SummaryWe have established a detailed framework for the process of shoot regeneration from Arabidopsis root and hypocotyl explants grown in vitro. Using transgenic plant lines in which the GUS or GFP genes were fused to promoters of developmental genes (WUS, CLV1, CLV3, STM, CUC1, PLT1, RCH1, QC25), or to promoters of genes encoding indicators of the auxin response (DR5) or transport (PIN1), cytokinin (CK) response (ARR5) or synthesis (IPT5), or mitotic activity (CYCB1), we showed that regenerated shoots originated directly or indirectly from the pericycle cells adjacent to xylem poles. In addition, shoot regeneration appeared to be partly similar to the formation of lateral root meristems (LRMs). During pre-culture on a 2, 4-dichlorophenoxyacetic acid (2, 4-D)-rich callus-inducing medium (CIM), xylem pericycle reactivation established outgrowths that were not true calli but had many characteristics of LRMs. Transfer to a CK-rich shoot-inducing medium (SIM) resulted in early LRM-like primordia changing to shoot meristems. Direct origin of shoots from the xylem pericycle occurred upon direct culture on CK-containing media without prior growth on CIM. Thus, it appeared that the xylem pericycle is more pluripotent than previously thought. This pluripotency was accompanied by the ability of pericycle derivatives to retain diploidy, even after several rounds of cell division. In contrast, the phloem pericycle did not display such developmental plasticity, and responded to CKs with only periclinal divisions. Such observations reinforce the view that the pericycle is an 'extended meristem' that comprises two types of cell populations. They also suggest that the founder cells for LRM initiation are not initially fully specified for this developmental pathway.

In Arabidopsis, the basic leucine zipper transcription factor ABI5 activates several late embryogenesis-abundant genes, including AtEm1 and AtEm6 . However, the expression of many other seed maturation genes is independent of ABI5 . We investigated the possibility that ABI5 homologs also participate in the regulation of gene expression during seed maturation. We identified 13 ABI5 -related genes in the Arabidopsis genomic sequence. RNA gel blot analysis showed that seven of these genes are active during seed maturation and that they display distinct expression kinetics. We isolated and characterized two mutant alleles of one of these genes, AtbZIP12/EEL . Unlike abi5 , the eel mutations did not inhibit the expression of any of the maturation marker genes that we monitored. On the contrary, the accumulation of the AtEm1 and AtEm6 mRNAs was enhanced in eel mutant seeds compared with wild-type seeds. Gel mobility shift assays, combined with analysis of the genetic interactions among the eel and abi5 mutations, indicated that ABI5 and EEL compete for the same binding sites within the AtEm1 promoter. This study illustrates how two homologous transcription factors can play antagonistic roles to fine-tune gene expression.

The three amino acid loop extension (TALE) homeodomain superfamily, which comprises the KNOTTED-like and BEL1-like families, plays a critical role in regulating meristem activity. We previously demonstrated a function for KNAT6 (for KNOTTED-like from Arabidopsis thaliana 6) in shoot apical meristem and boundary maintenance during embryogenesis. KNAT2, the gene most closely related to KNAT6, does not play such a role. To investigate the contribution of KNAT6 and KNAT2 to inflorescence development, we examined their interactions with two TALE genes that regulate internode patterning, BREVIPEDICELLUS (BP) and PENNYWISE (PNY). Our data revealed distinct and overlapping interactions of KNAT6 and KNAT2 during inflorescence development. Removal of KNAT6 activity suppressed the pny phenotype and partially rescued the bp phenotype. Removal of KNAT2 activity had an effect only in the absence of both BP and KNAT6 or in the absence of both BP and PNY. Consistent with this, KNAT6 and KNAT2 expression patterns were enlarged in both bp and pny mutants. Thus, the defects seen in pny and bp are attributable mainly to the misexpression of KNAT6 and to a lesser extent of KNAT2. Hence, our data showed that BP and PNY restrict KNAT6 and KNAT2 expression to promote correct inflorescence development. This interaction was also revealed in the carpel.

