CHROMOPLASTS OF TOMATO FRUITS. II. THE RED TOMATO

Plastid transformation at the ultrastructural level in a normal red tomato 'Pearson' is reported. During ripening, lycopene aggregates on thylakoid membranes. These thylakoids lengthen and sometimes are associated in concentrically layered groups forming very elongated or irregular crystalloids. Thylakoid plexes often occur. Comparisons of the mature chromoplasts of the normal red tomato with those of other pigment lines indicate that differences in carotenoids markedly affect ultrastructure. IN THE first part of this study (Harris and Spurr, 1969), carotene analyses during ripening of lowpigment (r/r), high-beta (B/B), and normal red (r+/r+) lines of 'Pearson' tomato and the ultrastructural transformations of plastids of the first two lines were reported. In this second report the fine structural development of chromoplasts in the normal red tomato, where lycopene is the predominant carotene, is described. Materials and methods and the ripeness stages were described in the preceding report. Light-microscope studies have shown that when lycopene is present in large amounts characteristic crystalline bodies are present. Duggar (1913) noted that red tomatoes with large amounts of "lycopersicin" had long needle-shaped pigment bodies. Howard (1925) observed a similar situation in other fruits containing lycopene. Purcell, Carra, and DeGruy (1963) noted needle-shaped pigment bodies in the chromoplasts of pink grapefruit. However, as beta-carotene increased, pigment crystals with a more rectangular "platelet" form increased in number. Oztig (1962) noted that red tomatoes contain needle-shaped pigment bodies as well as roundish chromoplasts. Grilli (1965a, b, c) studied several types of chromoplasts, including those in the tomato, with the electron microscope. Elongated crystalloids and membrane systems were evident in her material, but the study did not include developmental stages. The present studv was undertaken to determine: (1) the development of tomato chromoplasts at the fine structural level, (2) the relationship between lycopene and chromoplast structure, and (3) the effect of genes controlling fruit pig1 Received for publication 31 May 1968. This study is part of a thesis submitted by the first author in partial fulfillment of the requirements for the degree of Doctor of Philosophy, University of California. The authors thank Miss Dora G. Hunt for reading the manuscript. 2 Present address: Department of Botany and Bacteriology, University of Arkansas, Fayetteville, Arkansas 72701. ment on chromoplast ultrastructure and carotene development. OBSERVATIONS-The chloroplasts of the mature green fruits of the normal red tomato (Fig. 1) were similar to those reported for most higher plants. Starch was commonly present at this stage and the cells were highly vacuolated. As the fruits began to ripen globules increased in the chloroplasts and frequently a dark granular body appeared (Fig. 2). The first indication that a chloroplast was changing into a chromoplast was the appearance of electron-dense lines associated with granal and intergranal membranes (Fig. 3-5). These dark lines have been interpreted as early carotenoid depositions (Rosso, in press) on the basis of theoretical affinities of oxidizing agents for carotenes and polarizing light studies. In our own observations of fresh material under the light microscope the size of the red crystals corresponded to the dark-staining areas seen in electron micrographs. Pure lycopene crystals have been shown to reduce 0S04 vapors on filter paper (Harris and Spurr, 1969). Thus these electron-dense lines were interpreted as carotene crystalloids. With glutaraldehyde-osmium fixation procedures the pigment crystalloid region assumed a wavy or sinuous outline (Fig. 3, 4, 6). The electron-transparent regions adjacent to the wavy lines probably represented areas where the membranes became disassociated from the carotene crvstalloid during alcohol dehydration which extracted some of the lycopene. Apparently, OS04 is able to penetrate and fix smaller crystalloids (Fig. 3, 4, 8, 16). However, the larger accumulations of carotene were lost during dehydration; light areas traversed by undulating membranes indicate where the carotenes were. These regions are termed crystalloid remnants (CrR). With glutaraldehydeKMnO4 fixation the membranes remained associated with the crystalloids which showed no undulations (Fig. 5). Though it was difficult to designate precisely where accumulations of lyco-