Changes in Gene Expression during Tomato Fruit Ripening

Total proteins from pericarp tissue of different chronological ages from normally ripening tomato (Lycopersicon esculentum Mill. cv Rutgers) fruits and from fruits of the isogenic ripening-impaired mutants rin, nor, and Nr were extracted and separated by sodium dodecylsulfate-polyacrylamide gel electrophoresis. Analysis of the stained bands revealed increases in 5 polypeptides (94, 44, 34, 20, and 12 kilodaltons), decreases in 12 polypeptides (106, 98, 88, 76, 64, 52, 48, 45, 36, 28, 25, and 15 kilodaltons), and fluctuations in 5 polypeptides (85, 60, 26, 21, and 16 kilodaltons) as normal ripening proceeded. Several polypeptides present in ripening normal pericarp exhibited very low or undetectable levels in developing mutant pericarp. Total RNAs extracted from various stages of Rutgers pericarp and from 60 to 65 days old rin, nor, and Nr pericarp were fractionated into poly(A)⁺ and poly(A)⁻ RNAs. Peak levels of total RNA, poly(A)⁺ RNA, and poly(A)⁺ RNA as percent of total RNA occurred between the mature green to breaker stages of normal pericarp. In vitro translation of poly(A)⁺ RNAs from normal pericarp in rabbit reticulocyte lysates revealed increases in mRNAs for 9 polypeptides (116, 89, 70, 42, 38, 33, 31, 29, and 26 kilodaltons), decreases in mRNAs for 2 polypeptides (41 and 35 kilodaltons), and fluctuations in mRNAs for 5 polypeptides (156, 53, 39, 30, and 14 kilodaltons) during normal ripening. Analysis of two-dimensional separation of in vitro translated polypeptides from poly(A)⁺ RNAs isolated from different developmental stages revealed even more extensive changes in mRNA populations during ripening. In addition, a polygalacturonase precursor (54 kilodaltons) was immunoprecipitated from breaker, turning, red ripe, and 65 days old Nr in vitro translation products.     

香蕉果实成熟软化过程中细胞壁物质的变化

系统研究了香蕉果实软化过程中细胞壁物质―醇不溶性固形物(AIS)以及3种不同性质的果胶物质:水溶性果胶(WSP)、酸溶性果胶(HP)和碱溶性果胶(OHP)含量的变化。结果表明:随果实的成熟软化,AIS的含量不断降低,且在呼吸跃变时急剧降低;WSP的含量不断增加,HP和OHP的含量不断减少,且均表现出在早期变化量少,在果实硬度迅速降低时变化明显。该研究进一步证明细胞壁物质的变化是导致香蕉果实软化的主要原因。     

Polyribosomes from Pear Fruit: Changes during Ripening and Senescence

Polysome profiles were examined from lyophilized peel tissue of ripening pear (Pyrus communis, L. var. Passe-Crassane). Messenger RNA chains bearing up to eight ribosomes (octamers) were resolved and exhibited the highest absorption peak when ribonuclease activity was eliminated during extraction. Neither normal ripening nor the increase of large polyribosomes that normally accompanies ripening and senescence of the fruit occurred when pretreatment at 0 C was omitted. Normal ripening and increase of large polyribosomes would, however, be initiated by an ethylene treatment. The size distribution of the polyribosomes remained essentially constant throughout a 4-month cold storage; there was, however, a large increase in ribosomes by the 12th week of storage.     

Changes in mRNA and Protein during Ripening in Apple Fruit (Malus domestica Borkh. cv Golden Delicious)

Poly(A)⁺ RNA was extracted from cortical tissue of apple fruit (Malus domestica Borkh. cv Golden Delicious), and in vitro translation products were analyzed by two-dimensional gel electrophoresis. As fruit internal ethylene concentration increased from a basal level of 0.0 to 0.1 microliter per liter to 1 to 5 microliters/liter, substantial changes in the pattern of in vitro translation products were observed. More subtle changes were observed as fruit continued to ripen and internal ethylene concentration increased to 80 to 100 microliters/liter. Overall, the levels of at least six mRNAs were found to increase, while one mRNA decreased. Analysis of proteins extracted from ripening fruit indicated that the level of at least three proteins increased with ripening.     