The homeobox gene family plays a crucial role during the development of multicellular organisms. The KNOTTED-like genes from Arabidopsis thaliana (KNAT6 and KNAT2) are close relatives of the meristematic genes SHOOT MERISTEMLESS (STM) and BREVIPEDICELLUS, but their function is not currently known. To investigate their role, we identified null alleles of KNAT6 and KNAT2. We demonstrate that KNAT6 contributes redundantly with STM to the maintenance of the shoot apical meristem (SAM) and organ separation. Consistent with this role, the expression domain of KNAT6 in the SAM marks the boundaries between the SAM and cotyledons. The lack of meristematic activity in the knat6 stm-2 double mutant and the fusion of cotyledons were linked to the modulation of CUP-SHAPED COTYLEDON (CUC) activity. During embryogenesis, KNAT6 is expressed later than STM and CUC. In agreement with this fact, CUC1 and CUC2 were redundantly required for KNAT6 expression. These data provide the basis for a model in which KNAT6 contributes to SAM maintenance and boundary establishment in the embryo via the STM/CUC pathway. KNAT2, although the closest related member of the family to KNAT6, did not have such a function.

ORCID IDs: 0000-0002-0050-4001 (B.C.S.); 0000-0002-3526-7982 (J.L.); 0000-0001-7896-6049 (M.P.); 0000-0001-7144-1274 (D.X.); 0000-0001-5026-095X (S.Ci.); 0000-0002-6496-3792 (S.R.H.); 0000-0003-1808-5172 (V.P.).In the model plant Arabidopsis (Arabidopsis thaliana), endogenous and environmental signals acting on the shoot apical meristem cause acquisition of inflorescence meristem fate. This results in changed patterns of aerial development seen as the transition from making leaves to the production of flowers separated by elongated internodes. Two related BEL1-like homeobox genes, PENNYWISE (PNY) and POUND-FOOLISH (PNF), fulfill this transition. Loss of function of these genes impairs stem cell maintenance and blocks internode elongation and flowering. We show here that pny pnf apices misexpress lateral organ boundary genes BLADE-ON-PETIOLE1/2 (BOP1/2) and KNOTTED-LIKE FROM ARABIDOPSIS THALIANA6 (KNAT6) together with ARABIDOPSIS THALIANA HOMEOBOX GENE1 (ATH1). Inactivation of genes in this module fully rescues pny pnf defects. We further show that BOP1 directly activates ATH1, whereas activation of KNAT6 is indirect. The pny pnf restoration correlates with renewed accumulation of transcripts conferring floral meristem identity, including FD, SQUAMOSA PROMOTER-BINDING PROTEIN LIKE genes, LEAFY, and APETALA1. To gain insight into how this module blocks flowering, we analyzed the transcriptome of BOP1-overexpressing plants. Our data suggest a central role for the microRNA156-SQUAMOSA PROMOTER BINDING PROTEIN-LIKE-microRNA172 module in integrating stress signals conferred in part by promotion of jasmonic acid biosynthesis. These data reveal a potential mechanism by which repression of lateral organ boundary genes by PNY-PNF is essential for flowering.Plant development relies on the activity of the shoot apical meristem (SAM) as a continuous source of founder cells for production of new leaves, shoots, and internodes throughout the life cycle (for review, see Aichinger et al., 2012). A tight balance between the allocation of cells to developing primordia and the perpetuation of pluripotent stem cells in the central zone maintains the SAM at a constant size. In Arabidopsis (Arabidopsis thaliana), the vegetative SAM produces leaves in a spiral phyllotaxy with dormant axillary meristems. In conjunction, internode elongation is repressed, resulting in a basal rosette. The transition to flowering is governed by internal and external signals that converge at the SAM to promote acquisition of inflorescence meristem (IM) fate (for review, see Amasino and Michaels, 2010;Srikanth and Schmid, 2011;Andrés and Coupland, 2012). This process, known as floral evocation, results in new patterns of growth at the shoot apex, including production of flowers, and an increase in stem elongation, called bolting. Lateral organ boundaries are specialized domains of restricted growth that separate meristem and organ compartments and produce axillary meristems (for review, see Aida and Tasaka, 2006;Tian et al., 2014). Early in the transition t...