OBJECTIVE: This study aims to elucidate the functional role of IQGAP1 phosphorylation modification mediated by the SOX4/MAPK1 regulatory axis in developing pancreatic cancer through phosphoproteomics analysis.
METHODS: Proteomics and phosphoproteomics data of pancreatic cancer were obtained from the Clinical Proteomic Tumor Analysis Consortium (CPTAC) database. Differential analysis, kinase-substrate enrichment analysis (KSEA), and independent prognosis analysis were performed on these datasets. Subtype analysis of pancreatic cancer patients was conducted based on the expression of prognostic-related proteins, and the prognosis of different subtypes was evaluated through prognosis analysis. Differential analysis of proteins in different subtypes was performed to identify differential proteins in the high-risk subtype. Clinical correlation analysis was conducted based on the expression of prognostic-related proteins, pancreatic cancer typing results, and clinical characteristics in the pancreatic cancer proteomics dataset. Functional pathway enrichment analysis was performed using GSEA/GO/KEGG, and most module proteins correlated with pancreatic cancer were selected using WGCNA analysis. In cell experiments, pancreatic cancer cells were grouped, and the expression levels of SOX4, MAPK1, and the phosphorylation level of IQGAP1 were detected by RT-qPCR and Western blot experiments. The effect of SOX4 on MAPK1 promoter transcriptional activity was assessed using a dual-luciferase assay, and the enrichment of SOX4 on the MAPK1 promoter was examined using a ChIP assay. The proliferation, migration, and invasion functions of grouped pancreatic cancer cells were assessed using CCK-8, colony formation, and Transwell assays. In animal experiments, the impact of SOX4 on tumor growth and metastasis through the regulation of MAPK1-IQGAP1 phosphorylation modification was studied by constructing subcutaneous and orthotopic pancreatic cancer xenograft models, as well as a liver metastasis model in nude mice.
RESULTS: Phosphoproteomics and proteomics data analysis revealed that the kinase MAPK1 may play an important role in pancreatic cancer progression by promoting IQGAP1 phosphorylation modification. Proteomics analysis classified pancreatic cancer patients into two subtypes, C1 and C2, where the high-risk C2 subtype was associated with poor prognosis, malignant tumor typing, and enriched tumor-related pathways. SOX4 may promote the occurrence of the high-risk C2 subtype of pancreatic cancer by regulating MAPK1-IQGAP1 phosphorylation modification. In vitro cell experiments confirmed that SOX4 promoted IQGAP1 phosphorylation modification by activating MAPK1 transcription while silencing SOX4 inhibited the proliferation, migration, and invasion of pancreatic cancer cells by reducing the phosphorylation level of MAPK1-IQGAP1. In vivo, animal experiments further confirmed that silencing SOX4 suppressed the growth and metastasis of pancreatic cancer by reducing the phosphorylation level of MAPK1-IQGAP1.
CONCLUSIONS: The findings of this study suggest that SOX4 promotes the phosphorylation modification of IQGAP1 by activating MAPK1 transcription, thereby facilitating the growth and metastasis of pancreatic cancer.

The dynamic crosstalk between tumor and stromal cells is a major determinant of cancer aggressiveness. The tumor-suppressor DAB2IP (Disabled homolog 2 interacting protein) plays an important role in this context, since it modulates cell responses to multiple extracellular inputs, including inflammatory cytokines and growth factors. DAB2IP is a RasGAP and negatively controls Ras-dependent mitogenic signals. In addition, it modulates other major oncogenic pathways, including TNFα/NF-κB, WNT/β-catenin, PI3K/AKT, and androgen receptor signaling. In line with its tumor-suppressive role, DAB2IP is frequently inactivated in cancer by transcriptional and post-transcriptional mechanisms, including promoter methylation, microRNA-mediated downregulation, and protein-protein interactions. Intriguingly, some observations suggest that downregulation of DAB2IP in cells of the tumor stroma could foster establishment of a pro-metastatic microenvironment. This review summarizes recent insights into the tumor-suppressive functions of DAB2IP and the consequences of its inactivation in cancer. In particular, we explore potential approaches aimed at reactivating DAB2IP, or augmenting its expression levels, as a novel strategy in cancer treatment. We suggest that reactivation or upregulation of DAB2IP would concurrently attenuate multiple oncogenic pathways in both cancer cells and the tumor microenvironment, with implications for improved treatment of a broad spectrum of tumors.

