RESUMO
Mitochondria play an important role in the initiation of apoptosis. However, whether cisplatin can induce apoptosis by initiating a mitochondrial fission pathway and the mechanism underlying this effect remain poorly understood. In this study, we show that the mitochondrial fission protein FIS1 is upregulated upon cisplatin treatment in tongue squamous cell carcinoma (TSCC) cells. FIS1 knockdown can attenuate mitochondrial fission and cisplatin sensitivity. We found that FIS1 is a direct target of miR-483-5p and that miR-483-5p can inhibit mitochondrial fission and cisplatin sensitivity in vitro and in vivo. Furthermore, we found that miR-483-5p and FIS1 are significantly associated with cisplatin sensitivity and with overall survival in patients with TSCC in a retrospective analysis of multiple centers. This study revealed that a novel mitochondrial fission pathway composed of miR-483-5p and FIS1 regulates cisplatin sensitivity. The modulation of miR-483-5p and FIS1 levels may provide a new approach for increasing cisplatin sensitivity.
Assuntos
Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/genética , Cisplatino/farmacologia , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/genética , Proteínas de Membrana/genética , MicroRNAs/genética , Dinâmica Mitocondrial/genética , Proteínas Mitocondriais/genética , Neoplasias da Língua/tratamento farmacológico , Neoplasias da Língua/genética , Animais , Antineoplásicos/farmacologia , Carcinoma de Células Escamosas/metabolismo , Linhagem Celular Tumoral , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/metabolismo , Dinâmica Mitocondrial/efeitos dos fármacos , Proteínas Mitocondriais/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço , Neoplasias da Língua/metabolismo , Transfecção , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The IκB kinases IKKα and IKKß and the IKK-related kinases TANK-binding kinase 1 (TBK1) and IKKε are the master regulators of the NF-κB signaling pathway. Although this pathway has been extensively studied in mammals, less attention has been paid in crustaceans, which have significant economic value. Here, we report the cloning and functional studies of two IKK homologs, LvIKKß and LvIKKε, from Pacific white shrimp, Litopenaeus vannamei. LvIKKß and LvIKKε mRNAs are widely expressed in different tissues and are responsive to white spot syndrome virus (WSSV) infection. When overexpressed in Drosophila S2 cells, LvIKKß but not LvIKKε activates the promoters of NF-κB pathway-controlled antimicrobial peptide genes (AMPs), such as the Penaeidins (PENs). In HEK 293T cells, both LvIKKß and LvIKKε activate an NF-κB reporter. The silencing of LvIKKß or LvIKKε using double-stranded RNA (dsRNA)-mediated RNA interference (RNAi) decreases the expression of L. vannamei AMPs, including PENs, lysozyme and crustins. Intriguingly, LvIKKß- or LvIKKε-silenced L. vannamei are resistant to WSSV infection. We hypothesized that successful infection with WSSV requires the activation of the IKK-NF-κB signaling pathway to modulate viral gene expression. We constructed luciferase reporters for 147 WSSV genes. By screening, we found that the WSV051, WSV059, WSV069, WSV083, WSV090, WSV107, WSV244, WSV303, WSV371 and WSV445 promoters can be activated by LvIKKß or LvIKKε in Drosophila S2 cells. Taken together, our results reveal that LvIKKß and LvIKKε may participate in the regulation of shrimp AMPs and that WSSV may subvert the L. vannamei IKK-NF-κB signaling pathway to facilitate viral gene expression.
Assuntos
Peptídeos Catiônicos Antimicrobianos/genética , Regulação Viral da Expressão Gênica , Quinase I-kappa B/genética , Penaeidae/imunologia , Transdução de Sinais , Vírus da Síndrome da Mancha Branca 1/genética , Animais , Peptídeos Catiônicos Antimicrobianos/imunologia , Peptídeos Catiônicos Antimicrobianos/metabolismo , Linhagem Celular , Clonagem Molecular , Drosophila melanogaster/citologia , Inativação Gênica , Genes Reporter , Interações Hospedeiro-Patógeno , Humanos , Quinase I-kappa B/antagonistas & inibidores , Quinase I-kappa B/imunologia , Luciferases/genética , Luciferases/metabolismo , Muramidase/genética , Muramidase/metabolismo , NF-kappa B/genética , NF-kappa B/imunologia , NF-kappa B/metabolismo , Penaeidae/genética , Penaeidae/virologia , Regiões Promotoras Genéticas , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Vírus da Síndrome da Mancha Branca 1/imunologia , Vírus da Síndrome da Mancha Branca 1/metabolismoRESUMO
Invertebrates rely on innate immunity as the first line defense against microbes. In Drosophila, the inducible antimicrobial peptides (AMPs) regulated by the Toll and immune deficiency (Imd) pathways are important effectors in innate immunity. Here we report an immune deficiency homolog (LvIMD) from the white shrimp, Litopenaeus vannamei. The full-length cDNA of LvIMD is 758 bp with an open reading frame of 483 bp that encodes a putative protein of 160 amino acids including a death domain at the C-terminus. LvIMD death domain shows similarity to that of Drosophila IMD and human receptor interacting protein 1 (RIP1) of the tumor necrosis factor receptor (TNFR) pathway, with 27.9% and 26.4% identity, respectively. Phylogenetic analysis shows that LvIMD clusters with a predicted protein from the starlet sea anemone (Nematostella vectensis) independent to insect IMDs and vertebrates RIP1s. LvIMD mRNA is expressed in most tissues and is induced in hepatopancreas and hemocytes after immune challenge. Luciferase reporter assays confirm that LvIMD is able to induce the expression of AMP genes, including Drosophila Attacin A and shrimp Penaeidin 4 in S2 cells. To our knowledge, this is the first report that LvIMD participates in innate signaling to activate the expression of AMP genes in shrimp.
