The Petunia CHANEL Gene is a ZEITLUPE Ortholog Coordinating Growth and Scent Profiles

The floral perianth, comprising sepals and petals, conceals the sexual organs and attracts pollinators. The coordination of growth and scent emission is not fully understood. We have analyzed the effect of knocking down CHANEL (PhCHL), the ZEITLUPE ortholog in petunia (PhCHL) by hairpin RNAs. Plants with low PhCHL mRNA had overall decreased size. Growth evaluation using time lapse image analysis showed that early leaf movement was not affected by RNAi:PhCHL, but flower angle movement was modified, moving earlier during the day in knockdown plants than in wild types. Despite differences in stem length, growth rate was not significantly affected by loss of PhCHL. In contrast, petal growth displayed lower growth rate in RNAi:PhCHL. Decreased levels of PhCHL caused strongly modified scent profiles, including changes in composition and timing of emission resulting in volatile profiles highly divergent from the wild type. Our results show a role of PhCHL in controlling growth and development of vegetative and reproductive organs in petunia. The different effects of PhCHL on organ development indicate an organ-specific interpretation of the down regulation of PhCHL. Through the control of both timing and quantitative volatile emissions, PhCHL appears to be a major coordinator of scent profiles.

[1]  T. Masci,et al.  Diurnal regulation of scent emission in rose flowers , 2007, Planta.

[2]  Klaus Schliep,et al.  Phylogenetic Analysis in R , 2015 .

[3]  Xiaomin Wei,et al.  Cloning and Characterization of a Putative TAC1 Ortholog Associated with Leaf Angle in Maize (Zea mays L.) , 2011, PloS one.

[4]  P. Waterhouse,et al.  Constructs and methods for high-throughput gene silencing in plants. , 2003, Methods.

[5]  Caroline Dean,et al.  Protocol: A simple phenol-based method for 96-well extraction of high quality RNA from Arabidopsis , 2011, Plant Methods.

[6]  D. Inzé,et al.  Growth rate rather than growth duration drives growth heterosis in maize B104 hybrids , 2017, Plant, Cell and Environment.

[7]  D. E. Somers,et al.  Control of circadian rhythms and photoperiodic flowering by the Arabidopsis GIGANTEA gene. , 1999, Science.

[8]  M. Lenhard,et al.  Regulation of plant lateral-organ growth by modulating cell number and size. , 2014, Current opinion in plant biology.

[9]  I. Baldwin,et al.  Fitness consequences of altering floral circadian oscillations for Nicotiana attenuata. , 2017, Journal of integrative plant biology.

[10]  Baohong Zhang,et al.  miRDeepFinder: a miRNA analysis tool for deep sequencing of plant small RNAs , 2012, Plant Molecular Biology.

[11]  Thomas D. Schmittgen,et al.  Analyzing real-time PCR data by the comparative CT method , 2008, Nature Protocols.

[12]  M. Egea-Cortines,et al.  Quantitative levels of Deficiens and Globosa during late petal development show a complex transcriptional network topology of B function. , 2012, The Plant journal : for cell and molecular biology.

[13]  I. Baldwin,et al.  Identification and characterization of circadian clock genes in a native tobacco, Nicotiana attenuata , 2012, BMC Plant Biology.

[14]  N. Blüthgen,et al.  Floral scents repel potentially nectar-thieving ants , 2008 .

[15]  M. Barton,et al.  Twenty years on: the inner workings of the shoot apical meristem, a developmental dynamo. , 2010, Developmental biology.

[16]  Gerrit T.S. Beemster,et al.  Variation in Growth Rate between Arabidopsis Ecotypes Is Correlated with Cell Division and A-Type Cyclin-Dependent Kinase Activity1 , 2002, Plant Physiology.

[17]  A. Millar,et al.  The Circadian Clock That Controls Gene Expression in Arabidopsis Is Tissue Specific1 , 2002, Plant Physiology.

[18]  A. Millar,et al.  Organ specificity in the plant circadian system is explained by different light inputs to the shoot and root clocks , 2016, The New phytologist.

