Silencing Arabidopsis CARBOXYL-TERMINAL DOMAIN PHOSPHATASE-LIKE 4 induces cytokinin-oversensitive de novo shoot organogenesis

De novo shoot organogenesis (DNSO) is a post-embryonic development programme that has been widely exploited by plant biotechnology. DNSO is a hormonally regulated process in which auxin and cytokinin (CK) coordinate suites of genes encoding transcription factors, general transcription factors, and RNA metabolism machinery. Here we report that silencing Arabidopsis thaliana carboxyl-terminal domain (CTD) phosphatase-like 4 (CPL4(RNAi)) resulted in increased phosphorylation levels of RNA polymerase II (pol II) CTD and altered lateral root development and DNSO efficiency of the host plants. Under standard growth conditions, CPL4(RNAi) lines produced no or few lateral roots. When induced by high concentrations of auxin, CPL4(RNAi) lines failed to produce focused auxin maxima at the meristem of lateral root primordia, and produced fasciated lateral roots. In contrast, root explants of CPL4(RNAi) lines were highly competent for DNSO. Efficient DNSO of CPL4(RNAi) lines was observed even under 10 times less the CK required for the wild-type explants. Transcriptome analysis showed that CPL4(RNAi), but not wild-type explants, expressed high levels of shoot meristem-related genes even during priming on medium with a high auxin/CK ratio, and during subsequent shoot induction with a lower auxin/CK ratio. Conversely, CPL4(RNAi) enhanced the inhibitory phenotype of the shoot redifferentiation defective2-1 mutation, which affected snRNA biogenesis and formation of the auxin gradient. These results indicated that CPL4 functions in multiple regulatory pathways that positively and negatively affect DNSO.