The plant cuticle is an important plant-atmosphere boundary, the synthesis and maintenance of which represents a significant metabolic cost. Only limited information regarding cuticle dynamics is available. We determined the composition and dynamics of Clusia rosea cuticular waxes and matrix using 13 CO2 labelling, compound-specific and bulk isotope ratio mass spectrometry. Collodion was used for wax collection; gas exchange techniques to test for any collodion effects on living leaves. Cutin matrix (MX) area density did not vary between young and mature leaves and between leaf sides. Only young leaves incorporated new carbon into their MX. Collodion-based sampling discriminated between epicuticular (EW) and intracuticular wax (IW) effectively. Epicuticular differed in composition from IW. The newly synthetised wax was deposited in IW first and later in EW. Both young and mature leaves synthetised IW and EW. The faster dynamics in young leaves were due to lower wax coverage, not a faster synthesis rate. Longer-chain alkanes were deposited preferentially on the abaxial, stomatous leaf side, producing differences between leaf sides in wax composition. We introduce a new, sensitive isotope labelling method and demonstrate that cuticular wax is renewed during leaf ontogeny of C. rosea. We discuss the ecophysiological significance of the new insights.

Department of Ecosystem Biology Faculty of Science University of South Bohemia Branišovská 1760 31 České Budějovice Czech Republic

Department of Experimental Plant Biology Faculty of Science University of South Bohemia Branišovská 1760 31 České Budějovice Czech Republic

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Baker EA. 1974. The influence of environment on leaf wax development in Brassica oleracea var. gemmifera. New Phytologist 73: 955-966.

Barthlott W, Neinhuis C. 1997. Purity of the sacred lotus, or escape from contamination in biological surfaces. Planta 202: 1-8.

Berhin A, de Belllis D, Franke RB, Andrade Buono R, Nowack M, Nawrath C. 2019. The root cap cuticle: a cell wall structure for seedling establishment and lateral root formation. Cell 176: 1367-1378.

Bernard A, Joubès J. 2013. Arabidopsis cuticular waxes: advances in synthesis, export and regulation. Progress in Lipid Research 52: 110-129.

Bird SM, Gray JE. 2003. Signals from the cuticle affect epidermal cell differentiation. New Phytologist 157: 9-23.

Bourgault R, Matschi S, Vasquez M, Qiao P, Sonntag A, Charlebois C, Mohammadi M, Scanlon MJ, Smith LG, Molina I. 2020. Constructing functional cuticles: analysis of relationships between cuticle lipid composition, ultrastructure and water barrier function in developing adult maize leaves. Annals of Botany 125: 79-91.

Boyer JS. 2015. Turgor and the transport of CO2 and water across the cuticle (epidermis) of leaves. Journal of Experimental Botany 66: 2625-2633.

Buchanan BB, Gruissem W, Jones RL, eds. 2015. Biochemistry and molecular biology of plants, 2nd edn. Chichester, UK: John Wiley & Sons.

Budke JM, Goffinet B. 2016. Comparative cuticle development reveals taller sporophytes are covered by thicker calyptra cuticles in mosses. Frontiers in Plant Science 7: 832.

Ensikat HJ, Neinhuis C, Barthlott W. 2000. Direct access to plant epicuticular wax crystals by a new mechanical isolation method. International Journal of Plant Sciences 161: 143-148.

Fernández V, Bahamonde HA, Javier Peguero-Pina J, Gil-Pelegrín E, Sancho-Knapik D, Gil L, Goldbach HE, Eichert T. 2017. Physico-chemical properties of plant cuticles and their functional and ecological significance. Journal of Experimental Botany 68: 5293-5306.

Fernández V, Guzmán-Delgado P, Graça J, Santos S, Gil L. 2016. Cuticle structure in relation to chemical composition: re-assessing the prevailing model. Frontiers in Plant Science 7: 1-14.

Fich EA, Segerson NA, Rose JKC. 2016. The plant polyester cutin: biosynthesis, structure, and biological roles. Annual Review of Plant Biology 67: 207-233.

Flore JA, Bukovac MJ. 1974. Pesticide effects on the plant cuticle: I. Response of Brassica oleracea L. to EPTC as indexed by epicuticular wax production. Journal of the American Society for Horticultural Science 99: 34-37.

Gao L, Burnier A, Huang Y. 2012. Quantifying instantaneous regeneration rates of plant leaf waxes using stable hydrogen isotope labeling. Rapid Communications in Mass Spectrometry 26: 115-122.

Gniwotta F, Vogg G, Gartmann V, Carver TLW, Riederer M, Jetter R. 2005. what do microbes encounter at the plant surface? Chemical composition of pea leaf cuticular waxes. Plant Physiology 139: 519-530.

Gustafsson MHG, Bittrich V, Stevens PF. 2002. Phylogeny of Clusiaceae based on rbc L sequences. International Journal of Plant Sciences 163: 1045-1054.

