
α-Catenin (alpha-catenin) functions as the primary protein link between cadherins and the actin cytoskeleton. It has been reported that the actin binding proteins vinculin and α-actinin can bind to alpha-catenin. It has been suggested that alpha-catenin does not bind with high affinity to both actin filaments and the E-cadherin-beta-catenin complex at the same time. It has been observed that when α-catenin is not in a molecular complex with β-catenin, it dimerizes and functions to regulate actin filament assembly, possibly by competing with Arp2/3 protein. α-Catenin exhibits significant protein dynamics. However, a protein complex including a cadherin, actin, β-catenin and α-catenin has not been isolated.
Cell division control protein 42 homolog is a protein that in humans is encoded by the CDC42 gene. Cdc42 is involved in regulation of the cell cycle. It was originally identified in S. cerevisiae (yeast) as a mediator of cell division, and is now known to influence a variety of signaling events and cellular processes in a variety of organisms from yeast to mammals.
Rac1, also known as Ras-related C3 botulinum toxin substrate 1, is a protein found in human cells. It is encoded by the RAC1 gene. This gene can produce a variety of alternatively spliced versions of the Rac1 protein, which appear to carry out different functions.
Ezrin also known as cytovillin or villin-2 is a protein that in humans is encoded by the EZR gene.
LIM domain kinase 1 is an enzyme that in humans is encoded by the LIMK1 gene.
Mitogen-activated protein kinase kinase kinase 11 is an enzyme that in humans is encoded by the MAP3K11 gene.
Cytoplasmic protein NCK1 is a protein that in humans is encoded by the NCK1 gene.
Rho GTPase-activating protein 1 is an enzyme that in humans is encoded by the ARHGAP1 gene.
Alpha-actinin-1 is a protein that in humans is encoded by the ACTN1 gene.
Rac2 is a small signaling G protein, and is a member of the Rac subfamily of the family Rho family of GTPases. It is encoded by the gene RAC2.
Rho guanine nucleotide exchange factor 7 is a protein that in humans is encoded by the ARHGEF7 gene.
Formin-binding protein 1 is a protein that in humans is encoded by the FNBP1 gene.
Receptor-type tyrosine-protein phosphatase mu is an enzyme that in humans is encoded by the PTPRM gene.
Rho GTPase-activating protein 32 is a protein that in humans is encoded by the RICS gene. RICS has two known isoforms, RICS that are expressed primarily at neurite growth cones, and at the post synaptic membranes, and PX-RICS which is more widely expressed in the endoplasmic reticulum, Golgi apparatus and endosomes. The only known domain of the RICS is the RhoGAP domain, whilst PX-RICS has an additional Phox homology and SH3 domain.
Ras GTPase-activating-like protein IQGAP2 is an enzyme that in humans is encoded by the IQGAP2 gene.
mDia1 is a member of the protein family called the formins and is a Rho effector. It is the mouse version of the diaphanous homolog 1 of Drosophila. mDia1 localizes to cells' mitotic spindle and midbody, plays a role in stress fiber and filopodia formation, phagocytosis, activation of serum response factor, formation of adherens junctions, and it can act as a transcription factor. mDia1 accelerates actin nucleation and elongation by interacting with barbed ends of actin filaments. The gene encoding mDia1 is located on Chromosome 18 of Mus musculus and named Diap1.
Long-term potentiation (LTP), thought to be the cellular basis for learning and memory, involves a specific signal transmission process that underlies synaptic plasticity. Among the many mechanisms responsible for the maintenance of synaptic plasticity is the cadherin–catenin complex. By forming complexes with intracellular catenin proteins, neural cadherins (N-cadherins) serve as a link between synaptic activity and synaptic plasticity, and play important roles in the processes of learning and memory.
In molecular biology, the IMD domain is a BAR-like domain of approximately 250 amino acids found at the N-terminus in the insulin receptor tyrosine kinase substrate p53 (IRSp53/BAIAP2) and in the evolutionarily related IRSp53/MIM (MTSS1) family. In IRSp53, a ubiquitous regulator of the actin cytoskeleton, the IMD domain acts as conserved F-actin bundling domain involved in filopodium formation. Filopodium-inducing IMD activity is regulated by Cdc42 and Rac1 and is SH3-independent. The IRSp53/MIM family is a novel F-actin bundling protein family that includes invertebrate relatives:
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