Macropinocytosis mediates the non-selective bulk uptake of extracellular fluid, enabling cells to survey the environment and obtain nutrients. A conserved set of signaling proteins orchestrates the actin dynamics that lead to membrane ruffling and macropinosome formation across various eukaryotic organisms. At the center of this signaling network are Ras GTPases, whose activation potently stimulates macropinocytosis. However, how Ras signaling is initiated and spatiotemporally regulated during macropinocytosis is not well understood. By using the model system Dictyostelium and a proteomics-based approach to identify regulators of macropinocytosis, we uncovered Leep2, consisting of Leep2A and Leep2B, as a RasGAP complex. The Leep2 complex specifically localizes to emerging macropinocytic cups and nascent macropinosomes, where it modulates macropinosome formation by regulating the activities of three Ras family small GTPases. Deletion or overexpression of the complex, as well as disruption or sustained activation of the target Ras GTPases, impairs macropinocytic activity. Our data reveal the critical role of fine-tuning Ras activity in directing macropinosome formation.

Acute myeloid leukemia (AML) is fatal in the majority of adults. Identification of new therapeutic targets and their pharmacologic modulators are needed to improve outcomes. Previous studies had shown that immunization of rabbits with normal peripheral WBCs that had been incubated with fluorodinitrobenzene elicited high titer antibodies that bound to a spectrum of human leukemias. We report that proteomic analyses of immunoaffinity-purified lysates of primary AML cells showed enrichment of scaffolding protein IQGAP1. Immunohistochemistry and gene-expression analyses confirmed IQGAP1 mRNA overexpression in various cytogenetic subtypes of primary human AML compared to normal hematopoietic cells. shRNA knockdown of IQGAP1 blocked proliferation and clonogenicity of human leukemia cell-lines. To develop small molecules targeting IQGAP1 we performed in-silico screening of 212,966 compounds, selected 4 hits targeting the IQGAP1-GRD domain, and conducted SAR of the \'fittest hit\' to identify UR778Br, a prototypical agent targeting IQGAP1. UR778Br inhibited proliferation, induced apoptosis, resulted in G2/M arrest, and inhibited colony formation by leukemia cell-lines and primary-AML while sparing normal marrow cells. UR778Br exhibited favorable ADME/T profiles and drug-likeness to treat AML. In summary, AML shows response to IQGAP1 inhibition, and UR778Br, identified through in-silico studies, selectively targeted AML cells while sparing normal marrow.

We previously found IQ motif containing GTPase activating protein (IQGAP1) to be consistently elevated in lung fibroblasts (LF) isolated from patients with scleroderma (systemic sclerosis, SSc)-associated interstitial lung disease (ILD) and reported that IQGAP1 contributed to SSc by regulating expression and organization of α-smooth muscle actin (SMA) in LF. The aim of this study was to compare the development of ILD in the presence and absence of IQGAP1. Pulmonary fibrosis was induced in IQGAP1 knockout (KO) and wild-type (WT) mice by a single-intratracheal instillation of bleomycin. Two and three weeks later, mice were euthanized and investigated. We observed that the IQGAP1 KO mouse was characterized by a reduced rate of actin polymerization with reduced accumulation of actin in the lung compared to the WT mouse. After exposure to bleomycin, the IQGAP1 KO mouse demonstrated decreased contractile activity of LF, reduced expression of SMA, TGFβ, and collagen, and lowered overall fibrosis scores compared to the WT mouse. The numbers of inflammatory cells and expression of pro-inflammatory cytokines in lung tissue were not significantly different between IQGAP1 KO and WT mice. We conclude that IQGAP1 plays an important role in the development of lung fibrosis induced by bleomycin, and the absence of IQGAP1 reduces the contractile activity of lung fibroblast and bleomycin-induced pulmonary fibrosis. Thus, IQGAP1 may be a potential target for novel anti-fibrotic therapies for lung fibrosis.

Cell processes require precise regulation of actin polymerization that is mediated by plus-end regulatory proteins. Detailed mechanisms that explain plus-end dynamics involve regulators with opposing roles, including factors that enhance assembly, e.g., the formin mDia1, and others that stop growth (capping protein, CP). We explore IQGAP1\'s roles in regulating actin filament plus-ends and the consequences of perturbing its activity in cells. We confirm that IQGAP1 pauses elongation and interacts with plus ends through two residues (C756 and C781). We directly visualize the dynamic interplay between IQGAP1 and mDia1, revealing that IQGAP1 displaces the formin to influence actin assembly. Using four-color TIRF, we show that IQGAP1\'s displacement activity extends to formin-CP \"decision complexes,\" promoting end-binding protein turnover at plus-ends. Loss of IQGAP1 or its plus-end activities disrupts morphology and migration, emphasizing its essential role. These results reveal a new role for IQGAP1 in promoting protein turnover on filament ends and provide new insights into how plus-end actin assembly is regulated in cells.