[19]  A. Aharoni,et al.  Terpenoid Metabolism in Wild-Type and Transgenic Arabidopsis Plants Online version contains Web-only data. Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.016253. , 2003, The Plant Cell Online.

[20]  A comparison of semi-quantitative methods suitable for establishing volatile profiles , 2018, Plant Methods.

[21]  B. Krizek.  Ectopic expression of AINTEGUMENTA in Arabidopsis plants results in increased growth of floral organs. , 1999, Developmental genetics.

[22]  Thomas Altmann,et al.  Versatile gene-specific sequence tags for Arabidopsis functional genomics: transcript profiling and reverse genetics applications. , 2004, Genome research.

[23]  Motomu Endo.  Tissue-specific circadian clocks in plants. , 2016, Current opinion in plant biology.

[24]  Richard Kennaway,et al.  Generation of Diverse Biological Forms through Combinatorial Interactions between Tissue Polarity and Growth , 2011, PLoS Comput. Biol..

[25]  Pedro J. Navarro,et al.  Machine Learning and Computer Vision System for Phenotype Data Acquisition and Analysis in Plants , 2016, Sensors.

[26]  David K. Smith,et al.  ggtree: an r package for visualization and annotation of phylogenetic trees with their covariates and other associated data , 2017 .

[27]  Robert C Schuurink,et al.  Regulation of floral scent production in petunia revealed by targeted metabolomics. , 2003, Phytochemistry.

[28]  Pedro M. Valero-Mora,et al.  ggplot2: Elegant Graphics for Data Analysis , 2010 .

[29]  D. E. Somers,et al.  ZEITLUPE Encodes a Novel Clock-Associated PAS Protein from Arabidopsis , 2000, Cell.

[30]  N. Dudareva,et al.  Developmental Regulation of Methyl Benzoate Biosynthesis and Emission in Snapdragon Flowers , 2000, Plant Cell.

[31]  W. Kao,et al.  Tropic leaf movements, photosynthetic gas exchange, leaf δ13C and chlorophyll a fluorescence of three soybean species in response to water availability , 1998 .

[32]  Eran Pichersky,et al.  The formation and function of plant volatiles: perfumes for pollinator attraction and defense. , 2002, Current opinion in plant biology.

[33]  Carlos Fernández Andrés,et al.  Development of a Configurable Growth Chamber with a Computer Vision System to Study Circadian Rhythm in Plants , 2012, Sensors.

[34]  E. M. Farré,et al.  The regulation of plant growth by the circadian clock. , 2012, Plant biology.

[35]  F. Speleman,et al.  Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes , 2002, Genome Biology.

[36]  R. Durbin,et al.  GeneWise and Genomewise. , 2004, Genome research.

[37]  T. Stuessy,et al.  A survey of floral traits, breeding systems, floral visitors, and pollination systems of the angiosperms of the Juan Fernández Islands (Chile) , 2001, The Botanical Review.

[38]  K. Yamato,et al.  Co-option of a photoperiodic growth-phase transition system during land plant evolution , 2014, Nature Communications.

[39]  Korbinian Strimmer,et al.  APE: Analyses of Phylogenetics and Evolution in R language , 2004, Bioinform..

[40]  Ajay Jain,et al.  Aphid-repellent pheromone E-β-farnesene is generated in transgenic Arabidopsis thaliana over-expressing farnesyl diphosphate synthase2. , 2015, Annals of botany.

[41]  Klaus Peter Schliep,et al.  phangorn: phylogenetic analysis in R , 2010, Bioinform..

[42]  J. Gershenzon,et al.  Protective perfumes: the role of vegetative volatiles in plant defense against herbivores. , 2009, Current opinion in plant biology.

[43]  Hans Sommer,et al.  Ternary complex formation between the MADS‐box proteins SQUAMOSA, DEFICIENS and GLOBOSA is involved in the control of floral architecture in Antirrhinum majus , 1999, The EMBO journal.

[44]  Rémy Bruggmann,et al.  Insight into the evolution of the Solanaceae from the parental genomes of Petunia hybrida , 2016, Nature Plants.

[45]  M. Pfaffl,et al.  Determination of stable housekeeping genes, differentially regulated target genes and sample integrity: BestKeeper – Excel-based tool using pair-wise correlations , 2004, Biotechnology Letters.

