RLF, a cytochrome b5-like heme/steroid binding domain protein, controls lateral root formation independently of ARF7/19-mediated auxin signaling in Arabidopsis thaliana

Lateral root (LR) formation is important for the establishment of root architecture in higher plants. Recent studies have revealed that LR formation is regulated by an auxin signaling pathway that depends on auxin response factors ARF7 and ARF19, and auxin/indole-3-acetic acid (Aux/IAA) proteins including SOLITARY-ROOT (SLR)/IAA14. To understand the molecular mechanisms of LR formation, we isolated a recessive mutant rlf (reduced lateral root formation) in Arabidopsis thaliana. The rlf-1 mutant showed reduction of not only emerged LRs but also LR primordia. Analyses using cell-cycle markers indicated that the rlf-1 mutation inhibits the first pericycle cell divisions involved in LR initiation. The rlf-1 mutation did not affect auxin-induced root growth inhibition but did affect LR formation over a wide range of auxin concentrations. However, the rlf-1 mutation had almost no effect on auxin-inducible expression of LATERAL ORGAN BOUNDARIES-DOMAIN16/ASYMMETRIC LEAVES2-LIKE18 (LBD16/ASL18) and LBD29/ASL16 genes, which are downstream targets of ARF7/19 for LR formation. These results indicate that ARF7/19-mediated auxin signaling is not blocked by the rlf-1 mutation. We found that the RLF gene encodes At5g09680, a protein with a cytochrome b5-like heme/steroid binding domain. RLF is ubiquitously expressed in almost all organs, and the protein localizes in the cytosol. These results, together with analysis of the genetic interaction between the rlf-1 and arf7/19 mutations, indicate that RLF is a cytosolic protein that positively controls the early cell divisions involved in LR initiation, independent of ARF7/19-mediated auxin signaling.