Guzmán P, Fernández V, García ML, Khayet M, Fernández A, Gil L. 2014. Localization of polysaccharides in isolated and intact cuticles of eucalypt, poplar and pear leaves by enzyme-gold labelling. Plant Physiology and Biochemistry 76: 1-6.

Haas K, Rentschler I. 1984. Discrimination between epicuticular and intracuticular wax in blackberry leaves: ultrastructural and chemical evidence. Plant Science Letters 36: 143-147.

Hauke V, Schreiber L. 1998. Ontogenetic and seasonal development of wax composition and cuticular transpiration of ivy (Hedera helix L.) sun and shade leaves. Planta 207: 67-75.

Holloway PJ, Hunt GM, Baker EA, Macey MJK. 1977. Chemical composition and ultrastructure of the epicuticular wax in four mutants of Pisum sativum (L). Chemistry and Physics of Lipids 20: 141-155.

Jeffree CE. 2018. The fine structure of the plant cuticle. In: Riederer M, Müller C, eds. Annual plant reviews online, vol. 23. Chichester, UK: John Wiley & Sons, 11-125.

Jetter R, Kunst L, Samuels AL. 2018. Composition of plant cuticular waxes. In: Annual plant reviews online. Chichester, UK: John Wiley & Sons, 145-181.

Jetter R, Schäffer S. 2001. Chemical composition of the Prunus laurocerasus leaf surface. Dynamic changes of the epicuticular wax film during leaf development. Plant Physiology 126: 1725-1737.

Jetter R, Schäffer S, Riederer M. 2000. Leaf cuticular waxes are arranged in chemically and mechanically distinct layers: evidence from Prunus laurocerasus L. Plant, Cell & Environment 23: 619-628.

Kahmen A, Dawson TE, Vieth A, Sachse D. 2011. Leaf wax n-alkane δD values are determined early in the ontogeny of Populus trichocarpa leaves when grown under controlled environmental conditions. Plant, Cell & Environment 34: 1639-1651.

Karabourniotis G, Tzobanoglou D, Nikolopoulos D, Liakopoulos G. 2001. Epicuticular phenolics over guard cells: exploitation for in situ stomatal counting by fluorescence microscopy and combined image analysis. Annals of Botany 87: 631-639.

Kerstiens G. 1996a. Cuticular water permeability and its physiological significance. Journal of Experimental Botany 47: 1813-1832.

Kerstiens G. 1996b. Signalling across the divide: a wider perspective of cuticular structure-function relationships. Trends in Plant Science 1: 125-129.

Kerstiens G. 1997. In vivo manipulation of cuticular water permeance and its effect on stomatal response to air humidity. New Phytologist 137: 473-480.

Koch K, Neinhuis C, Ensikat H, Barthlott W. 2004. Self assembly of epicuticular waxes on living plant surfaces imaged by atomic force microscopy (AFM). Journal of Experimental Botany 55: 711-718.

Kong L, Liu Y, Zhi P, Wang X, Xu B, Gong Z, Chang C. 2020. Origins and evolution of cuticle biosynthetic machinery in land plants. Plant Physiology 184: 1998-2010.

Kubásek J, Hájek T, Duckett J, Pressel S, Šantrůček J. 2021. Moss stomata do not respond to light and CO2 concentration but facilitate carbon uptake by sporophytes: a gas exchange, stomatal aperture and 13C labelling study. New Phytologist 230: 1815-1828.

Lewandowska M, Keyl A, Feussner I. 2020. Wax biosynthesis in response to danger: its regulation upon abiotic and biotic stress. New Phytologist 227: 698-713.

Lüttge U. 2008. Clusia: holy grail and enigma. Journal of Experimental Botany 59: 1503-1514.

Macková J, Vašková M, Macek P, Hronková M, Schreiber L, Šantrůček J. 2013. Plant response to drought stress simulated by ABA application: changes in chemical composition of cuticular waxes. Environmental and Experimental Botany 86: 70-75.

Matos TM, Peralta DF, Roma LP, dos Santos DYAC. 2021. The morphology and chemical composition of cuticular waxes in some Brazilian liverworts and mosses. Journal of Bryology 43: 129-137.

Medina E, Aguiar G, Gómez M, Aranda J, Medina JD, Winter K. 2006. Taxonomic significance of the epicuticular wax composition in species of the genus Clusia from Panama. Biochemical Systematics and Ecology 34: 319-326.

Medina E, Aguiar G, Gómez M, Medina JD. 2004. Patterns of leaf epicuticular waxes in species of clusia: taxonomical implications. Interciencia 29: 579-582.

Neinhuis C, Koch K, Barthlott W. 2001. Movement and regeneration of epicuticular waxes through plant cuticles. Planta 213: 427-434.

Niklas KJ, Cobb ED, Matas AJ. 2017. The evolution of hydrophobic cell wall biopolymers: from algae to angiosperms. Journal of Experimental Botany 68: 5261-5269.

Onoda Y, Richards L, Westoby M. 2012. The importance of leaf cuticle for carbon economy and mechanical strength. New Phytologist 196: 441-447.

