ABSTRACT: We investigated the role of the LysR-type transcriptional regulator VtlR/LsrB (Atu2186) from Agrobacterium tumefaciens C58 (alias A. fabrum C58), which regulates the expression of small-regulatory-RNA (sRNA) genes such as AbcR1 and the expression of genes encoding small proteins with a domain of unknown function DUF1127. For this purpose, we constructed a vtlR/lsrB deletion mutant. Total RNA was isolated from this mutant as well as from an AbcR1 deletion mutant (Wilms et al., 2011) at optical densities (OD) of 0.5 or 0.5 and 1.5, respectively. Sequencing was done on the Illumina NovaSeq 6000 platform. In addition to annotated genes (NCBI genome: 13606), almost 700 previously described sRNA genes (Wilms et al., 2012; Lee et al., 2013; Möller et al., 2014) were also considered within our data analysis. Wild-type samples that derived from the same experiment were published previously (Kraus et al., 2020) (GEO accession: GSE150941). In the vtlR/lsrB mutant a total of 542 genes were downregulated and 665 were upregulated at least three-fold (p value ≤ 0.05). Among those were 25 downregulated sRNA genes (including AbcR1) and 77 upregulated sRNA genes. In the AbcR1 mutant 8 or 20 genes were downregulated at ODs 0.5 or 1.5, respectively, and 72 or 31 were upregulated. A total of 39 differentially expressed genes from the vtlR/lsrB mutant also displayed identical regulation in the AbcR1 mutant and thus are controlled indirectly by VtlR/LsrB via the sRNA AbcR1. Overall, the results demonstrate the complex network of VtlR/LsrB and regulatory sRNAs, which has a great impact on the transcriptome of A. tumefaciens. Kraus, A., Weskamp, M., Zierles, J., Balzer, M., Busch, R., Eisfeld, J., et al. (2020) Arginine-rich small proteins with a domain of unknown function DUF1127 play a role in phosphate and carbon metabolism of Agrobacterium tumefaciens. J Bacteriol 202: e00309-20. Lee, K., Huang, X., Yang, C., Lee, D., Ho, V., Nobuta, K., et al. (2013) A genome-wide survey of highly expressed non-coding RNAs and biological validation of selected candidates in Agrobacterium tumefaciens. PLoS One 8: e70720. Möller, P., Overlöper, A., Förstner, K.U., Wen, T.N., Sharma, C.M., Lai, E.M., and Narberhaus, F. (2014) Profound impact of Hfq on nutrient acquisition, metabolism and motility in the plant pathogen Agrobacterium tumefaciens. PLoS One 9: e110427. Wilms, I., Overlöper, A., Nowrousian, M., Sharma, C.M., and Narberhaus, F. (2012) Deep sequencing uncovers numerous small RNAs on all four replicons of the plant pathogen Agrobacterium tumefaciens. RNA Biol 9: 446–57. Wilms, I., Voss, B., Hess, W.R., Leichert, L.I., and Narberhaus, F. (2011) Small RNA-mediated control of the Agrobacterium tumefaciens GABA binding protein. Mol Microbiol 80: 492–506.
