The development of plastids in the subepidermis of fruits of Cucurbita pepo var. patisson starts with the stage of normal proplastids. In the process of fruit ripening the proplastids are transformed into fully-grown leucoplasts, which are similar in ultrastructure to the chromoplasts of the globular type. In the process of fruit decaying the leucoplasts are completely disintegrated. The problem of plastid terminology has been touched upon.Razvoj plastida u subepidermi plodova varijeteta Cucurbita pepo var. pâtisson započinje sa stadijem normalno građenih proplastida. Sazrijevanjem ploda proplastidi prelaze u odrasle bezbojne plastide — leukoplaste, čija fina građa odgovara onoj globularnog tipa kromoplasta. Starenjem ploda u leukoplastima se razgrade sve membranske tvorevine i dolazi do potpune dezintegracije plastida. Istaknute su teškoće terminologije u tom slučaju

Nikola Ljubešić

Razvoj plastida u subepidermi plodova varijeteta Cucurbita pepo var. pâtisson započinje sa stadijem normalno građenih proplastida. Sazrijevanjem ploda proplastidi prelaze u odrasle bezbojne plastide — leukoplaste, čija fina građa odgovara onoj globularnog tipa kromoplasta. Starenjem ploda u leukoplastima se razgrade sve membranske tvorevine i dolazi do potpune dezintegracije plastida. Istaknute su teškoće terminologije u tom slučaju.The development of plastids in the subepidermis of fruits of Cucurbita pepo var. patisson starts with the stage of normal proplastids. In the process of fruit ripening the proplastids are transformed into fully-grown leucoplasts, which are similar in ultrastructure to the chromoplasts of the globular type. In the process of fruit decaying the leucoplasts are completely disintegrated. The problem of plastid terminology has been touched upon

