RESUMEN
In this study, we describe a new post-translational modification at position -1 of the PDZ-binding motif in the mutated in colorectal cancer (MCC) protein and its role in lamellipodia formation. Serine 828 at position -1 of this motif is phosphorylated, which is predicted to increase MCC binding affinity with the polarity protein Scrib. We show that endogenous MCC localizes at the active migratory edge of cells, where it interacts with Scrib and the non-muscle motor protein Myosin-IIB. Expression of MCC harboring a phosphomimetic mutation MCC-S828D strongly impaired lamellipodia formation and resulted in accumulation of Myosin-IIB in the membrane cortex fraction. We propose that MCC regulates lamellipodia formation by binding to Scrib and its downstream partner Myosin-IIB in a multiprotein complex. Importantly, we propose that the function of this complex is under the regulation of a newly described phosphorylation of the PDZ-binding motif at position -1.
Asunto(s)
Colon/citología , Células Epiteliales/metabolismo , Dominios PDZ , Fosfoserina/metabolismo , Seudópodos/metabolismo , Proteínas Supresoras de Tumor/química , Proteínas Supresoras de Tumor/metabolismo , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Sustitución de Aminoácidos/genética , Línea Celular Tumoral , Membrana Celular/metabolismo , Polaridad Celular , Humanos , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Datos de Secuencia Molecular , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Mutación/genética , Miosina Tipo IIB no Muscular/metabolismo , Fosforilación , Unión Proteica , Transporte de Proteínas , Relación Estructura-Actividad , Fracciones Subcelulares/metabolismoRESUMEN
Mutations in the RUNX1 gene have been associated with chemotherapy resistance and poor prognosis in patients with acute myeloid leukemia (AML), T-cell acute lymphoblastic leukemia, and myelodysplastic syndromes. However, the underlying mechanisms connecting RUNX1 to the success of therapy remain elusive. Here we explore the hypothesis that RUNX1 is directly involved in the response of hematopoietic cells to cytotoxic agents. RUNX1 was upregulated posttranscriptionally by cytotoxic agents in C57BL/6 mice in vivo and hematopoietic cell lines. Upregulation was also seen in primary human AML cells after treatment with cytarabine in vitro Upon overexpression, RUNX1 restricted proliferation, promoted apoptosis, and augmented the DNA damage response. This unknown activity of RUNX1 required an intact runt homology domain (RHD), a domain where most leukemia-associated point mutations cluster. Consistent with this, two RHD-defective RUNX1 proteins lacked any antiproliferative or apoptotic activity, and RHD-defective (K83N, N109D) mutant RUNX1 conferred resistance to ionizing radiation when overexpressed in Ba/F3 cells under certain conditions. Our experiments reveal a novel function of RUNX1 and offer an explanation for the link between RUNX1 mutations and chemotherapy and radiation resistance. Moreover, these data suggest that pharmacologic modulation of RUNX1 might be an attractive new approach to treat hematologic malignancies. Cancer Res; 77(24); 6818-24. ©2017 AACR.