Extended Data Fig. 2: Discovery and characterization of homopolymeric DNA products elicited by DRT9 immune systems.
From: Protein-primed homopolymer synthesis by an antiviral reverse transcriptase
a, Unmapped reads from a cDIP-seq dataset of SenDRT9-expressing cells infected with T5 phage, showing uninterrupted strings of poly-dT and poly-dA. b, RIP-seq and cDIP-seq coverage tracks (top to bottom) for either WT or RT-inactive (MUT) PsaDRT9, in the presence of T5 phage infection. A schematic of the genomic locus is shown below; data are normalized for sequencing depth and plotted as counts per million reads (CPM). c, MEME analysis results revealing a poly-dT motif enriched in unmapped reads from the WT + T5 cDIP-seq dataset in b. E, E-value significance; n, number of contributing sites. d, Methylene blue-stained membrane used for the Southern blot shown in Fig. 1h. Representative data are shown for experiments repeated at least two times with similar results. e, Schematic of oligo spike-in experiment to address potential bias in next-generation sequencing-based detection of poly-dA and poly-dT on the AVITI platform; P, phosphorylated 5′ end. f, Bar graph of dA25 and dT25 counts from total DNA sequencing of oligo spike-in experiments schematized in e; the apparent bias against poly-dA capture and sequencing leads to artificially elevated levels of dT25-containing reads relative to dA25-containing reads. g, Bar graph of normalized homopolymer counts from total RNA-seq datasets of WT and MUT SenDRT9-expressing cells +/– T5 phage infection. These data demonstrate that poly-dA and poly-dT cDNAs are not transcribed, in contrast to the cDNA products of DRT2. Data in f,g are shown as mean ± s.d. for n = 3 independent biological replicates.