Rectal adenocarcinoma (READ) is a common malignant tumor of the digestive tract. Growing studies have confirmed Ras GTPase-activating proteins are involved in the progression of several tumors. This study aimed to explore the expression and function of Ras GTPase-activating proteins in READ. In this study, we analyzed RNA sequencing data from 165 patients with READ and 789 normal tissue samples, identifying 5603 differentially expressed genes (DEGs), including 2937 upregulated genes and 2666 downregulated genes. Moreover, we also identified two dysregulated genes, RASA4 and SYNGAP1, among six Ras GTPase-activating proteins. High NF1 expression was associated with longer overall survival, while high SYNGAP1 expression showed a trend towards extended overall survival. Further analysis revealed the mutation frequency and copy number variations of Ras GTPase-activating proteins in various cancer samples. Additionally, DNA methylation analysis demonstrated a negative correlation between DNA methylation of Ras GTPase-activating proteins and their expression. Moreover, among Ras GTPase-activating proteins, we focused on SYNGAP1, and experimental validation confirmed that the overexpression of SYNGAP1 in READ significantly suppressed READ cell proliferation and increased apoptosis via regulating the Wnt/β-Catenin signaling pathway. These findings underscored the potential significance of SYNGAP1 in READ and provide new insights for further research and treatment.

IQGAP3 (IQ Motif Containing GTPase Activating Protein 3) is member of the IQGAP family of scaffold proteins, which are essential for assembling multiprotein complexes that coordinate various intracellular signaling pathways. Previous research has shown that IQGAP3 is overexpressed in psoriatic skin lesions. Given its involvement in processes like cell proliferation and chemokine signaling, we sought to explore its molecular role in driving the psoriatic phenotype of keratinocytes. By conducting transcriptome profiling of HaCaT keratinocytes, we identified numerous psoriasis-associated pathways that were affected when IQGAP3 was knocked down. These included alterations in NFkB signaling, EGFR signaling, activation of p38/MAPK and ERK1/ERK2, lipid metabolism, cytokine production, and the response to inflammatory cytokine stimulation. Real-time analysis further revealed changes in cell growth dynamics, including proliferation and wound healing. The balance between cell proliferation and apoptosis was altered, as were skin barrier functions and the production of IL-6 and IFNγ. Despite these significant findings, the diversity of the alterations observed in the knockdown cells led us to conclude that IQGAP3 may not be the best target for the therapeutic inhibition to normalize the phenotype of keratinocytes in psoriasis.

Lower-grade gliomas (GBMLGG) are common, fatal, and difficult-to-treat cancers. The current treatment choices have impressive efficacy constraints. As a result, the development of effective treatments and the identification of new therapeutic targets are urgent requirements. Disulfide metabolism is the cause of the non-apoptotic programmed cell death known as disulfideptosis, which was only recently discovered. The mRNA expression data and related clinical information of GBMLGG patients downloaded from public databases were used in this study to investigate the prognostic significance of genes involved in disulfideptosis. In the Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) cohort, our findings showed that many disulfidptosis-related genes were expressed differently in normal and GBMLGG tissues. It was discovered that IQ motif-containing GTPase-activating protein 1 (IQGAP1) is a key gene that influences the outcome of GBMLGG. Besides, a nomogram model was built to foresee the visualization of GBMLGG patients. In addition, in vivo and in vitro validation of IQGAP1\'s cancer-promoting function was done. In conclusion, we discovered a gene signature associated with disulfideptosis that can effectively predict OS in GBMLGG patients. As a result, treating disulfideptosis may be a viable alternative for GBMLGG patients.

Organelles of the endomembrane system contain Rab GTPases as identity markers. Their localization is determined by guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs). It remains largely unclear how these regulators are specifically targeted to organelles and how their activity is regulated. Here, we focus on the GAP Gyp7, which acts on the Rab7-like Ypt7 protein in yeast, and surprisingly observe the protein exclusively in puncta proximal to the vacuole. Mistargeting of Gyp7 to the vacuole strongly affects vacuole morphology, suggesting that endosomal localization is needed for function. In agreement, efficient endolysosomal transport requires Gyp7. In vitro assays reveal that Gyp7 requires a distinct lipid environment for membrane binding and activity. Overexpression of Gyp7 concentrates Ypt7 in late endosomes and results in resistance to rapamycin, an inhibitor of the target of rapamycin complex 1 (TORC1), suggesting that these late endosomes are signaling endosomes. We postulate that Gyp7 is part of regulatory machinery involved in late endosome function.