Next-Generation Sequencing of Small RNAs from HIV-Infected Cells Identifies Phased microRNA Expression Patterns and Candidate Novel microRNAs Differentially Expressed upon Infection

Chang, Stewart T.; Thomas, Matthew J.; Sova, Pavel; Green, Richard R.; Palermo, Robert E.; Katze, Michael G.

External data also supported the existence of a microRNA at this location. Small RNA-Seq data from the ENCODE project showed that other immune cell types-including GM12878, a lymphoblastoid cell line, and K562, a leukemia-derived cell line (16)-expressed a small RNA mapping to the same location and strand detected by miRDeep (Fig. 4) . This indicated that the EPB41L2-encoded microRNA was robustly expressed and detected, despite differences in cell type, sample processing, and read mapping. In addition, evolutionary comparisons of aligned sequences from primates, mammals, and other vertebrates showed that the sequence for the EPB41L2-encoded microRNA was conserved in higher primates, particularly Old World monkeys and apes, with the exception of gorillas, but not other vertebrates (Fig. 4) . External data sets provided additional evidence that viral infection results in the downregulation of the EPBL41L2-encoded candidate novel microRNA. We obtained raw small RNA-Seq data from a separate infection model system, human cytomegalovirus (HCMV) infection of human fibroblasts (17) , and observed reads mapping to the location of the candidate microRNA in each of four samples (mock-and HCMV-infected at 24 and 72 hpi), suggesting that the microRNA was expressed in human fibroblasts. Moreover, quantification of these reads suggested that HCMV downregulated the expression of this microRNA at 24 hpi (from 13.2 rpm to 5.6 rpm, a change of 58%) and at 72 hpi (from 34.4 rpm to 7.0 rpm, a change of 80%).