On 2016 Oct 03, Duke RNA Biology Journal Club commented:

Overall, we were very impressed with the thoroughness of the paper - we rarely found an experiment that did not have orthogonal experimental validation along with the necessary controls. Leaving the discussion, we were convinced that NRAV plays a role in IAV host immune response and possibly a broader viral response, which control experiments hinted at.

More critically, we felt the paper focused on creating a broad understanding of NRAV’s function in immunity while leaving many loose ends in mechanistic understanding. For instance, an undeveloped part of this paper was the role of ZONAB in NRAV transcriptional repression. While the pulldown assays in Fig 6 I and J suggest ZONAB acts as a transcription factor of MxA, it’s not clear how this is regulated in the presence of NRAV. Perhaps performing a mass spec analysis of all proteins interacting with NRAV, see Fig 6H, would help uncover the rest of the mechanism. Additionally, our group had concerns with Fig 7. The determination of such a long RNA with structure prediction software is risky since longer RNA is more likely to be predicted to fold into many different structures with similar free energy compared to a much smaller RNA, the general query molecules for these programs. We would prefer the authors confirm these predictions with biochemical assays such as RNAse protection and SHAPE-based assays. We were also curious why the NRAV truncation mutants were not used to determine the ZONAB interaction site using pulldown methods.

To summarize, this paper scratches the surface of NRAV’s role in viral host immunity. We hope the authors continue to provide more depth to the story by determining how NRAV is regulated, how NRAV changes histone methylation patterns at ISG transcription sites and how ZONAB specifically fits into this response. We look forward to learning more about the role of lncRNA in the immune response and will be interested to see how the other lncRNAs from Fig 1A fit into this emerging field.

On 2016 Oct 03, Duke RNA Biology Journal Club commented:

Overall, we were very impressed with the thoroughness of the paper - we rarely found an experiment that did not have orthogonal experimental validation along with the necessary controls. Leaving the discussion, we were convinced that NRAV plays a role in IAV host immune response and possibly a broader viral response, which control experiments hinted at.

More critically, we felt the paper focused on creating a broad understanding of NRAV’s function in immunity while leaving many loose ends in mechanistic understanding. For instance, an undeveloped part of this paper was the role of ZONAB in NRAV transcriptional repression. While the pulldown assays in Fig 6 I and J suggest ZONAB acts as a transcription factor of MxA, it’s not clear how this is regulated in the presence of NRAV. Perhaps performing a mass spec analysis of all proteins interacting with NRAV, see Fig 6H, would help uncover the rest of the mechanism. Additionally, our group had concerns with Fig 7. The determination of such a long RNA with structure prediction software is risky since longer RNA is more likely to be predicted to fold into many different structures with similar free energy compared to a much smaller RNA, the general query molecules for these programs. We would prefer the authors confirm these predictions with biochemical assays such as RNAse protection and SHAPE-based assays. We were also curious why the NRAV truncation mutants were not used to determine the ZONAB interaction site using pulldown methods.

To summarize, this paper scratches the surface of NRAV’s role in viral host immunity. We hope the authors continue to provide more depth to the story by determining how NRAV is regulated, how NRAV changes histone methylation patterns at ISG transcription sites and how ZONAB specifically fits into this response. We look forward to learning more about the role of lncRNA in the immune response and will be interested to see how the other lncRNAs from Fig 1A fit into this emerging field.