Mechanisms and functions of poxvirus-induced micro-RNA degradation

• Applicant: Dr. Simone Backes
• Subject Area: Virology
• Term: since 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 447157729

MicroRNAs (miRNAs) are genome-encoded small non-coding RNAs that play fundamental roles in many cellular processes by inhibiting the expression of complementary messenger RNAs (mRNAs). Despite the potential to specifically target small RNAs for treatment and prophylaxis of infectious diseases, the role of small non-coding RNAs, expressed by either the host or by viral pathogens, is not sufficiently understood. We have discovered that the poxviral poly(A) polymerase VP55 induces the specific, sequence-independent poly(A)-tailing of all miRNAs, which results in their rapid degradation by a yet poorly characterized but highly conserved cellular pathway. This first demonstration of viral antagonism of global host miRNA function suggests a functional role of small RNA-mediated silencing during infection and raises several questions about the evolution of viral strategies to counteract and to exploit these pathways. Here, we aim to determine the importance of small RNA antagonism for the poxviral life-cycle and to utilize VP55-induced miRNA polyadenylation as a model system to identify factors and mechanisms involved in polyadenylation-induced miRNA degradation . In Aim 1, we will analyze replication and gene expression of the prototypical poxvirus Vaccinia (VACV) in the presence or absence of functional small RNAs. In addition, we will investigate the role of virus-derived small interfering RNAs (vsiRNAs), which are miRNA-like molecules generated from viral RNAs that have well-established antiviral roles in plants and invertebrates. It is currently unknown whether vsiRNAs also contribute to the antiviral response in mammals. Moreover, we will employ VP55 as a tool to induce the conditional and synchronized global miRNA degradation with the aim to identify and to study the involved cellular proteins and pathways (Aim 2). We have started to identify cellular VP55-interacting proteins and to determine the impact of these interactions for VACV-induced miRNA degradation and for viral replication. Moreover, we aim to structurally and biochemically analyze miRNA-containing ribonucleoprotein complexes undergoing VP55-induced polyadenylation and degradation.Despite the broad relevance of miRNAs during cellular differentiation, infection, inflammation and cancer, the mechanisms regulating miRNA stability and decay remain poorly understood. Our preliminary data demonstrate that our approach enables the experimental manipulation of miRNA turnover, and thereby overcomes current limitations to study these pathways. The proposed studies will significantly increase our understanding of the role of small non-coding RNAs and the pathways mediating their function and turnover, and will thereby facilitate the identification of novel molecular targets for the therapy and prophylaxis of infectious diseases.

• DFG Programme: Research Grants