Project description:The mouse hepatitis virus (MHV) genomic and sub-genomic RNAs have 3’ poly(A) tails. The terminal addition of uridines to poly(A) tails has been shown to be a widespread modification. Here, we investigated the presence of 3' end additions on the MHV RNA poly(A) tails. To this end, we infected NCTC cells with MHV and isolated RNA at 24-hours post-infection (hpi). While the median poly(A) tail length of the MHV RNAs is around 50 nucleotides, we observed a peak of uridylation in transcripts with poly(A) tails about 40 nucleotides long
Project description:The mouse hepatitis virus (MHV) genomic RNA has a poly(A) tail required for replication. Here we investigated the presence of terminal poly(A) tail uridylation and guanylation in the virion RNA. After isolating the viral RNA followed by sequencing, we found a mean poly(A) tail of 66 nucleotides long with a peak of terminal uridylation on tails 55 to 66 nucleotides long.
Project description:The mouse hepatitis virus (MHV) genomic and sub-genomic RNAs are 3’ polyadenylated. The length of the 3’ poly(A) tail is known to change during infection, but little is known about the molecular mechanisms driving this change. Here, we investigate the presence of terminal uridylation and terminal guanylation on the MHV poly(A) tail during infection. We infected 17-CL1 cells with MHV and isolated RNA at 24- and 48-hour post-infection (hpi). We observed that MHV RNAs poly(A) tails shortened during infection and that short poly(A) tails of about 20 nucleotides are highly uridylated.
Project description:Terminal uridylation of mRNAs poly(A) tails by Terminal Uridylyl Transferases (TUTs) promote transcript decay. Here, we investigate the role of two functionally redundant mammalian TUTs, TUT4 and TUT7 (TUT4/7), in mouse hepatitis virus (MHV) RNA processing. We generated a TUT4/7 knock-down 17-CL1 cell line using lentivirus encoding shRNAs against Tut4 and Tut7 transcripts (shTUT4/7) and a control 17-CL1 cell line (shCTL) with a non-targeting shRNA. We then infected the shCTL and shTUT4/7 cells with MHV and isolated RNA at 24- and 48-hours post-infection (hpi). We found that viral RNA poly(A) tails of about 20 and 40 nucleotides are highly uridylated and that the uridylation of the 20 nucleotide long tails is dependent on TUT4/7. Depletion of TUT4/7 also resulted in an accumulation of the viral RNA.
Project description:Investigation of whole genome gene expression level changes in a Bacteroides fragilis NCTC 9343 delta-gmd-fcl delta-fkp mutant strain and a Bacteroides fragilis NCTC 9343 delta-lfg mutant strain, each as compared to the wild-type strain. The mutations engineered into these strains interfere with B. fragilis protein glycosylation.