Search

RAS-ERK (extracellular signal–regulated kinase) pathway signals are modulated by scaffold proteins that assemble the components of different kinase tiers into a sequential phosphorylation cascade. In the prevailing model scaffold proteins function as isolated entities, where the flux of phosphorylation events progresses downstream linearly, to achieve ERK phosphorylation. We show that different types of scaffold proteins, specifically KSR1 (kinase suppressor of Ras 1) and IQGAP1 (IQ motif-containing guanosine triphosphatase activating protein 1), can bind to each other, forming a complex whereby phosphorylation reactions occur across both species. MEK (mitogen-activated protein kinase kinase) bound to IQGAP1 can phosphorylate ERK docked at KSR1, a process that we have named “trans-phosphorylation.” We also reveal that ERK trans-phosphorylation participates in KSR1-regulated adipogenesis, and it also underlies the modest cytotoxicity exhibited by KSR-directed inhibitors. Overall, we identify interactions between scaffold proteins and trans-phosphorylation as an additional level of regulation in the ERK cascade, with broad implications in signaling and the design of scaffold protein–aimed therapeutics.

ERK 1/2 proteregulatory proteins like scaffold proteins, that link the components of MAP kinases sequential tiers to form a multienzymatic complex. This way ERK signals are fine-tune (amplitude, intensity and duration), afford signal fidelity and have spatial selectivity. Some examples of scaffold proteins are KSR1, that bind C/BRAF, MEK1/2 and ERK1/2, and IQGAP1, that binds BRAF, MEK1/2 AND ERK1/2. In both cases depletion or blockade of this scaffold functions prevents RAS-ERK pathway-driven neoplasia. We demostrated that ERK can binds constitutively to scaffold, to be cis-phosphorylated therein by scaffold-bound MEK which would enable its binding to a free, phosphorylated ERK monomer to form a dimer. We have found that KSR1 y IQGAP1 can functionally associate with each other. As a consecuence, ERK docked at KSR1 can be phosphorylated by MEK bound to IQGAP1, a process named ¿trans-phosphorylarion¿. The results identify interactions betweeen scaffold proteins as a higher-order level of regulation in the ERK cascade. ins are major actors in the transduction of signals conveyed by external stimuli to the interior of the cell, being regulators of biochemical processes such as proliferation, differentiation or survival. ERK signals are modulated by different

The transcription factor MYC regulates cell proliferation, transformation, and survival in response to growth factor signaling that is mediated in part by the kinase activity of ERK2. Because ERK2 can also bind to DNA to modify gene expression, we investigated whether it more directly regulates MYC transcription. We identified ERK2 binding sites in the MYC promoter and detected ERK2 at the promoter in various serum-stimulated cell types. Expression of nuclear-localized ERK2 constructs in serum-starved cells revealed that ERK2 in the nucleus—regardless of its kinase activity—increased MYC mRNA expression and MYC protein abundance. ERK2 bound to the promoter through its amino-terminal insert domain and to the cyclin-dependent kinase CDK9 (which activates RNA polymerase II) through its carboxyl-terminal conserved docking domain. Both interactions were essential for ERK2-induced MYC expression, and depleting ERK impaired CDK9 occupancy and RNA polymerase II progression at the MYC promoter. Artificially tethering CDK9 to the MYC promoter by fusing it to the ERK2 insert domain was sufficient to stimulate MYC expression in serum-starved cells. Our findings demonstrate a role for ERK2 at the MYC promoter acting as a kinase-independent anchor for the recruitment of CDK9 to promote MYC expression.

Metastatic melanoma is a highly immunogenic tumor with very poor survival rates due to immune system escape-mechanisms. Immune checkpoint inhibitors (ICIs) targeting the cytotoxic T-lymphocyte-associated protein 4 (CTLA4) and the programmed death-1 (PD1) receptors, are being used to impede immune evasion. This immunotherapy entails an increment in the overall survival rates. However, melanoma cells respond with evasive molecular mechanisms. ERK cascade inhibitors are also used in metastatic melanoma treatment, with the RAF activity blockade being the main therapeutic approach for such purpose, and in combination with MEK inhibitors improves many parameters of clinical efficacy. Despite their efficacy in inhibiting ERK signaling, the rewiring of the melanoma cell-signaling results in disease relapse, constituting the reinstatement of ERK activation, which is a common cause of some resistance mechanisms. Recent studies revealed that the combination of RAS–ERK pathway inhibitors and ICI therapy present promising advantages for metastatic melanoma treatment. Here, we present a recompilation of the combined therapies clinically evaluated in patients.

Background: RAS-to-ERK signaling is crucial for the onset and progression of advanced thyroid carcinoma, and blocking ERK dimerization provides a therapeutic benefit in several human carcinomas. Here we analyzed the effects of DEL-22379, a relatively specific ERK dimerization inhibitor, on the activation of the RAS-to-ERK signaling cascade and on tumor-related processes in vitro and in vivo. Methods: We used a panel of four human anaplastic thyroid carcinoma (ATC) cell lines harboring BRAF or RAS mutations to analyze ERK dynamics and tumor-specific characteristics. We also assessed the impact of DEL-22379 on the transcriptional landscape of ATC cell lines using RNA-sequencing and evaluated its therapeutic efficacy in an orthotopic mouse model of ATC. Results: DEL-22379 impaired upstream ERK activation in BRAF- but not RAS-mutant cells. Cell viability and metastasis-related processes were attenuated by DEL-22379 treatment, but mostly in BRAF-mutant cells, whereas in vivo tumor growth and dissemination were strongly reduced for BRAF-mutant cells and mildly reduced for RAS-mutant cells. Transcriptomics analyses indicated that DEL-22379 modulated the transcriptional landscape of BRAF- and RAS-mutant cells in opposite directions. Conclusions: Our findings establish that BRAF- and RAS-mutant thyroid cells respond differentially to DEL-22379, which cannot be explained by the previously described mechanism of action of the inhibitor. Nonetheless, DEL-22379 demonstrated significant anti-tumor effects against BRAF-mutant cells in vivo with an apparent lack of toxicity, making it an interesting candidate for the development of combinatorial treatments. Our data underscore the differences elicited by the specific driver mutation for thyroid cancer onset and progression, which should be considered for experimental and clinical approaches.

Thyroid cancer is mostly an ERK-driven carcinoma, as up to 70% of thyroid carcinomas are caused by mutations that activate the RAS/ERK mitogenic signaling pathway. The incidence of thyroid cancer has been steadily increasing for the last four decades; yet, there is still no effective treatment for advanced thyroid carcinomas. Current research efforts are focused on impairing ERK signaling with small-molecule inhibitors, mainly at the level of BRAF and MEK. However, despite initial promising results in animal models, the clinical success of these inhibitors has been limited by the emergence of tumor resistance and relapse. The RAS/ERK pathway is an extremely complex signaling cascade with multiple points of control, offering many potential therapeutic targets: from the modulatory proteins regulating the activation state of RAS proteins to the scaffolding proteins of the pathway that provide spatial specificity to the signals, and finally, the negative feedbacks and phosphatases responsible for inactivating the pathway. The aim of this review is to give an overview of the biology of RAS/ERK regulators in human cancer highlighting relevant information on thyroid cancer and future areas of research.