The influenza A virus endoribonuclease PA-X usurps host mRNA processing machinery to limit host gene expression

Lea Gaucherand; Brittany K. Porter; Summer K. Schmaling; Christopher Harley Rycroft; Yuzo Kevorkian; Craig McCormick; Denys A. Khaperskyy; Marta Maria Gaglia

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. used, except --library-type fr-firststrand and -u, and a gtf of the hg19 annotation was provided as 664 reference. Only previously annotated RNAs with a level of >1 FPKM in all the control samples 665 (A549 + dox or mock-infected A549) were used in further analyses. The FPKMs for these RNAs 666 were converted to attomoles of RNA based on the known concentration of the spike-in controls. 667 Table S2 shows the high correlation in measured RNA levels between replicate samples. The 668 absolute RNA levels in the replicates were averaged, and the relative RNA levels (RNA ratio) in 669 infected vs. mock infected or PA-X-expressing vs. control cells was computed. All downstream 670 analysis was carried out on the relative levels (ratio) in log2 scale. Tables S3 and S4 summarize 671 the results of the RNAseq analyses. RNAseq data is available on the GEO database, entry 672 GSE120183 (raw read data and processed data included here as Tables S3 and S4). The hg19 673 annotation was used to derive the number of exons, length of transcripts and GC content for the 674 analyses in Figures 4, S3 . For analysis of intronless RNAs, only RNAs that were longer than 300 675 nt were used, to exclude small RNAs that are transcribed by Pol III or are produced through 676 processing of longer transcripts. The length distribution of the remaining intronless RNAs was 677 similar to that of the spliced RNAs. All RNAs were used for the analysis of PA-X down-678 regulation vs. length, exon number, and GC content. k-means clustering analysis was carried out 679 using the Cluster 3.0 program (http://bonsai.hgc.jp/~mdehoon/software/cluster/software.htm). 680