Panikashvili D, Shi JX, Schreiber L, Aharoni A. 2011. The Arabidopsis ABCG13 transporter is required for flower cuticle secretion and patterning of the petal epidermis. New Phytologist 190: 113-124.

Petit J, Bres C, Mauxion J-P, Bakan B, Rothan C. 2017. Breeding for cuticle-associated traits in crop species: traits, targets, and strategies. Journal of Experimental Botany 68: 5369-5387.

Pighin JA, Zheng H, Balakshin LJ, Goodman IP, Western TL, Jetter R, Kunst L, Samuels AL. 2004. Plant cuticular lipid export requires an ABC transporter. Science 306: 702-704.

Proctor MCF, Tuba Z. 2002. Poikilohydry and homoihydry: antithesis or spectrum of possibilities? New Phytologist 156: 327-349.

Riederer M, Müller C. 2006. Biology of the plant cuticle. Oxford, UK: Blackwell.

Riederer M, Schreiber L. 2001. Protecting against water loss: analysis of the barrier properties of plant cuticles. Journal of Experimental Botany 52: 2023-2032.

Sachse D, Kahmen A, Gleixner G. 2009. Significant seasonal variation in the hydrogen isotopic composition of leaf-wax lipids for two deciduous tree ecosystems (Fagus sylvativa and Acer pseudoplatanus). Organic Geochemistry 40: 732-742.

Samuels L, Kunst L, Jetter R. 2008. Sealing plant surfaces: cuticular wax formation by epidermal cells. Annual Review of Plant Biology 59: 683-707.

Schimmelmann A, Qi H, Coplen TB, Brand WA, Fong J, Meier-Augenstein W, Kemp HF, Toman B, Ackermann A, Assonov S et al. 2016. Organic reference materials for hydrogen, carbon, and nitrogen stable isotope-ratio measurements: caffeines, n -alkanes, fatty acid methyl esters, glycines, l-valines, polyethylenes, and oils. Analytical Chemistry 88: 4294-4302.

Schönherr J, Riederer M. 1986. Plant cuticles sorb lipophilic compounds during enzymatic isolation. Plant, Cell & Environment 9: 459-466.

Schönherr J, Schmidt HW. 1979. Water permeability of plant cuticles: dependence of permeability coefficients of cuticular transpiration on vapor pressure saturation deficit. Planta 144: 391-400.

Schreiber L, Skrabs M, Hartmann KD, Diamantopoulos P, Simanova E, Santrucek J. 2001. Effect of humidity on cuticular water permeability of isolated cuticular membranes and leaf disks. Planta 214: 274-282.

Seale M. 2020. The fat of the land: cuticle formation in terrestrial plants. Plant Physiology 184: 1622-1624.

Skoss JD. 1955. Structure and composition of plant cuticle in relation to environmental factors and permeability. Botanical Gazette 117: 55-72.

Terashima I, Hanba YT, Tholen D, Niinemets Ü. 2011. Leaf functional anatomy in relation to photosynthesis. Plant Physiology 155: 108-116.

Tsubaki S, Ozaki Y, Yonemori K, Azuma J. 2012. Mechanical properties of fruit-cuticular membranes isolated from 27 cultivars of Diospyros kaki Thunb. Food Chemistry 132: 2135-2139.

Vráblová M, Vrábl D, Sokolová B, Marková D, Hronková M. 2020. A modified method for enzymatic isolation of and subsequent wax extraction from Arabidopsis thaliana leaf cuticle. Plant Methods 16: 129.

Wang X, Kong L, Zhi P, Chang C. 2020. Update on cuticular wax biosynthesis and its roles in plant disease resistance. International Journal of Molecular Sciences 21: 5514.

Wen M, Jetter R. 2009. Composition of secondary alcohols, ketones, alkanediols, and ketols in Arabidopsis thaliana cuticular waxes. Journal of Experimental Botany 60: 1811-1821.

Woolley JT. 1967. Relative permeabilities of plastic films to water and carbon dioxide. Plant Physiology 42: 641-643.

Yeats TH, Buda GJ, Wang Z, Chehanovsky N, Moyle LC, Jetter R, Schaffer AA, Rose JKC. 2012. The fruit cuticles of wild tomato species exhibit architectural and chemical diversity, providing a new model for studying the evolution of cuticle function. The Plant Journal 69: 655-666.

Zeisler V, Schreiber L. 2016. Epicuticular wax on cherry laurel (Prunus laurocerasus) leaves does not constitute the cuticular transpiration barrier. Planta 243: 65-81.

Zeisler-Diehl V, Migdal B, Schreiber L. 2017. Quantitative characterization of cuticular barrier properties: methods, requirements, and problems. Journal of Experimental Botany 68: 5281-5291.

Zeisler-Diehl V, Müller Y. 2018. Epicuticular wax on leaf cuticles does not establish the transpiration barrier, which is essentially formed by intracuticular wax. Journal of Plant Physiology 227: 66-74.

Amphistomy: stomata patterning inferred from 13C content and leaf-side-specific deposition of epicuticular wax