BackgroundDifferential RNA-Seq (dRNA-Seq) is a recently developed method of performing primary transcriptome analyses that allows for the genome-wide mapping of transcriptional start sites (TSSs) and the identification of novel transcripts. Although the transcriptomes of diverse bacterial species have been characterized by dRNA-Seq, the transcriptome analysis of archaeal species is still rather limited. Therefore, we used dRNA-Seq to characterize the primary transcriptome of the model archaeon Haloferax volcanii.ResultsThree independent cultures of Hfx. volcanii grown under optimal conditions to the mid-exponential growth phase were used to determine the primary transcriptome and map the 5′-ends of the transcripts. In total, 4749 potential TSSs were detected. A position weight matrix (PWM) was derived for the promoter predictions, and the results showed that 64 % of the TSSs were preceded by stringent or relaxed basal promoters. Of the identified TSSs, 1851 belonged to protein-coding genes. Thus, fewer than half (46 %) of the 4040 protein-coding genes were expressed under optimal growth conditions. Seventy-two percent of all protein-coding transcripts were leaderless, which emphasized that this pathway is the major pathway for translation initiation in haloarchaea. A total of 2898 of the TSSs belonged to potential non-coding RNAs, which accounted for an unexpectedly high fraction (61 %) of all transcripts. Most of the non-coding TSSs had not been previously described (2792) and represented novel sequences (59 % of all TSSs). A large fraction of the potential novel non-coding transcripts were cis-antisense RNAs (1244 aTSSs). A strong negative correlation between the levels of antisense transcripts and cognate sense mRNAs was found, which suggested that the negative regulation of gene expression via antisense RNAs may play an important role in haloarchaea. The other types of novel non-coding transcripts corresponded to internal transcripts overlapping with mRNAs (1153 iTSSs) and intergenic small RNA (sRNA) candidates (395 TSSs).ConclusionThis study provides a comprehensive map of the primary transcriptome of Hfx. volcanii grown under optimal conditions. Fewer than half of all protein-coding genes have been transcribed under these conditions. Unexpectedly, more than half of the detected TSSs belonged to several classes of non-coding RNAs. Thus, RNA-based regulation appears to play a more important role in haloarchaea than previously anticipated.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2920-y) contains supplementary material, which is available to authorized users.

Background: CRISPR/Cas systems allow archaea and bacteria to resist invasion by foreign nucleic acids.Results: The CRISPR/Cas system in Haloferax recognized six different PAM sequences that could trigger a defense response.Conclusion: The PAM sequence specificity of the defense response in type I CRISPR systems is more relaxed than previously thought.Significance: The PAM sequence requirements for interference and adaptation appear to differ markedly.

Accurate tRNA 3¢ end maturation is essential for aminoacylation and thus for protein synthesis in all organisms. Here we report the ®rst identi®cation of protein and DNA sequences for tRNA 3¢-processing endonucleases (RNase Z). Puri®cation of RNase Z from wheat identi®ed a 43 kDa protein correlated with the activity. Peptide sequences obtained from the puri®ed protein were used to identify the corresponding gene. In vitro expression of the homologous proteins from Arabidopsis thaliana and Methanococcus janaschii con®rmed their tRNA 3¢-processing activities. These RNase Z proteins belong to the ELAC1/2 family of proteins and to the cluster of orthologous proteins COG 1234. The RNase Z enzymes from A.thaliana and M.janaschii are the ®rst members of these families to which a function can now be assigned. Proteins with high sequence similarity to the RNase Z enzymes from A.thaliana and M.janaschii are present in all three kingdoms.

In contrast to Escherichia coli, where the 3¢ ends of tRNAs are primarily generated by exoribonucleases, maturation of the 3¢ end of tRNAs is catalysed by an endoribonuclease, known as RNase Z (or 3¢ tRNase), in many eukaryotic and archaeal systems. RNase Z cleaves tRNA precursors 3¢ to the discriminator base.Here we show that this activity, previously unsuspected in bacteria, is encoded by the yqjK gene of Bacillus subtilis. Decreased yqjK expression leads to an accumulation of a population of B.subtilis tRNAs in vivo, none of which have a CCA motif encoded in their genes, and YqjK cleaves tRNA precursors with the same speci®city as plant RNase Z in vitro. We have thus renamed the gene rnz. A CCA motif downstream of the discriminator base inhibits RNase Z activity in vitro, with most of the inhibition due to the ®rst C residue. Lastly, tRNAs with long 5¢ extensions are poor substrates for cleavage, suggesting that for some tRNAs, processing of the 5¢ end by RNase P may have to precede RNase Z cleavage.

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