不同品种苹果采后后熟软化过程中细胞壁多糖的降解

以2种苹果为试材,提取了不同贮藏时期果实的细胞壁物质和8种细胞壁多糖组分,并采用气相色谱法分析了细胞壁多糖组分的单糖组成。结果表明,在贮藏过程中,‘金星’苹果果肉的硬度下降明显,在贮藏第10天前后出现明显的乙烯释放量高峰,而耐贮藏性‘富士’苹果在贮藏期间只释放极少量的乙烯。‘金星’苹果的Na2CO3-1溶性果胶多糖组分的减少尤为显著。这些结果表明,苹果果实Na2CO3-1溶性果胶多糖组分侧链成分的酶降解,是引起苹果细胞壁多糖网络结构的变化,进而导致果实软化的重要原因之一。     

Changes in Cell Wall Polysaccharides During Fruit Ripening

Received 20 June 2000/ Accepted in revised form 20 July 2000     

The Fine Structure of Apple, Pear, and Plum Fruit Surfaces, their Changes during Ripening, and their Response to Polishing

The appearance of a fruit depends upon the way in which lightis reflected and scattered at its surface, and this is partlydetermined by the form of the surface waxes. The fine fibrilspredominating on the surface of plums scatter light much morethan the platelets present on apples and pears, and cause thecharacteristic and prominent bloom. Polishing crushes the wax elements together, giving a much smootherreflecting surface, and sometimes the wax layer completely recrystallizes.The bloom, however, begins to regenerate within a few days,but it does not attain its original prominence. Changes comparable with those caused by polishing occur duringthe ripening of some varieties of apple.     

Changes in Structural Features of Free N-Glycan and Endoglycosidase Activity during Tomato Fruit Ripening

In developing plants, free N-glycans occur ubiquitously at micromolar concentrations. Such oligosaccharides have been proposed to be signaling molecules in plant development. As a part of a study to elucidate the physiological roles of de-N-glycosylation machinery involved in fruit ripening, we analyzed changes in the amounts and structural features of free N-glycans in tomato fruits at four ripening stages. The amount of high-mannose type free N-glycans increased significantly in accordance with fruit ripening, and the relative amounts of high-molecular size N-glycans, such as Man_8-9GlcNAc₁, became predominant. These observations suggest that the de-N-glycosylation machinery, including endo-β-N-acetylglucosaminidase (ENGase) activity, is stimulated in the later stages of fruit ripening. But contrary to expectation, we found that total ENGase activities in the tomato fruits did not vary significantly with the ripening process, suggesting that ENGase activity must be maintained at a certain level, and that the expression of α-mannosidase involved in the clearance of free N-glycans decreases during tomato fruit ripening.     

Differential Protein Accumulation in Banana Fruit during Ripening

Banana (Musa acuminata, cv Dwarf Cavendish) proteins were extracted from pulp tissue at different stages of ripening and analyzed by two-dimensional electrophoresis. The results provide evidence of differential protein accumulation during ripening. Two sets of polypeptides have been detected that increase substantially in ripe fruit. These polypeptides were characterized as glycoproteins by western blotting and concanavalin A binding assays. Antibodies againts tomato polygalacturonase cross-react with one of these sets of proteins.     

Effects of Calmodulin Antagonists on the Mitochondrial and Microsomal Ca2$ Uptake in Apple Fruit

In apple fruit, active ATP-dependent microsomal Ca^2$ uptakeand respiration-dependent mitochondrial Ca^2$ uptake were observed. The mitochondrial Ca^2$ uptake was depressed by the calmodulinantagonists chlorpromazine hydrochloride (CPZ) and N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamidehydrochloride (W-7). The Ca^2$-ATPase from apple mitochondriawas also inhibited by CPZ or W-7. The apparent K_m value forCa^2$ in mitochondrial Ca^2$ uptake (K_m=0.35 mM) was similar tothat of mitochondrial Ca^2$-ATPase (K_m=0.32 mM). The inhibitoryeffect of W-7 on the activity of the mitochondrial Ca^2$ uptakewas closely correlated with the inhibition by W-7 of mitochondrialCa^2$-ATPase (r=0.996). These findings indicate that the mitochondrialuptake of Ca^2$ in apple fruit depends on the calmodulin-mediatedactivation of Ca^2$-ATPase. The microsomal Ca^2$ uptake was depressed by CPZ, suggestingthat the microsomal Ca^2$ uptake may also be modulated by calmodulin. ¹ Contribution No. C-72, Fruit Tree Research Station. (Received June 7, 1982; Accepted October 19, 1982)     

