RESUMEN
Epidermal growth factor (EGF) activates the EGF receptor (EGFR) and stimulates its internalization and trafficking to lysosomes for degradation. However, a percentage of EGFR undergoes ligand-independent endocytosis and is rapidly recycled back to the plasma membrane. Importantly, alterations in EGFR recycling are a common hallmark of cancer, and yet, our understanding of the machineries controlling the fate of endocytosed EGFR is incomplete. Intersectin-s is a multi-domain adaptor protein that is required for internalization of EGFR Here, we discover that intersectin-s binds DENND2B, a guanine nucleotide exchange factor for the exocytic GTPase Rab13, and this interaction promotes recycling of ligand-free EGFR to the cell surface. Intriguingly, upon EGF treatment, DENND2B is phosphorylated by protein kinase D and dissociates from intersectin-s, allowing for receptor targeting to degradation. Our study thus reveals a novel mechanism controlling the fate of internalized EGFR with important implications for cancer.
Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , Receptores ErbB/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Transducción de Señal , Proteínas Supresoras de Tumor/metabolismo , Membrana Celular/metabolismo , Endocitosis , Factor de Crecimiento Epidérmico/farmacología , Receptores ErbB/efectos de los fármacos , Receptores ErbB/genética , Factores de Intercambio de Guanina Nucleótido/genética , Células HEK293 , Humanos , Neoplasias/fisiopatología , Fosforilación , Unión Proteica , Proteína Quinasa C/metabolismo , Transporte de Proteínas , Proteínas Supresoras de Tumor/genética , Proteínas de Unión al GTP rab/metabolismoRESUMEN
During adipocyte differentiation, significant epigenomic changes occur in association with the implementation of the adipogenic program. We have previously shown that histone acetylation increases during differentiation in a manner dependent on acetyl coenzyme A (acetyl-CoA) production by the enzyme ATP-citrate lyase (ACL). Whether ACL regulates nuclear targets in addition to histones during differentiation is not clear. In this study, we report that DNA methyltransferase 1 (DNMT1) levels in adipocytes are controlled in part by ACL and that silencing of DNMT1 can accelerate adipocyte differentiation. DNMT1 gene expression is induced early in 3T3-L1 adipocyte differentiation during mitotic clonal expansion and is critical for maintenance of DNA and histone H3K9 methylation patterns during this period. In the absence of DNMT1, adipocyte-specific gene expression and lipid accumulation occur precociously. Later in differentiation, DNMT1 levels decline in an ACL-dependent manner. ACL-mediated suppression of DNMT1 occurs at least in part by promoting expression of microRNA 148a (miR-148a), which represses DNMT1. Ectopic expression of miR-148a accelerates differentiation under standard conditions and can partially rescue a hypermethylation-mediated differentiation block. The data suggest a role for DNMT1 in modulating the timing of differentiation and describe a novel ACL-miR-148a-dependent mechanism for regulating DNMT1 during adipogenesis.