Hydrogen Peroxide Is Involved in β-Cyclodextrin-hemin Complex-Induced Lateral Root Formation in Tomato Seedlings

Although previous results showed that β-cyclodextrin-hemin complex (β-CDH) could induce tomato lateral root (LR) formation, the corresponding downstream messengers are still not fully understood. In this report, similar to the inducing effects of exogenously applied hydrogen peroxide (H₂O₂), we discovered that β-CDH elicited RBOH1 transcript upregulation, endogenous H₂O₂ accumulation, and thereafter tomato LR development. Above responses were sensitive to dimethylthiourea (DMTU) and ascorbic acid (AsA), two membrane-permeable scavengers of H₂O₂, showing that accumulation of H₂O₂ and LR formation were significantly blocked. The test with diphenyleneiodonium (DPI; the inhibitor of NADPH oxidase) revealed that H₂O₂ mainly produced by NADPH oxidase, might be involved in LR formation triggered by β-CDH. qPCR combined with pharmacological and anatomical analyses showed that β-CDH-modulated several marker genes responsible for LR formation, such as CYCA3;1, CYCA2;1, CYCD3;1, and CDKA1 (four cell cycle regulatory genes), ARF7 and RSI-1 (two auxin signaling genes), LAX3 (an auxin influx carrier), IAA14 (encoding a member of the Aux/IAA protein family), PIN3 and PIN7 (two auxin efflux carriers), isocitrate dehydrogenase [NADP], NADH-cytochrome b, and L-ascorbate oxidase homolog genes (two reactive oxygen species-associated genes and one LR formation-related gene), were causally related to above H₂O₂ signaling. Particularly, representative proteins related to H₂O₂ metabolism and lateral rooting, were specifically induced in β-CDH-treated tomato seedlings. Overall, the results clearly suggested a vital role of H₂O₂ in the β-CDH-induced tomato LR formation, and β-CDH-elicited H₂O₂-related target proteins responsible for LR formation might be, at least partially, regulated at transcriptional and translational levels.