RESUMO
Aberrant activation of endoplasmic reticulum (ER) stress by extrinsic and intrinsic factors contributes to tumorigenesis and resistance to chemotherapies in various cancer types. Our previous studies have shown that the downregulation of PHLPP, a novel family of Ser/Thr protein phosphatases, promotes tumor initiation, and progression. Here we investigated the functional interaction between the ER stress and PHLPP expression in colon cancer. We found that induction of ER stress significantly decreased the expression of PHLPP proteins through a proteasome-dependent mechanism. Knockdown of PHLPP increased the phosphorylation of eIF2α as well as the expression of autophagy-associated genes downstream of the eIF2α/ATF4 signaling pathway. In addition, results from immunoprecipitation experiments showed that PHLPP interacted with eIF2α and this interaction was enhanced by ER stress. Functionally, knockdown of PHLPP improved cell survival under ER stress conditions, whereas overexpression of a degradation-resistant mutant PHLPP1 had the opposite effect. Taken together, our studies identified ER stress as a novel mechanism that triggers PHLPP downregulation; and PHLPP-loss promotes chemoresistance by upregulating the eIF2α/ATF4 signaling axis in colon cancer cells.
Assuntos
Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Regulação para Baixo/genética , Resistencia a Medicamentos Antineoplásicos , Estresse do Retículo Endoplasmático , Fator de Iniciação 2 em Eucariotos/metabolismo , Proteínas Nucleares/genética , Fosfoproteínas Fosfatases/genética , Fator 4 Ativador da Transcrição/metabolismo , Autofagia/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Estresse do Retículo Endoplasmático/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Irinotecano/farmacologia , Irinotecano/uso terapêutico , Proteínas Nucleares/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Fosforilação , Complexo de Endopeptidases do Proteassoma/metabolismo , Inibidores de Proteassoma/farmacologia , Ligação Proteica/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Tunicamicina/farmacologia , Tunicamicina/uso terapêuticoRESUMO
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.
Assuntos
Carcinogênese/patologia , Neoplasias Colorretais/patologia , Proteínas Oncogênicas/metabolismo , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/metabolismo , Via de Sinalização Wnt , Animais , Linhagem Celular Tumoral , Proliferação de Células/genética , Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Células-Tronco Neoplásicas/patologia , Proteínas Oncogênicas/genética , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Cancer cells are known to upregulate aerobic glycolysis to promote growth, proliferation, and survival. However, the role of mitochondrial respiration in tumorigenesis remains elusive. Here we report that inhibition of mitochondrial function by silencing TFAM, a key transcription factor essential for mitochondrial DNA (mtDNA) replication and the transcription of mtDNA-encoded genes, markedly reduced tumor-initiating potential of colon cancer cells. Knockdown of TFAM significantly decreased mitochondrial respiration in colon cancer cells; however, the cellular levels of ATP remained largely unchanged as a result of increased glycolysis. This metabolic alteration rendered cancer cells highly susceptible to glucose deprivation. Interestingly, upregulation of glycolysis was independent of hypoxia-inducible factor-1 (HIF1) as TFAM knockdown cells fail to stabilize HIF1α under hypoxic conditions. Moreover, knockdown of TFAM results in decreased expression of genes-associated cancer stem cells downstream of Wnt/ß-catenin signaling. Metabolic analysis reveals that the level of α-ketoglutarate (α-KG) was significantly upregulated in TFAM knockout cells. Silencing of prolyl hydroxylase domain-containing protein 2 (PHD2), a α-KG-dependent dioxyenase, rescued the expression of target genes of both HIF1α and Wnt/ß-catenin. Furthermore, intestinal-specific knockout of TFAM prevents tumor formation in Apc-mutant mouse models of colon cancer. Taken together, our findings identify a novel role of mitochondria-mediated retrograde signaling in regulating Wnt signaling and tumor initiation in colon cancer.
Assuntos
Neoplasias do Colo/genética , Mitocôndrias/metabolismo , Via de Sinalização Wnt/genética , Animais , Carcinogênese , Humanos , Camundongos , Transdução de SinaisRESUMO
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.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Carcinogênese/genética , Proliferação de Células/genética , Neoplasias Colorretais/genética , Proteínas Quinases/genética , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteína da Polipose Adenomatosa do Colo/genética , Animais , Caderinas/genética , Movimento Celular/genética , Polaridade Celular/genética , Neoplasias Colorretais/patologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Knockout , Via de Sinalização Wnt/genética , Quinases raf/genética , Proteínas ras/genéticaRESUMO
Sterol regulatory element-binding proteins (SREBPs) belong to a family of transcription factors that regulate the expression of genes required for the synthesis of fatty acids and cholesterol. Three SREBP isoforms, SREBP1a, SREBP1c, and SREBP2, have been identified in mammalian cells. SREBP1a and SREBP1c are derived from a single gene through the use of alternative transcription start sites. Here we investigated the role of SREBP-mediated lipogenesis in regulating tumor growth and initiation in colon cancer. Knockdown of either SREBP1 or SREBP2 decreased levels of fatty acids as a result of decreased expression of SREBP target genes required for lipid biosynthesis in colon cancer cells. Bioenergetic analysis revealed that silencing SREBP1 or SREBP2 expression reduced the mitochondrial respiration, glycolysis, as well as fatty acid oxidation indicating an alteration in cellular metabolism. Consequently, the rate of cell proliferation and the ability of cancer cells to form tumor spheroids in suspension culture were significantly decreased. Similar results were obtained in colon cancer cells in which the proteolytic activation of SREBP was blocked. Importantly, knockdown of either SREBP1 or SREBP2 inhibited xenograft tumor growth in vivo and decreased the expression of genes associated with cancer stem cells. Taken together, our findings establish the molecular basis of SREBP-dependent metabolic regulation and provide a rationale for targeting lipid biosynthesis as a promising approach in colon cancer treatment.
