New insights into the classification of the RAC1 P29S hotspot mutation in melanoma as an oncogene

The RAC1P29S hotspot mutation, prevalent in melanoma, drives tumorigenesis by enhancing molecular interactions and hyperactivating key signaling pathways, making it a compelling target for cancer therapy. This study provides a comprehensive biochemical characterization of RAC1P29S compared to wild-type RAC1 and mutations T17N and F28L. The P29S mutation significantly impairs nucleotide binding to guanosine triphosphate (GTP) and guanosine diphosphate, accelerating intrinsic nucleotide exchange. While minimally affecting regulation by guanosine dissociation inhibitor 1, RAC1P29S exhibits reduced activation via diffuse B-cell lymphoma family guanine nucleotide exchange factors but retains effective activation by dedicator of cytokinesis 2. Critically, the P29S mutation severely impairs GTPase-activating protein-stimulated GTP hydrolysis, most likely contributing to RAC1P29S hyperactivation by prolonging its GTP-bound form. RAC1P29S displays a stronger binding affinity for IQ motif-containing GTPase-activating protein 1 than for p21-activated kinase 1, highlighting the role of the former in scaffolding RAC1P29S-driven signaling. In serum-starved cells, RAC1P29S predominantly adopts an active GTP-bound state. RAC1P29S overexpression activates key cancer-associated pathways, including extracellular signal-regulated kinase and p38 mitogen-activated protein kinase, reinforcing its role as an oncogenic driver in melanoma. These insights suggest potential therapeutic targets for melanoma treatment, including RAC1 regulators and modulators.