The KNOTTED-like genes of peach (Prunus persica L. Batsch) are differentially expressed during drupe growth and the class 1 KNOPE1 contributes to mesocarp development.

The Knotted-like transcription factors (KNOX) contribute to plant organ development. The expression patterns of peach KNOX genes showed that the class 1 members act precociously (S1-S2 stages) and differentially during drupe growth. Specifically, the transcription of KNOPE1 and 6 decreased from early (cell division) to late (cell expansion) S1 sub-stages, whilst that of STMlike1, 2, KNOPE2, 2.1 ceased at early S1. The KNOPE1 role in mesocarp was further addressed by studying the mRNA localization in the pulp cells and vascular net at early and late S1. The message signal was first diffuse in parenchymatous cells and then confined to hypodermal cell layers, showing that the gene down-tuning accompanied cell expansion. As for bundles, the mRNA mainly featured in the procambium/phloem of collateral open types and subsequently in the phloem side of complex structures (converging bundles, ducts). The KNOPE1 overexpression in Arabidopsis caused fruit shortening, decrease of mesocarp cell size, diminution of vascular lignification together with the repression of the major gibberellin synthesis genes AtGA20ox1 and AtGA3ox1. Negative correlation between the expression of KNOPE1 and PpGA3ox1 was observed in four cultivars at S1, suggesting that the KNOPE1 repression of PpGA3ox1 may regulate mesocarp differentiation by acting on gibberellin homeostasis.

The peach (Prunus persica L. Batsch) genome harbours 10 KNOX genes, which are differentially expressed in stem development, and the class 1 KNOPE1 regulates elongation and lignification during primary growth.

The KNOTTED-like (KNOX) genes encode homeodomain transcription factors and regulate several processes of plant organ development. The peach (Prunus persica L. Batsch) genome was found to contain 10 KNOX members (KNOPE genes); six of them were experimentally located on the Prunus reference map and the class 1 KNOPE1 was found to link to a quantitative trait locus (QTL) for the internode length in the peach×Ferganensis population. All the KNOPE genes were differentially transcribed in the internodes of growing shoots; the KNOPE1 mRNA abundance decreased progressively from primary (elongation) to secondary growth (radial expansion). During primary growth, the KNOPE1 mRNA was localized in the cortex and in the procambium/metaphloem zones, whereas it was undetected in incipient phloem and xylem fibres. KNOPE1 overexpression in the Arabidopsis bp4 loss-of-function background (35S:KNOPE1/bp genotype) restored the rachis length, suggesting, together with the QTL association, a role for KNOPE1 in peach shoot elongation. Several lignin biosynthesis genes were up-regulated in the bp4 internodes but repressed in the 35S:KNOPE1/bp lines similarly to the wild type. Moreover, the lignin deposition pattern of the 35S:KNOPE1/bp and the wild-type internodes were the same. The KNOPE1 protein was found to recognize in vitro one of the typical KNOX DNA-binding sites that recurred in peach and Arabidopsis lignin genes. KNOPE1 expression was inversely correlated with that of lignin genes and lignin deposition along the peach shoot stems and was down-regulated in lignifying vascular tissues. These data strongly support that KNOPE1 prevents cell lignification by repressing lignin genes during peach stem primary growth.

The peach (Prunus persica [L.] Batsch) homeobox gene KNOPE3, which encodes a class 2 knotted-like transcription factor, is regulated during leaf development and triggered by sugars.

Class 1 KNOTTED1-like transcription factors (KNOX) are known to regulate plant development, whereas information on class 2 KNOX has been limited. The peach KNOPE3 gene was cloned, belonged to a family of few class 2 members and was located at 66 cM in the Prunus spp. G1 linkage-group. The mRNA localization was diversified in leaf, stem, flower and drupe, but recurred in all organ sieves, suggesting a role in sap nutrient transport. During leaf development, the mRNA earliest localized to primordia sieves and subsequently to mesophyll cells of growing leaves. Consistently, its abundance augmented with leaf expansion. The transcription was monitored in leaves responding to darkening, supply and transport block of sugars. It peaked at 4 h after darkness and dropped under prolonged obscurity, showing a similar kinetic to that of sucrose content variation. Feeding leaflets via the transpiration stream caused KNOPE3 up-regulation at 3 h after fructose, glucose and sucrose absorption and at 12 h after sorbitol. In girdling experiments, leaf KNOPE3 was triggered from 6 h onwards along with sucrose and sorbitol raise. Both the phloem-associated expression and sugar-specific gene modulation suggest that KNOPE3 may play a role in sugar translocation during the development of agro-relevant organs such as drupe.

