Transcriptomic Analysis Reveals That Reactive Oxygen Species and Genes Encoding Lipid Transfer Protein Are Associated with Tobacco Hairy Root Growth and Branch Development

Abstract

The hairy root, a specialized plant tissue that emerges from a cell transformed with transfer DNA (T-DNA) from Agrobacterium rhizogenes, can be used to study root biology and utilized in biotechnological applications. The rol genes are known to participate in the generation of hairy roots; however, the means by which the rol genes contribute to the initiation and the maintenance of hairy roots remains largely unknown. We demonstrated that tobacco hairy roots lacking either rolB or rolC exhibited fewer branch roots and lost their growth ability in long-term subculture. Additionally, a microarray analysis revealed that the expression of several genes encoding lipid transfer proteins (LTP) and reactive oxygen species (ROS)-related genes was significantly suppressed in rolB- or rolC-deficient hairy roots. We found that hairy root clones that exhibited greater branching expressed higher levels of RolB or RolC and the genes encoding LTP identified from the microarray. When hairy roots were compared with intact roots, the expression levels of LTP-encoding genes were dramatically different. In addition, ROS were present at lower levels in rolB- and rolC-deficient hairy roots. We therefore suggest that upregulating LTP and increasing the level of ROS is important for hairy root growth.

Efficient CRISPR-mediated base editing in Agrobacterium spp.

“…S8). Our observations are in line with the previously reported crucial roles of rolB and rolC in inducing hairy root formation (10,47,49,50).…”

“…For K599, we designed sgRNAs to target the T-DNA-encoded genes rolB (B0909_24535), rolC (B0909_24530), and orf13 (B0909_24525). The Agrobacterium phenotypic plasticity (plast) genes rolB and rolC were selected because of their significant contribution to the development of the hairy root phenotype (10,47). We also included orf13, another plast gene for which the contribution to the initiation and development of hairy roots is unclear (10,48).…”

Proteomic Analysis of Proteins Related to Defense Responses in Arabidopsis Plants Transformed with the rolB Oncogene

“…Upregulated proteins involved in lipid metabolism are represented by 3-ketoacyl-CoA thiolase 2 (KAT2), UDP-sulfoquinovose synthase, and short-chain dehydrogenase/reductase SDRA (Table 2). This data is consistent with Wang et al [12], who proposed a role for rolB in lipid metabolism. Interestingly, KAT2 is involved in inflorescence development by regulating peroxisomal β-oxidation [47] and benzoylated glucosinolate biosynthesis [48], i.e., KAT2 may be involved in both processes affected by rolB.…”

“…It has been shown that important ROS generators, NADPH oxidase genes AtRbohD and AtRbohF and corresponding proteins, are upregulated in rolB-expressing cells [90], and the deletion of rolB from wild-type hairy roots decreases ROS level in rolB-deficient hairy roots [12]. This data could indicate that ROS levels in rolB-transformed plants should be increased.…”

Agrobacterium Rhizogenes -Mediated Hairy Roots Transformation as a Tool For Exploring Aluminum-Responsive Genes Function

“…The suppression pattern of hairy roots of NtSTOP1 -KD is very similar to that of intact roots of tobacco (Figure 2C & Supplementary Figure 2A). However, the expression patterns of the genes relating to reactive oxygen species (ROS) are different between the intact and the hairy roots, possibly because they are hairy roots [31]. Moreover, Al-tolerant cultured cells of tobacco showed Al tolerance and enhanced citrate excretion [32].…”