RESUMO
Non-coding RNA (ncRNA) regulatory networks are emerging as critical regulators of gene expression. These intricate networks of ncRNA:ncRNA interactions modulate multiple cellular pathways and impact the development and progression of multiple diseases. Herpesviruses, including Kaposi's sarcoma-associated herpesvirus, are adept at utilising ncRNAs, encoding their own as well as dysregulating host ncRNAs to modulate virus gene expression and the host response to infection. Research has mainly focused on unidirectional ncRNA-mediated regulation of target protein-coding transcripts; however, we identify a novel host ncRNA regulatory network essential for KSHV lytic replication in B cells. KSHV-mediated upregulation of the host cell circRNA, circHIPK3, is a key component of this network, functioning as a competing endogenous RNA of miR-30c, leading to increased levels of the miR-30c target, DLL4. Dysregulation of this network highlights a novel mechanism of cell cycle control during KSHV lytic replication in B cells. Importantly, disruption at any point within this novel ncRNA regulatory network has a detrimental effect on KSHV lytic replication, highlighting the essential nature of this network and potential for therapeutic intervention.
Assuntos
Herpesvirus Humano 8 , MicroRNAs , Linfócitos B , Herpesvirus Humano 8/genética , MicroRNAs/genética , RNA Circular/genética , Regulação para CimaRESUMO
Human papillomaviruses (HPV) are a major cause of malignancy worldwide. They are the aetiological agents of almost all cervical cancers as well as a sub-set of other anogenital and head and neck cancers. Hijacking of host cellular pathways is essential for virus pathogenesis; however, a major challenge remains to identify key host targets and to define their contribution to HPV-driven malignancy. The Hippo pathway regulates epithelial homeostasis by down-regulating the function of the transcription factor YAP. Increased YAP expression has been observed in cervical cancer but the mechanisms driving this increase remain unclear. We found significant down-regulation of the master Hippo regulatory kinase STK4 (also termed MST1) in cervical disease samples and cervical cancer cell lines compared with healthy controls. Re-introduction of STK4 inhibited the proliferation of HPV positive cervical cells and this corresponded with decreased YAP nuclear localization and decreased YAP-dependent gene expression. The HPV E6 and E7 oncoproteins maintained low STK4 expression in cervical cancer cells by upregulating the oncomiR miR-18a, which directly targeted the STK4 mRNA 3'UTR. Interestingly, miR-18a knockdown increased STK4 expression and activated the Hippo pathway, significantly reducing cervical cancer cell proliferation. Our results identify STK4 as a key cervical cancer tumour suppressor, which is targeted via miR-18a in HPV positive tumours. Our study indicates that activation of the Hippo pathway may offer a therapeutically beneficial option for cervical cancer treatment.
Assuntos
Transformação Celular Viral , MicroRNAs/metabolismo , Papillomaviridae/metabolismo , Infecções por Papillomavirus/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , RNA Neoplásico/metabolismo , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismo , Neoplasias do Colo do Útero/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , MicroRNAs/genética , Proteínas Oncogênicas Virais/genética , Proteínas Oncogênicas Virais/metabolismo , Papillomaviridae/genética , Infecções por Papillomavirus/genética , Infecções por Papillomavirus/patologia , Infecções por Papillomavirus/virologia , Proteínas Serina-Treonina Quinases/genética , RNA Neoplásico/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/genética , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/virologia , Proteínas de Sinalização YAPRESUMO
Historically, ribosomes were viewed as unchanged homogeneous macromolecular machines with no regulatory capacity for mRNA translation. An emerging concept is that heterogeneity of ribosomal composition exists, exerting a regulatory function or specificity in translational control. This is supported by recent discoveries identifying compositionally distinct specialised ribosomes that actively regulate mRNA translation. Viruses lack their own translational machinery and impose high translational demands on the host during replication. We explore the possibility that KSHV manipulates ribosome biogenesis producing specialised ribosomes which preferentially translate viral transcripts. Quantitative proteomic analysis identified changes in the stoichiometry and composition of precursor ribosomal complexes during the switch from latent to lytic replication. We demonstrate the enhanced association of ribosomal biogenesis factors BUD23 and NOC4L, and the KSHV ORF11 protein, with small ribosomal subunit precursor complexes during lytic replication. BUD23 depletion resulted in significantly reduced viral gene expression, culminating in dramatic reduction of infectious virion production. Ribosome profiling demonstrated BUD23 is essential for reduced association of ribosomes with KSHV uORFs in late lytic genes, required for the efficient translation of the downstream coding sequence. Results provide mechanistic insights into KSHV-mediated manipulation of cellular ribosome composition inducing a population of specialised ribosomes facilitating efficient translation of viral mRNAs.
Assuntos
Herpesvirus Humano 8 , Herpesvirus Humano 8/genética , Replicação Viral/genética , Proteômica , Ribossomos/genética , Regulação Viral da Expressão GênicaRESUMO
The epitranscriptomic modification N6-methyladenosine (m6A) is a ubiquitous feature of the mammalian transcriptome. It modulates mRNA fate and dynamics to exert regulatory control over numerous cellular processes and disease pathways, including viral infection. Kaposi's sarcoma-associated herpesvirus (KSHV) reactivation from the latent phase leads to the redistribution of m6A topology upon both viral and cellular mRNAs within infected cells. Here we investigate the role of m6A in cellular transcripts upregulated during KSHV lytic replication. Our results show that m6A is crucial for the stability of the GPRC5A mRNA, whose expression is induced by the KSHV latent-lytic switch master regulator, the replication and transcription activator (RTA) protein. Moreover, we demonstrate that GPRC5A is essential for efficient KSHV lytic replication by directly regulating NFκB signalling. Overall, this work highlights the central importance of m6A in modulating cellular gene expression to influence viral infection.
Assuntos
Herpesvirus Humano 8 , Herpesvirus Humano 8/genética , Latência Viral/genética , Linhagem Celular Tumoral , Transdução de Sinais , RNA Mensageiro/genética , Replicação Viral , Regulação Viral da Expressão GênicaRESUMO
Circular RNAs (circRNAs) are a group of noncoding RNAs, formed mostly through a unique backsplicing mechanism. Originally proposed to be a byproduct from errors in splicing, recent studies have shown they exhibit a range of roles in regulating gene expression, including sponging of microRNAs (miRNAs), interactions with RNAbinding proteins and regulation of transcription. Though research is still in its infancy, evidence suggests circRNA levels are tightly regulated in the cell, reinforced by dysregulated circRNAs levels being implicated in a range of diseases, including cancer and viral infection. There is growing interest in circRNAs playing specific roles in cancers, either oncogenic or as tumour suppressors, with particular focus on their potential as novel biomarkers. This review will provide an overview of circRNA biogenesis and regulation, and their potential roles in the cell, with a focus on their dysregulation in cancer.