Ljubešić, Nikola

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Acta Bot. Croat. 32 (1973) 59— 62T R A N S F O R M A T I O N S  O F P L A S T I D S  I N W H I T E  P U M P K I N  F R U I T SNIKOLA LJUBESlC(Laboratory o f  E lectron M icroscopy, R uder B oskovic Institute, Zagreb) R eceived  January 24, 1973I n t r o d u c t i o nThe transformation of proplastids into leucoplasts occurs very fre­quently in the underground parts of the plant. The proplastids might have been transformed into chloroplasts but the light was missing. If these parts were exposed to light, the leucoplasts would synthesize chlorophyll and become chloroplasts (De R e z e n d e - P i n t o  1962). Colourless plastids (leucoplasts) occur also in the parts of the plant which are exposed to the sun light but have a genetic defect or are otherwise prevented from synthesizing chlorophyll ( K i r k  and T i l n e y - B a s -  s e t t  1967).The majority of the fruits of pumpkin species are pigmented, and contain many types of coloured plastids ( D é v i d é  1970a, 1970b, L j u -  b e s i c  1970a, 1970b, 1972). However, in the stage of ripening and withering of Cucurbita pepo var. pâtisson fruit, colourless plastids are present showing the transformations of proplastids into leucoplasts.M a t e r i a l  and  M e t h o d sThe plastids of the outer cell layers (subepidermis) of the Cucurbita pepo var. pâtisson fruits were investigated. The plants were grown in garden conditions. The samples for investigation were taken at deter­mined intervals, i.e. immediately after the flowering, during the ripening and before decaying of the fruit.The material was investigated by light and electron microscopy. For electron microscopy the material was fixed in l°/o glutaraldehyde, postfixed in 1%> 0s04 and embedded in araldite. The sections were made on Reichert Om U2 ultramicrotome, stained with uranyl acetate and lead citrate and observed in a Siemens Elmiskop I (at the Institute of Biology, University of Zagreb).59The tissue samples taken for microscopy were also tested for pig­ments. The pigment compounds were examined by means of thin-layer chromatography on silica gel G ( B o l l i g e r  and K o n i g  1969; solvent: petrol ether : ethyl acetate : diethylamine — 58 : 30 : 12).Re s ul t s  and Di s c us s i o nThe transformations of plastids in subepidermal cell layers of Cu­curbita pepo var. pâtisson may be divided into three periods:1. T h e  P e r i o d  o f  F r u i t  D e v e l o p m e n t .  This period begins with flowering and ends with the process of fruit ripening. During this period the fruits are of a pale-green colour. Thin-layer chromato­graphy revealed small amounts of chlorophyll and carotenoids. However, since during the process of growing the fruits enlarge their volume considerably, and new amounts of pigments are not synthesized, the quantity of pigment per unit of fresh weight is diminished. In this period very small and oval plastids can be observed in the light micro­scope. Their colour is hardly greenish. This greenish colour is possibly a result of diffraction of light on the plastids, and not of presence of chlorophyll. No fluorescence could be noticed in the fluorescence micro­scope. The plastids show no visible structures. With the ripening of the fruits their volume increases, but the number of plastids per cell and their size do not change.The presence of a small amount of chlorophyll in young fruits is evident but no chloroplasts could be observed by the electron microscope. In the subepidermis only typical proplastids could be found (Fig. 1). The plastids are oval in shape and about 2 pm in diameter. The stroma is dense and rich with ribosomes. Only a few long single thylakoids------------------------ ---------------------------------------------------------------------------►Figs. 1. —  4. Cucurbita pepo var. pâtisson. Plastids from the outer cell layers of the fruit:SI. 1. —  4. Cucurbita pepo var. pâtisson. Plastidi iz subepidermskog sloja ploda:Fig. 1. Proplastids from young fruits. Big starch grains are present. 40,000 :1. Si. 1. Proplastidi iz mladog ploda. Prisutna su velika škrobna zrnca. 40 000 :1.Fig. 2. Leucoplast from the fruit at the beginning of ripening period. Pla- stoglobules are surrounded by a few long thylakoids. 30,000 :1.SI. 2. Leukoplast iz ploda na početku perioda zriobe. Oko nakupine plasto- globula nalaze se dugi tilakoidi. 30 000 :1.Fig. 3. Leucoplast from the ripe fruit. In the central part of the leucoplast there is a large accumulation of plastoglobules. 30,000 :1.SI. 3. Leukoplast zrelog ploda. U srednjem dijelu leukoplasta je velika na­kupina plastoglobula. 30 000 :1.Fig. 4. Plastid (leucoplast) from the fruit during the decaying period. Only remnants of thylakoids are present. 32,000 :1.SI. 4. Plastid (leukoplast) iz perioda propadanja ploda. Prisutni su samo ostaci tilakoida. 32 000 :1.60Fig. 1— 2. SL 1— 2.Fig. 3— 4, SI. 3— 4.are present. Grana have never been found. Usually a big grain of starch is situated in the middle of the plastid. Such plastids are similar to amyloplasts. The plastoglobules are small and scarce.2. T h e  P e r i o d  o f  R i p e  F r u i t .  