Fig. 7. The role of TENT-5 in innate immunity is evolutionarily conserved.

(A) Heatmap showing expression levels of Tent5 genes in BMDMs isolated from the wild-type mice. Stimulation of BMDMs with LPS led to increased expression levels of Tent5a/c. (B) TENT5A/C were elevated in macrophages stimulated with LPS as illustrated by anti-FLAG immunoblots on extracts prepared from wild-type (WT), TENT5A-3xFLAG (AF), and TENT5C-3xFLAG (CF) BMDMs. iNOS and CD80 levels serve as a control of BMDM activation. The lines and asterisks indicate specific and unspecific bands, respectively. (C) Plot showing expression levels and difference in the median poly(A) tail length of mRNAs that were isolated from Tent5a^−/− Tent5c^−/− and wild-type BMDMs. TENT5A/C-regulated transcripts (n = 98) are marked in red (FDR < 0.05). (D) Top 20 (ordered by adjusted P value) GO terms of TENT5A/C substrate mRNAs (data S6). (E) Bee swarm plots of poly(A) tail length distributions for the top 20 TENT5A/C molecular substrates ordered by the median poly(A) tail length fold change (data S6). (F) Western blots showing that the lack of TENT5A/C leads to the lower abundance of indicated peptides both in unstimulated (LPS; 0 hours) and stimulated (LPS; 8 hours) BMDMs. Glyceraldehyde phosphate dehydrogenase (GAPDH) serves as a loading control. Shown is the representative result of three biological replicates (fig. S7K). Graphs on the right show values of the band intensity ratio between tested protein and GAPDH. Points indicate values of each of three biological replicates, and lines denote the average value for biological replicas. *P ≤ 0.05 (two-tailed t test).