RESUMEN

Human pancreatic ductal adenocarcinoma (PDAC) harboring one KRAS mutant allele often displays increasing genomic loss of the remaining wild-type (WT) allele (known as LOH at KRAS) as tumors progress to metastasis, yet the molecular ramification of this WT allelic loss is unknown. In this study, we showed that the restoration of WT KRAS expression in human PDAC cell lines with LOH at KRAS significantly attenuated the malignancy of PDAC cells both in vitro and in vivo, demonstrating a tumor-suppressive role of the WT KRAS allele. Through RNA-Seq, we identified the HIPPO signaling pathway to be positively regulated by WT KRAS in PDAC cells. In accordance with this observation, PDAC cells with LOH at KRAS exhibited increased nuclear localization and activation of transcriptional co-activator YAP1. Mechanistically, we discovered that WT KRAS expression sequestered YAP1 from the nucleus, through enhanced 14-3-3zeta interaction with phosphorylated YAP1 at S127. Consistently, expression of a constitutively-active YAP1 mutant in PDAC cells bypassed the growth inhibitory effects of WT KRAS. In patient samples, we found that the YAP1-activation genes were significantly upregulated in tumors with LOH at KRAS, and YAP1 nuclear localization predicted poor survival for PDAC patients. Collectively, our results reveal that the WT allelic loss leads to functional activation of YAP1 and enhanced tumor malignancy, which explains the selection advantage of the tumor cells with LOH at KRAS during pancreatic cancer clonal evolution and progression to metastasis, and should be taken into consideration in future therapeutic strategies targeting KRAS.

Asunto(s)

Biomarcadores de Tumor/metabolismo , Carcinoma Ductal Pancreático/patología , Regulación Neoplásica de la Expresión Génica , Pérdida de Heterocigocidad , Neoplasias Pancreáticas/patología , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Señalizadoras YAP/metabolismo , Animales , Apoptosis , Biomarcadores de Tumor/genética , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Proliferación Celular , Femenino , Factores de Transcripción Forkhead/fisiología , Humanos , Ratones , Ratones Desnudos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Pronóstico , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto , Proteínas Señalizadoras YAP/genética

RESUMEN

HHLA2 is a newly identified member of the B7 immune checkpoint family, but its function and crosstalk with immune cells is not fully understood. To gain insights into the HHLA2 expression profile and to determine the clinical significance and function of HHLA2 in pancreatic cancer, we performed immunohistochemistry (IHC) analyses on tissue microarrays (TMAs) of pancreatic ductal adenocarcinoma (PDAC, n = 92) with matched peritumoral tissues as well as in cohorts of precancerous lesions: pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasm (IPMN). We found that HHLA2 was rarely detected in normal acinar, islet, and ductal cells but widely expressed from early pancreatic precancerous lesions to invasive PDAC. The overall HHLA2 positivity was 95% (19/20) in low grade PanIN and 70.73% (29/41) in IPMN. HHLA2 expression was detected in 77.17% (71/92) of the PDAC cases and was significantly associated with better prognosis in this cohort. Our findings suggest that HHLA2 may behave as a costimulatory ligand in pancreatic cancer, which differs from other B7 family members that are largely characterized as checkpoint inhibitors. Further investigation of the HHLA2 signaling pathway and its receptors is warranted by our data and may lead to novel therapeutic interventions.

Asunto(s)

Biomarcadores de Tumor/análisis , Carcinoma in Situ/inmunología , Carcinoma Ductal Pancreático/inmunología , Inmunoglobulinas/análisis , Neoplasias Intraductales Pancreáticas/inmunología , Neoplasias Pancreáticas/inmunología , Carcinoma in Situ/mortalidad , Carcinoma in Situ/patología , Carcinoma in Situ/cirugía , Carcinoma Ductal Pancreático/mortalidad , Carcinoma Ductal Pancreático/patología , Carcinoma Ductal Pancreático/cirugía , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Pancreatectomía , Neoplasias Intraductales Pancreáticas/mortalidad , Neoplasias Intraductales Pancreáticas/patología , Neoplasias Intraductales Pancreáticas/cirugía , Neoplasias Pancreáticas/mortalidad , Neoplasias Pancreáticas/patología , Neoplasias Pancreáticas/cirugía , Estudios Retrospectivos , Factores de Tiempo , Análisis de Matrices Tisulares , Resultado del Tratamiento , Regulación hacia Arriba

RESUMEN

REV1 is one of the major Y-family DNA polymerases. It not only functions as a scaffold protein to mediate other specialized DNA polymerases to sites of lesions, but also inserts deoxycytidine across the lesion strand during translesion DNA synthesis (TLS). Meanwhile, REV1 has been reported to be involved in homologous recombination (HR) repair. Here we further explore the roles of REV1-interacting proteins RAD51 and RAD51C in REV1-mediated DNA double-strand break (DSB) repair. We found that RAD51 but not RAD51C regulates REV1 recruitment to DSB sites via pulsed laser microirradiation. Interestingly, immunofluorescence staining exhibits that REV1 also regulates RAD51 focus formation in response to CPT treatment. These results suggest that REV1 and RAD51 might be mutually dependent on each other in the REV1-related HR pathway.

Asunto(s)

Roturas del ADN de Doble Cadena , Proteínas Nucleares/metabolismo , Nucleotidiltransferasas/metabolismo , Recombinasa Rad51/metabolismo , Línea Celular , Reparación del ADN , Replicación del ADN , Humanos , Proteínas Nucleares/genética , Nucleotidiltransferasas/genética , Unión Proteica , Recombinasa Rad51/genética , Reparación del ADN por Recombinación