RESUMEN
Wnt signaling dysregulation promotes tumorigenesis in colorectal cancer (CRC). We investigated the role of PTPRF, a receptor-type tyrosine phosphatase, in regulating Wnt signaling in CRC. Knockdown of PTPRF decreased cell proliferation in patient-derived primary colon cancer cells and established CRC cell lines. In addition, the rate of proliferation as well as colony formation ability were significantly decreased in tumor organoids grown in 3D, whereas the number of differentiated tumor organoids were markedly increased. Consistently, knockdown of PTPRF resulted in a decrease in the expression of genes associated with cancer stem cells downstream of Wnt/ß-catenin signaling. Treating PTPRF knockdown cells with GSK3 inhibitor rescued the expression of Wnt target genes suggesting that PTPRF functions upstream of the ß-catenin destruction complex. PTPRF was found to interact with LRP6 and silencing PTPRF largely decreased the activation of LRP6. Interestingly, this PTPRF-mediated activation of Wnt signaling was blocked in cells treated with clathrin endocytosis inhibitor. Furthermore, knockdown of PTPRF inhibited xenograft tumor growth in vivo and decreased the expression of Wnt target genes. Taken together, our studies identify a novel role of PTPRF as an oncogenic protein phosphatase in supporting the activation of Wnt signaling in CRC.
Asunto(s)
Carcinogénesis/patología , Neoplasias Colorrectales/patología , Proteínas Oncogénicas/metabolismo , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/metabolismo , Vía de Señalización Wnt , Animales , Línea Celular Tumoral , Proliferación Celular/genética , Neoplasias Colorrectales/genética , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Ratones , Células Madre Neoplásicas/patología , Proteínas Oncogénicas/genética , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Messenger RNA polyadenylation is a universal aspect of gene expression in eukaryotes. In well-established model organisms, this process is mediated by a conserved complex of 15-20 subunits. To better understand this process in apicomplexans, a group of unicellular parasites that causes serious disease in humans and livestock, a computational and high throughput sequencing study of the polyadenylation complex and poly(A) sites in several species was conducted. BLAST-based searches for orthologs of the human polyadenylation complex yielded clear matches to only two-poly(A) polymerase and CPSF73-of the 19 proteins used as queries in this analysis. As the human subunits that recognize the AAUAAA polyadenylation signal (PAS) were not immediately obvious, a computational analysis of sequences adjacent to experimentally-determined apicomplexan poly(A) sites was conducted. The results of this study showed that there exists in apicomplexans an A-rich region that corresponds in position to the AAUAAA PAS. The set of experimentally-determined sites in one species, Sarcocystis neurona, was further analyzed to evaluate the extent and significance of alternative poly(A) site choice in this organism. The results showed that almost 80% of S. neurona genes possess more than one poly(A) site, and that more than 780 sites showed differential usage in the two developmental stages-extracellular merozoites and intracellular schizonts-studied. These sites affected more than 450 genes, and included a disproportionate number of genes that encode membrane transporters and ribosomal proteins. Taken together, these results reveal that apicomplexan species seem to possess a poly(A) signal analogous to AAUAAA even though genes that may encode obvious counterparts of the AAUAAA-recognizing proteins are absent in these organisms. They also indicate that, as is the case in other eukaryotes, alternative polyadenylation is a widespread phenomenon in S. neurona that has the potential to impact growth and development.
Asunto(s)
Apicomplexa/metabolismo , Neospora/metabolismo , ARN Mensajero/metabolismo , Sarcocystis/metabolismo , Toxoplasma/metabolismo , Apicomplexa/genética , Línea Celular , Biología Computacional , Estudio de Asociación del Genoma Completo , Humanos , Neospora/genética , Poliadenilación , Sarcocystis/genética , Toxoplasma/genética , Secuenciación Completa del GenomaRESUMEN
Erbin belongs to the LAP (leucine-rich repeat and PDZ domain) family of scaffolding proteins that plays important roles in orchestrating cell signaling. Here, we show that Erbin functions as a tumor suppressor in colorectal cancer. Analysis of Erbin expression in colorectal cancer patient specimens revealed that Erbin was downregulated at both mRNA and protein levels in tumor tissues. Knockdown of Erbin disrupted epithelial cell polarity and increased cell proliferation in 3D culture. In addition, silencing Erbin resulted in increased amplitude and duration of signaling through Akt and RAS/RAF pathways. Erbin loss induced epithelial-mesenchymal transition, which coincided with a significant increase in cell migration and invasion. Erbin interacted with kinase suppressor of Ras 1 (KSR1) and displaced it from the RAF/MEK/ERK complex to prevent signal propagation. Furthermore, genetic deletion of Erbin in Apc knockout mice promoted tumorigenesis and significantly reduced survival. Tumor organoids derived from Erbin/Apc double knockout mice displayed increased tumor initiation potential and activation of Wnt signaling. Results from gene set enrichment analysis revealed that Erbin expression associated positively with the E-cadherin adherens junction pathway and negatively with Wnt signaling in human colorectal cancer. Taken together, our study identifies Erbin as a negative regulator of tumor initiation and progression by suppressing Akt and RAS/RAF signaling in vivoSignificance: These findings establish the scaffold protein Erbin as a negative regulator of EMT and tumorigenesis in colorectal cancer through direct suppression of Akt and RAS/RAF signaling. Cancer Res; 78(17); 4839-52. ©2018 AACR.
