Characterization of the transcriptome of Haloferax volcanii, grown under four different conditions, with mixed RNA-Seq

• Published in: PloS one 2019-04, Vol.14 (4), p.e0215986-e0215986
• Main Authors: Laass, Sebastian, Monzon, Vivian A, Kliemt, Jana, Hammelmann, Matthias, Pfeiffer, Friedhelm, Förstner, Konrad U, Soppa, Jörg
• Format: Article
• Language: English
• Subjects: 3' Untranslated Regions - genetics; 5' Untranslated Regions - genetics; Antisense RNA; Biofilms; Bioinformatics; Biology; Biology and Life Sciences; Cell cycle; Computational biology; Deletion mutant; Gene clusters; Gene expression; Gene Expression Regulation, Archaeal; Genes; Genetic aspects; Genomes; Genomics; Halobacteria; Haloferax volcanii - genetics; Haloferax volcanii - growth & development; Isoforms; Messenger RNA; Multigene Family; Mutants; Open Reading Frames - genetics; Operon - genetics; Proteins; Regulation; Research and Analysis Methods; Ribonucleic acid; RNA; RNA sequencing; RNA, Messenger - genetics; RNA, Messenger - metabolism; RNA, Untranslated - genetics; RNA, Untranslated - metabolism; RNA-Seq; Transcription; Transcriptome - genetics
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0215986

Haloferax volcanii is a well-established model species for haloarchaea. Small scale RNomics and bioinformatics predictions were used to identify small non-coding RNAs (sRNAs), and deletion mutants revealed that sRNAs have important regulatory functions. A recent dRNA-Seq study was used to characterize the primary transcriptome. Unexpectedly, it was revealed that, under optimal conditions, H. volcanii contains more non-coding sRNAs than protein-encoding mRNAs. However, the dRNA-Seq approach did not contain any length information. Therefore, a mixed RNA-Seq approach was used to determine transcript length and to identify additional transcripts, which are not present under optimal conditions. In total, 50 million paired end reads of 150 nt length were obtained. 1861 protein-coding RNAs (cdRNAs) were detected, which encoded 3092 proteins. This nearly doubled the coverage of cdRNAs, compared to the previous dRNA-Seq study. About 2/3 of the cdRNAs were monocistronic, and 1/3 covered more than one gene. In addition, 1635 non-coding sRNAs were identified. The highest fraction of non-coding RNAs were cis antisense RNAs (asRNAs). Analysis of the length distribution revealed that sRNAs have a median length of about 150 nt. Based on the RNA-Seq and dRNA-Seq results, genes were chosen to exemplify characteristics of the H. volcanii transcriptome by Northern blot analyses, e.g. 1) the transcript patterns of gene clusters can be straightforward, but also very complex, 2) many transcripts differ in expression level under the four analyzed conditions, 3) some genes are transcribed into RNA isoforms of different length, which can be differentially regulated, 4) transcripts with very long 5'-UTRs and with very long 3'-UTRs exist, and 5) about 30% of all cdRNAs have overlapping 3'-ends, which indicates, together with the asRNAs, that H. volcanii makes ample use of sense-antisense interactions. Taken together, this RNA-Seq study, together with a previous dRNA-Seq study, enabled an unprecedented view on the H. volcanii transcriptome.