Early Stages in Wheat Endosperm Formation and Protein Body Initiation

The early stages of endosperm formation and protein body initiationare described for hard red winter wheat using light and transmissionelectron microscopy. Two days after flowering (DAF) the endospermwas a thin layer of coenocytic cytoplasm lining the embryo sac.By 4 DAF the endosperm had cellularized and completely filledthe embryo sac. Enough differentiation had occurred by 6 DAFto distinguish cells destined to become the aleurone layer,sub-aleurone region and central endosperm. Protein bodies wereinitiated at about 6–7 DAF and were first found near theGolgi apparatus. Wheat was ready for combine harvest at 34 DAF.Enlargement of the small protein bodies near the Golgi apparatusoccurred by several mechanisms: (1) fusion with one or moreof the dense Golgi vesicles or fusion with other protein bodies,(2) fusion with small electron-lucent Golgi-derived vesicles,(3) pinocytosis of a portion of the adjacent cytoplasm intothe developing protein body and (4) fusion of large proteinbodies with one another at later stages of grain development.Of the four mechanisms described, the pinocytotic vesicles andfusion of protein bodies were the most frequent and consistentprocesses observed. Direct connections between rough endoplasmicreticulum (RER) and protein bodies were not observed. The resultssuggest a rôle for the Golgi apparatus in the initiationof protein bodies. Also, the lack of RER derived vesicles suggestsa soluble mode of secretion of storage proteins involved inthe enlargement of protein bodies. Triticum aestivum, wheat endosperm, protein bodies Golgi apparatus     

Protein Body Inclusions in Developing Wheat Endosperm

Endosperm tissue of two wheat cultivars, Maris Freeman and Mardlerwas examined by light and electron microscopy from early stagesof development until maturity. Spherical electron-dense inclusionswere first seen embedded in the periphery of endosperm proteinbodies of both cultivars 11 days after anthesis. The inclusions,which gave a positive histochemical reaction for both proteinand lipid, persisted throughout endosperm development and werepresent in the protein matrix of mature grain. Two types ofmembranous inclusions were found. In Maris Freeman and Mardler,vesicles and myelin whorls were associated with the surfaceof the protein mass during development and maturation. In MarisFreeman, membrane lattices of branched tubules with a basiccubic repeat unit of 44.8 nm were found in close contact withthe protein mass, but these were not present in mature grain. Triticum aestivum L., wheat, endosperm, protein body inclusions, ultrastructure     

Pullulanase and Starch Synthase III Are Associated with Formation of Vitreous Endosperm in Quality Protein Maize

The opaque-2 (o2) mutation of maize increases lysine content, but the low seed density and soft texture of this type of mutant are undesirable. Lines with modifiers of the soft kernel phenotype (mo2) called “Quality Protein Maize” (QPM) have high lysine and kernel phenotypes similar to normal maize. Prior research indicated that the formation of vitreous endosperm in QPM might involve changes in starch granule structure. In this study, we focused on analysis of two starch biosynthetic enzymes that may influence kernel vitreousness. Analysis of recombinant inbred lines derived from a cross of W64Ao2 and K0326Y revealed that pullulanase activity had significant positive correlation with kernel vitreousness. We also found that decreased Starch Synthase III abundance may decrease the pullulanase activity and average glucan chain length given the same Zpu1 genotype. Therefore, Starch Synthase III could indirectly influence the kernel vitreousness by affecting pullulanase activity and coordinating with pullulanase to alter the glucan chain length distribution of amylopectin, resulting in different starch structural properties. The glucan chain length distribution had strong positive correlation with the polydispersity index of glucan chains, which was positively associated with the kernel vitreousness based on nonlinear regression analysis. Therefore, we propose that pullulanase and Starch Synthase III are two important factors responsible for the formation of the vitreous phenotype of QPM endosperms.     