Peach [Prunus persica (L.) Batsch] KNOPE1, a class 1 KNOX orthologue to Arabidopsis BREVIPEDICELLUS/KNAT1, is misexpressed during hyperplasia of leaf curl disease.

Class 1 KNOTTED-like (KNOX) transcription factors control cell meristematic identity. An investigation was carried out to determine whether they maintain this function in peach plants and might act in leaf curliness caused by the ascomycete Taphrina deformans. KNOPE1 function was assessed by overexpression in Arabidopsis and by yeast two-hybrid assays with Arabidopsis BELL proteins. Subsequently, KNOPE1 mRNA and zeatin localization was monitored during leaf curl disease. KNOPE1 and Arabidopsis BREVIPEDICELLUS (BP) proteins fell into the same phyletic group and recognized the same BELL factors. 35S:KNOPE1 Arabidopsis lines exhibited altered traits resembling those of BP-overexpressing lines. In peach shoot apical meristem, KNOPE1 was expressed in the peripheral and central zones but not in leaf primordia, identically to the BP expression pattern. These results strongly suggest that KNOPE1 must be down-regulated for leaf initiation and that it can control cell meristem identity equally as well as all class 1 KNOX genes. Leaves attacked by T. deformans share histological alterations with class 1 KNOX-overexpressing leaves, including cell proliferation and loss of cell differentiation. Both KNOPE1 and a cytokinin synthesis ISOPENTENYLTRANSFERASE gene were found to be up-regulated in infected curled leaves. At early disease stages, KNOPE1 was uniquely triggered in the palisade cells interacting with subepidermal mycelium, while zeatin vascular localization was unaltered compared with healthy leaves. Subsequently, when mycelium colonization and asci development occurred, both KNOPE1 and zeatin signals were scattered in sectors of cell disorders. These results suggest that KNOPE1 misexpression and de novo zeatin synthesis of host origin might participate in hyperplasia of leaf curl disease.

The gene geranylgeranyl reductase of peach (Prunus persica [L.] Batsch) is regulated during leaf development and responds differentially to distinct stress factors.

Plant geranylgeranyl hydrogenase (CHL P) reduces free geranylgeranyl diphosphate to phytil diphosphate, which provides the side chain to chlorophylls, tocopherols, and plastoquinones. In peach, the single copy gene (PpCHL P) encodes a deduced product of 51.68 kDa, which harbours a transit peptide for cytoplasm-to-chloroplast transport and a nicotinamide binding domain. The PpCHL P message was abundant in chlorophyll-containing tissues and flower organs, but barely detected in the roots and mesocarp of ripening fruits, suggesting that transcription was related to plastid types and maturation. The message was not revealed in shoot apical meristems, but spread thoroughly in leaf cells during the early stages and was located mainly in the palisade of mature leaves, which exhibited higher transcript levels than young ones. Hence, the transcription of PpCHL P was likely to be regulated during leaf development. Gene expression was monitored in leaves responding to natural dark, cold, wounding, stress by imposed darkening, and during the curl disease. Transcription was stimulated by light, but repressed by dark and cold stress. In darkened leaves, the PpCHL P message was augmented concomitantly with that of CATALASE. In wounded leaves, the message decreased, but recovered rapidly, whereas in curled leaves, a reduction in gene expression was related to leaf damage intensity. However, transcript signals increased locally both in cells mechanically wounded by a needle and in those naturally injured by the pathogenic fungus Taphrina deformans. These data suggest that PpCHL P expression was regulated by photosynthetic activity and was possibly involved in the defence response.