Small RNAs Derived from the T-DNA of Agrobacterium rhizogenes in Hairy Roots of Phaseolus vulgaris

Pablo Peláez,Alejandrina Hernández-López,Georgina Estrada-Navarrete,Federico Sanchez

doi:10.3389/fpls.2017.00096

Abstract

Agrobacterium rhizogenes is a pathogenic bacteria that causes hairy root disease by transferring bacterial DNA into the plant genome. It is an essential tool for industry and research due to its capacity to produce genetically modified roots and whole organisms. Here, we identified and characterized small RNAs generated from the transfer DNA (T-DNA) of A. rhizogenes in hairy roots of common bean (Phaseolus vulgaris). Distinct abundant A. rhizogenes T-DNA-derived small RNAs (ArT-sRNAs) belonging to several oncogenes were detected in hairy roots using high-throughput sequencing. The most abundant and diverse species of ArT-sRNAs were those of 21- and 22-nucleotides in length. Many T-DNA encoded genes constituted phasiRNA producing loci (PHAS loci). Interestingly, degradome analysis revealed that ArT-sRNAs potentially target genes of P. vulgaris. In addition, we detected low levels of ArT-sRNAs in the A. rhizogenes-induced calli generated at the wound site before hairy root emergence. These results suggest that RNA silencing targets several genes from T-DNA of A. rhizogenes in hairy roots of common bean. Therefore, the role of RNA silencing observed in this study has implications in our understanding and usage of this unique plant-bacteria interaction.

Highlights

Agrobacterium rhizogenes and Agrobacterium tumefaciens are plant pathogenic bacteria capable of inducing uncontrolled cell proliferation and development of hairy roots and crown gall tumors, respectively
In order to identify small RNAs derived from the transfer DNA (T-DNA) of A. rhizogenes in the hairy root disease, small RNA libraries from hairy roots, and calli were generated
3176 distinct ArTsRNAs corresponding to 17,000 small RNA reads per million (RPM) were detected

Summary

Introduction

Agrobacterium rhizogenes and Agrobacterium tumefaciens are plant pathogenic bacteria capable of inducing uncontrolled cell proliferation and development of hairy roots and crown gall tumors, respectively. A transferred DNA from the bacteria root inducing (Ri) or tumor-inducing (Ti) plasmids is integrated into the plant cells genome (Tzfira and Citovsky, 2006). Transformed cells produce amino acid and sugar derivatives (opines) that are used by bacteria as nutrients and trigger conjugal transfer of T-DNA-carrying plasmids (Subramoni et al, 2014). A. tumefaciens strains with Ti plasmids lacking T-DNA oncogenes do not induce tumor growth and have been used as efficient delivery systems for plant transformation (Krenek et al, 2015). Hairy roots harboring the A. rhizogenes Ri plasmid T-DNA have become an important

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Conclusion