Ripe fruits are white. It was not possible to prove the presence of chlorophyll by thin-layer chromatography and UV-light. A small amount of carotenoids was found. No changes could be observed in the light microscope- These oval plastids, of about 2 pm diameter, are colourless, and they exist in mature tissue and could be called leucoplasts.In the electron microscope these plastids (Figs. 2, 3) show a dense stroma rich with ribosomes. The central part of the plastids is filled with numerous plastoglobules (about 100 nm in diameter). The accumulat­ed plastoglobules enlarge in volume parallelly with the ripening of the fruit (Fig. 3). The plastoglobules are surrounded by a few long single thylakoids (Figs. 2, 3). Grana or grana-like structures are not present. The majority of thylakoids is perforated. The thylakoids are dilated and form osmiophilic spots. Along the border of the plastids numerous tubules occur. In fact these tubules are the invaginations of the inner part of the plastid membrane.There are some difficulties in respect of terminology. The ultra­structure of these leucoplasts is identical with that of chromoplasts of the globular type ( S t e f f e n  and W a l t e r  1958). In these leucoplasts numerous plastoglobules have a very low osmiophily. According to L i c h t e n t h a l e r  (1970) the low osmiophily of plastoglobules in­dicates an increased amount of carotenoids. However, these leucoplasts contain only traces of pigments and in spite of their ultrastructural similarity to chromoplasts, they are typical leucoplasts. The presence or absence of the pigments still seems to be a more important criterion for classification of plastids than morphological similarity or dissimilarity.3. T h e  D e c a y i n g  P e r i o d  o f  t h e  F r u i t .  Signs of sene­scence appear in the fruits five months after ripening, and one month later the fruit decays. An increase in the volume of leucoplasts can be observed in the light microscope. The disintegration of the thylakoidal system and the appearance of numerous plastoglobules are visible under the electron microscope (Fig. 4). The remnants of thylakoids in the form of smaller or bigger vesicles are present among the plastoglo­bules. The composition of pigments is the same as in a ripe fruit. In spite of the evident signs of fruit degeneration (disintegration of the thylakoids), ribosomes are visible in the leucoplast stroma of normal density.In the last stage of senescence the leucoplasts disintegrate completely. The disintegration of these leucoplasts is identical with that in chro­moplasts of other pumpkin varieties ( L j u b e s i c  1970a, 1972).*The author is indebted to Prof. Dr. Z. Dévidé and Dr. M. Wrischer, for reading the manuscript and helpful discussion.S u m m a r yThe development of plastids in the subepidermis of fruits of Cu- curbita pepo var. pâtisson starts with the stage of normal proplastids. In the process of fruit ripening the proplastids are transformed into61fully-grown leucoplasts, which are similar in ultrastructure to the chromoplasts of the globular type. In the process of fruit decaying the leucoplasts are completely disintegrated. The problem of plastid termino­logy has been touched upon.R e f e r e n c e sBolliger, H. and A. König, 1969: Vitamins, including carotenoids, chlorophylls and biologically active quinones. In: Thin-layer chromatography (Edi­tor: E. Stahl), Springer Verlag, Berlin —  Heidelberg —  New York.De Rezende-Pinto, M. C., 1962: Metamorphosis of amyloplasts of Solanum tuberosum into amylochloroplasts induced by light. Portug. Acta Biol. 6A, 239— 242.Dévidé, Z., 1970a: Ultrastructural changes of plastids in ripe fruit of Cu­curbita pepo cv. ovifera. Acta. Bot. Croat. 29, 57— 62.Dévidé, Z„ 1970b: Changes in fine structure of plastids in ripe fruit of Cu­curbita pepo cv. ovifera. Microscopie Électronique, Vol. Ill, 201— 202. 7-ième Congr. Intern. Micr. Électronique, Grenoble.Kirk, J. T. O. and R. A. E. Tilney-Bassett, 1967: The plastids. W. H. Freeman and Company, London —  San Francisco.Lichtenthaler, H. K., 1970: Die Lokalisation der Plastochinone und Carotinoide in den Chromopiasten der Petalen von Sarothamnus scoparius (L.) Wimm ex Koch. Planta 90, 142— 152.Ljubesic, N., 1970a: Fine structure of developing chromoplasts in outer yellow parts of Cucurbita pepo cv. pyriformis. Acta Bot. Croat. 29, 51— 56.Ljubesic, N., 1970b: Ultrastructural changes in chromoplasts of the fruit of Cucurbita pepo cv. pyriformis. Microscopie Électronique, Vol. Ill, 203—  — 204. 7-ième Congr. Intern. Micr. Électronique, Grenoble.Ljubesic, N., 1972: Ultrastructural changes of plastids during the yellowing of the fruit of Cucurbita pepo var. pyriformis. Acta Bot. Croat. 31, 47— 53.Steffen, K . und F. Walter, 1958: Die Chromopiasten von Solanum capsicastrum L. und ihre Genese. Planta 50, 640— 670.S A D R Ž A JPRETVORBE P LA STID A  U BIJELIM  PLODOVIM A BUNDEVENikola Ljubešić(Laboratorij za elektronsku m ikroskopiju , Institut »Ruđer B oškovlć«, Zagreb)Razvoj plastida u subepidermi plodova varijeteta Cucurbita pepo var. pâtisson započinje sa stadijem normalno građenih proplastida. Sa­zrijevanjem ploda proplastidi prelaze u odrasle bezbojne plastide — leukoplaste, čija fina građa odgovara onoj globularnog tipa kromoplasta. Starenjem ploda u leukoplastima se razgrade sve membranske tvorevine i dolazi do potpune dezintegracije plastida. Istaknute su teškoće termino­logije u tom slučaju.D r N ikola L ju bešić Institut »Ruđer B oškovlć«B ijen ička  5441 000 Zagreb (Jugoslavija)62

Pretvorbe plastida u bijelim plodovima bundeve

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