RESUMO
Autoimmune diseases such as ankylosing spondylitis (AS) can be driven by emerging neoantigens that disrupt immune tolerance. Here, we developed a workflow to profile posttranslational modifications involved in neoantigen formation. Using mass spectrometry, we identified a panel of cysteine residues differentially modified by carboxyethylation that required 3-hydroxypropionic acid to generate neoantigens in patients with AS. The lysosomal degradation of integrin αIIb [ITGA2B (CD41)] carboxyethylated at Cys96 (ITGA2B-ceC96) generated carboxyethylated peptides that were presented by HLA-DRB1*04 to stimulate CD4+ T cell responses and induce autoantibody production. Immunization of HLA-DR4 transgenic mice with the ITGA2B-ceC96 peptide promoted colitis and vertebral bone erosion. Thus, metabolite-induced cysteine carboxyethylation can give rise to pathogenic neoantigens that lead to autoreactive CD4+ T cell responses and autoantibody production in autoimmune diseases.
Assuntos
Autoanticorpos , Doenças Autoimunes , Cisteína , Cadeias HLA-DRB1 , Integrina alfa2 , Processamento de Proteína Pós-Traducional , Espondilite Anquilosante , Animais , Camundongos , Autoanticorpos/metabolismo , Doenças Autoimunes/genética , Doenças Autoimunes/metabolismo , Autoimunidade/genética , Autoimunidade/imunologia , Cisteína/metabolismo , Cadeias HLA-DRB1/genética , Cadeias HLA-DRB1/metabolismo , Camundongos Transgênicos , Integrina alfa2/metabolismo , Microbioma Gastrointestinal , Humanos , Espondilite Anquilosante/genética , Espondilite Anquilosante/metabolismoRESUMO
Esophageal squamous cell carcinoma (ESCC) is one of the most aggressive cancers and is highly resistant to current treatments. ESCC harbors a subpopulation of cells exhibiting cancer stem-like cell (CSC) properties that contribute to therapeutic resistance including radioresistance, but the molecular mechanisms in ESCC CSCs are currently unknown. Here, we report that ribosomal S6 protein kinase 4 (RSK4) plays a pivotal role in promoting CSC properties and radioresistance in ESCC. RSK4 was highly expressed in ESCC CSCs and associated with radioresistance and poor survival in patients with ESCC. RSK4 was found to be a direct downstream transcriptional target of ΔNp63α, the main p63 isoform, which is frequently amplified in ESCC. RSK4 activated the ß-catenin signaling pathway through direct phosphorylation of GSK-3ß at Ser9. Pharmacologic inhibition of RSK4 effectively reduced CSC properties and improved radiosensitivity in both nude mouse and patient-derived xenograft models. Collectively, our results strongly suggest that the ΔNp63α/RSK4/GSK-3ß axis plays a key role in driving CSC properties and radioresistance in ESCC, indicating that RSK4 is a promising therapeutic target for ESCC treatment.
Assuntos
Neoplasias Esofágicas/enzimologia , Carcinoma de Células Escamosas do Esôfago/enzimologia , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/biossíntese , Tolerância a Radiação , Proteínas Quinases S6 Ribossômicas 90-kDa/biossíntese , Transdução de Sinais , Animais , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Neoplasias Esofágicas/terapia , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/patologia , Carcinoma de Células Escamosas do Esôfago/terapia , Células HEK293 , Humanos , Camundongos , Proteínas de Neoplasias/genética , Proteínas Quinases S6 Ribossômicas 90-kDa/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
MiR-145 is a tumor-suppressive microRNA that participates in the malignant progression of colorectal cancer (CRC). Although miR-145 has been reported to inhibit proliferation and to induce apoptosis of CRC cells, the reports about its role in invasion and metastasis are controversial. The regulation of miR-145 its own expression also requires further elucidation. In this study, we firstly found that miR-145 is markedly downregulated in the metastatic tumors of CRC patients. Then through gain- and loss-of function studies, we demonstrated that miR-145 suppresses the invasion and metastasis of CRC cells. We also provided experimental evidences which include direct binding assays and verifications on tissue specimens to confirm that LIM and SH3 protein 1 (LASP1) is a direct target of miR-145. Furthermore, we identified the core promoter regions of miR-145 and observed the cooperation between histone methylation and transcription factors through binding to these core promoter regions to regulate the expression of miR-145 in CRC cells. Our study provides an insight into the regulatory network in CRC cells, thus offering new targets for treating CRC patients.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias do Colo/genética , Proteínas do Citoesqueleto/genética , Regulação Neoplásica da Expressão Gênica , Proteínas com Domínio LIM/genética , MicroRNAs/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Linhagem Celular Tumoral , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Proteínas do Citoesqueleto/metabolismo , Epigênese Genética , Transição Epitelial-Mesenquimal/genética , Feminino , Perfilação da Expressão Gênica , Células HCT116 , Células HEK293 , Histonas/metabolismo , Humanos , Proteínas com Domínio LIM/metabolismo , Masculino , Metilação , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Invasividade Neoplásica , Metástase Neoplásica , Transplante HeterólogoRESUMO
The purpose of this study was to explore the effect of proteasome inhibitor, bortezomib (Bzb), on osteoblast in pathologic status of myeloma bone disease. The myeloma bone disease was modeled by co-culture of mouse myeloma cell RPMI8226 with osteoblast line MC-3T3E1 from mouse calvaria, and intervenient culture of supernatant. The inhibitory effect of Bzb on proliferation of MC-3T3E1 assayed by modified MTT method, the apoptosis of MC-3T3E1 cells was determined by flow cytometry with Annexin V/PI staining, the expressions of osteoblast markers, Runx2/cbfa1, osteocalcin (OCN) and osterix (OSX) in MC-3T3E1 treated with Bzb were detected by RT-PCR and Western blot respectively. Experiments were divided into 3 group: single cultured, co-cultured and supernatant-interveniently cultured groups. The results showed the Bzb in higher concentration inhibited proliferation of MC-3T3E1 cells in a dose-dependent manner, with the IC(50) of 38.1 nmol/L for 48 hours, the Bzb in low concentration (5 nmol/L) did not show the inhibitory effect on proliferation of MC-3T3E1 in single cultured group (p>0.10), but could decrease apoptotic rate of MC-3T3E1 by 32.5% and 24.6% respectively in cocultured and supernatant-interveniently cultured groups, moreover increased the expression of osteoblast-related gene OSX, OCN mRNA and protein (p<0.05), while no obvious change of Runx2/cbfa1 expression was observed (p>0.05). It is concluded that the proteasome inhibitor, Bzb, in low concentration promotes the activity of osteoblast internal mechanisms, and prevents the apoptosis of osteoblasts induced by myeloma cells. In addition, it can up-regulate transcription and expression of osteoblast markers related to Runx2/cbfa1 path way, thus may protect osteoblasts in myeloma bone disease.