[46]  Michael H. Wilson,et al.  The circadian clock rephases during lateral root organ initiation in Arabidopsis thaliana , 2015, Nature Communications.

[47]  B. Kuluev,et al.  Role of AINTEGUMENTA-like gene NtANTL in the regulation of tobacco organ growth. , 2015, Journal of plant physiology.

[48]  J. Ludwig,et al.  grofit: Fitting Biological Growth Curves with R , 2010 .

[49]  Andrew J. Millar,et al.  The ELF4 gene controls circadian rhythms and flowering time in Arabidopsis thaliana , 2002, Nature.

[50]  Diego J. Aguila,et al.  Optimization of fragrance extraction: Daytime and flower age affect scent emission in simple and double narcissi , 2014 .

[51]  Karl Kornacker,et al.  JTK_CYCLE: An Efficient Nonparametric Algorithm for Detecting Rhythmic Components in Genome-Scale Data Sets , 2010, Journal of biological rhythms.

[52]  Juan A. Fernández,et al.  Diel pattern of circadian clock and storage protein gene expression in leaves and during seed filling in cowpea (Vigna unguiculata) , 2018, BMC Plant Biology.

[53]  Izaskun Mallona,et al.  pcrEfficiency: a Web tool for PCR amplification efficiency prediction , 2011, BMC Bioinformatics.

[54]  Kerstin Kaufmann,et al.  The 'ABC' of MADS domain protein behaviour and interactions. , 2010, Seminars in cell & developmental biology.

[55]  Gang Wu,et al.  MetaCycle: an integrated R package to evaluate periodicity in large scale data , 2016, bioRxiv.

[56]  Koji Goto,et al.  Complexes of MADS-box proteins are sufficient to convert leaves into floral organs , 2001, Nature.

[57]  Claus Lindbjerg Andersen,et al.  Normalization of Real-Time Quantitative Reverse Transcription-PCR Data: A Model-Based Variance Estimation Approach to Identify Genes Suited for Normalization, Applied to Bladder and Colon Cancer Data Sets , 2004, Cancer Research.

[58]  S. Thein,et al.  Selection of housekeeping genes for gene expression studies in human reticulocytes using real-time PCR , 2006, BMC Molecular Biology.

[59]  C. Kuhlemeier,et al.  Gain and Loss of Floral Scent Production through Changes in Structural Genes during Pollinator-Mediated Speciation , 2016, Current Biology.

[60]  Rodrigo Lopez,et al.  Clustal W and Clustal X version 2.0 , 2007, Bioinform..

[61]  Paloma Mas,et al.  The Circadian Clock Sets the Time of DNA Replication Licensing to Regulate Growth in Arabidopsis. , 2018, Developmental cell.

[62]  T. Imaizumi,et al.  Circadian clock gene LATE ELONGATED HYPOCOTYL directly regulates the timing of floral scent emission in Petunia , 2015, Proceedings of the National Academy of Sciences.

[63]  M. Egea-Cortines,et al.  Validation of Aintegumenta as a gene to modify floral size in ornamental plants. , 2014, Plant biotechnology journal.

[64]  I. Baldwin,et al.  Silencing Nicotiana attenuata LHY and ZTL alters circadian rhythms in flowers. , 2016, The New phytologist.

[65]  T. Imaizumi,et al.  Circadian Rhythms in Floral Scent Emission , 2016, Front. Plant Sci..

[66]  O. Vitek,et al.  Developmental Changes in the Metabolic Network of Snapdragon Flowers , 2012, PloS one.

[67]  C. Kuhlemeier,et al.  The composition and timing of flower odour emission by wild Petunia axillaris coincide with the antennal perception and nocturnal activity of the pollinator Manduca sexta , 2005, Planta.

[68]  Marcos Egea-Cortines,et al.  Validation of reference genes for quantitative real-time PCR during leaf and flower development in Petunia hybrida , 2010, BMC Plant Biology.

[69]  Steve A. Kay,et al.  The ELF4-ELF3-LUX Complex Links the Circadian Clock to Diurnal Control of Hypocotyl Growth , 2011, Nature.