ABSTRACT
The addition of chemical groups to cellular RNA to modulate RNA fate and/or function is summarized under the term epitranscriptomic modification. More than 170 different modifications have been identified on cellular RNA, such as tRNA, rRNA and, to a lesser extent, on other RNA types. Recently, epitranscriptomic modification of viral RNA has received considerable attention as a possible additional mechanism regulating virus infection and replication. N6-methyladenosine (m6A) and C5-methylcytosine (m5C) have been most broadly studied in different RNA viruses. Various studies, however, reported varying results with regard to number and extent of the modification. Here we investigated the m5C methylome of SARS-CoV-2, and we reexamined reported m5C sites in HIV and MLV. Using a rigorous bisulfite-sequencing protocol and stringent data analysis, we found no evidence for the presence of m5C in these viruses. The data emphasize the necessity for optimizing experimental conditions and bioinformatic data analysis.