Regulation of Ethylene Biosynthesis in Avocado Fruit during Ripening

Preclimacteric avocado (Persea americana Mill.) fruits produced very little ethylene and had only a trace amount of l-aminocyclopropane-1-carboxylic acid (ACC) and a very low activity of ACC synthase. In contrast, a significant amount of l-(malonylamino)cyclopropane-1-carboxylic acid (MACC) was detected during the preclimacteric stage. In harvested fruits, both ACC synthase activity and the level of ACC increased markedly during the climacteric rise reaching a peak shortly before the climacteric peak. The level of MACC also increased at the climacteric stage. Cycloheximide and cordycepin inhibited the synthesis of ACC synthase in discs excised from preclimacteric fruits. A low but measurable ethylene forming enzyme (EFE) activity was detected during the preclimacteric stage. During ripening, EFE activity increased only at the beginning of the climacteric rise. ACC synthase and EFE activities and the ACC level declined rapidly after the climacteric peak. Application of ACC to attached or detached fruits resulted in increased ethylene production and ripening of the fruits. Exogenous ethylene stimulated EFE activity in intact fruits prior to the increase in ethylene production. The data suggest that conversion of S-adenosylmethionine to ACC is the major factor limiting ethylene production during the preclimacteric stage. ACC synthase is first synthesized during ripening and this leads to the production of ethylene which in turn induces an additional increase in ACC synthase activity. Only when ethylene reaches a certain level does it induce increased EFE activity.     

Degradation of Cell Wall Polysaccharides during Tomato Fruit Ripening

Changes in neutral sugar, uronic acid, and protein content of tomato (Lycopersicon esculentum Mill) cell walls during ripening were characterized. The only components to decline in amount were galactose, arabinose, and galacturonic acid. Isolated cell walls of ripening fruit contained a water-soluble polyuronide, possibly a product of in vivo polygalacturonase action. This polyuronide and the one obtained by incubating walls from mature green fruit with tomato polygalacturonase contained relatively much less neutral sugar than did intact cell walls. The ripening-related decline in galactose and arabinose content appeared to be separate from polyuronide solubilization. In the rin mutant, the postharvest loss of these neutral sugars occurred in the absence of polygalacturonase and polyuronide solubilization. The enzyme(s) responsible for the removal of galactose and arabinose was not identified; a tomato cell wall polysaccharide containing galactose and arabinose (6:1) was not hydrolyzed by tomato β-galactosidase.     

Ultrastructure of the Mesocarp of Mature Avocado Fruit and Changes Associated with Ripening

The mesocarp tissue of ripening avocado fruits was studied byfreeze fracture, thin section and scanning electron microscopy.Carbon dioxide and ethylene production by individual fruit weremonitored, and samples were analysed at several stages of theripening process. The tissue is composed primarily of large, isodiametric, lipid-containingparenchyma cells. At maturity these cells contain the normalcomplement of cell organelles, and all membranes appear intact.When ripening begins, several changes in the ultrastructureoccur. The most obvious changes are a loosening and eventualbreakdown of the cell wall, and swelling and vesiculation ofthe rough endoplasmic reticulum. In freeze fracture replicasa significant increase in the number of intramembranous particlesin the EF face of the plasmamembrane was observed at the climactericpeak. In post-climacteric, soft fruit the particle density of theEF face of the plasmamembrane decreased to the density observedin the membrane of pre-climacteric cells. All of the organellesand membranes appear whole and intact whether examined by thinsection, freeze fracture or scanning electron microscopy. However,the cell walls in post-climacteric fruit have almost completelydisappeared. These results indicated that the ripening process per se inavocados does not involve a complete loss of compartmentationnor a breakdown of organelle and membrane integrity. It may,however, lead to these or similar senescence changes as a resultof the loss of the cell walls. The variations in particle densityof the plasmamembrane during ripening may reflect one or moreof several structural, compositional, or functional membranephenomena, and this aspect of ripening warrants further study. Persea americana Mill., avocado pear, freeze fracture, fruit ripening, scanning electron microscopy, senescence, ultrastructure     

Temperature-dependent Changes in the Polysomal Population of Senescent (Ripening) Pear Fruit

A loss and recovery of polysomes coincident with the temperature-dependent interruption and resumption of pear fruit (Pyrus communis, L.) senescence establishes a close correlation between the presence of protein-synthesizing machinery and the progression of senescence and ripening.     

Flow of Chromium into Apple Fruit during Development

A continuous flow of chromium into apples occurs throughout their growth. This ready movement of chromium into the apple is similar to that reported for essential trace elements such as boron, zinc, iron, copper, and manganese but differs from the restricted movement reported for mercury and cadmium.