玉米未成熟胚乳愈伤组织和器官的发生及其倍性变化的初步研究

本试验在附加和不附加外源激素的MS培养基上,均得到了玉米未成熟胚乳愈伤组织。愈伤组织在附加外源激素的MS培养基上达到器官分化,获得了发育正常的和许多畸形的胚乳植株。所得到的愈伤组织细胞和植株根尖细胞染色体数目和倍性是不稳定的,二者有相同的趋向,其中有整倍体的细胞(2n=10,20,30,40,50),也有各种非整倍体的细胞(2n=5—49)。     

Endosperm Protein Quality and Kernel Modification in the Quality Protein Maize Inbred Lines

Twenty-two selected quality protein maize (QPM) lines, including 13 lines developed in India (DMRQPM series) and nine lines released by CIMMYT, Mexico (CML series), were evaluated for their endosperm protein content and quality, besides kernel modification in terms of vitreousness. Endosperm protein contents in 13QPMlines were on par or better than that of the normal maize ‘checks’ (Trishulata and Parkash). The QPM endosperm proteins showed significantly higher % tryptophan as well as EF-1α (a multifunctional protein with a positive and highly significant correlation with lysine content in the endosperm) contents, in comparison with the normal maize genotypes. Evaluation of kernel modification revealed considerable scope for accumulation of endosperm modifiers in some of the QPM lines. Positive and highly significant correlation was revealed between tryptophan and EF-1α contents in the endosperm proteins, whereas the correlations between the quality parameters with kernel modification in the QPM genotypes were found to be non-significant. The study led to the identification of some promising QPM lines, such as DMRQPM-37, DMRQPM-44, CML176, CML142 and CML149, which could be effectively deployed in the QPM breeding programmes.     

Protein Body Degradation in the Starchy Endosperm of Germinating Sorghum

Taylor, J. R. N., Novellie, L. and Liebenberg, N. v. d. W. 1985.Protein body degradation in the starchy endosperm of germinatingsorghum.—J. exp. Bot. 36: 1287–1295. Transmission electron micrographs of starchy endosperms of germinatingsorghum indicate that the protein bodies are degraded predominantlyby progressive hydrolysis of prolamin from their surface. Theappearance of holes within partially degraded protein bodiesindicates that some internal hydrolysis also takes place. Chemicalanalyses of protein bodies isolated at different stages duringgermination showed that their amino acid composition and electrophoreticpattern remained relatively unchanged during hydrolysis. Theend result of protein body degradation was that the organellescompletely disappeared leaving empty starchy endosperm cells.The protein bodies did not swell prior to or during degradation.This mode of protein body degradation differs from that in germinatingdicotyledonous seeds and in the aleurone layer and embryo ofcereal seeds but was identical to the mode of prolamin proteinbody degradation in the starchy endosperm of germinating riceseeds. Key words: Sorghum bicolor, protein body degradation, prolamin     

Changes in Ultrastructure and Subunit Composition of Protein Body in Rice Endosperm during Germination

Changes in ultrastructure of protein bodies in subaleurone cells of rice endosperm during germination were studied by transmission electron microscopy. The subaleurone cells contained two different types of protein bodies: PB-I (spherical) and PB-II (crystalline). Both types of protein bodies were deconstructed during germination. But there was a considerable difference in digestibility between PB-I and PB-II. PB-II which did not have a dense core was easily digested from the central portion when germination began. At 6 days of germination, PB-II was almost deconstructed. On the other hand, PB-I which displayed concentric rings with a dense core was digested from the outside after 3 days of germination. At 9 days of germination, many kernels of the spherical protein bodies remained.

Changes in subunit composition of protein bodies during germination were investigated by SDS-polyacrylamide gel electrophoresis. Protein body fractions were isolated from germinating grains at various stages by enzymatic digestion and two-phase system, then subjected to SDS-polyacrylamide gel. As germination proceeded, 15 (b₁), 20 (d₁), 24 (e), 35 (f₁) and 37 (f₃) kdaltons subunits decreased. On the other hand, 16 (b₂), 21 (d₂) and 36 (f₂) k daltons subunits remained at the later stage of germination. We think that PB-I contains b₂, d₂ and f₂ subunits and is attacked only from the outside at middle and later stages of germination by de novo protease. On the contrary, PB-II contains b₁ d₁ e, f₁ and f₃ subunits is utilized at an early stage of germination. PB-II may possibly contain latent protease. The breakdown process of PB-I was by exo-type digestion, on the contrary, that of PB-II was by endo-type digestion.     

Embryo and Endosperm Phytochemicals from Polyembryonic Maize Kernels and Their Relationship with Seed Germination

Because of the growing worldwide demand for maize grain, new alternatives have been sought for breeding of this cereal, e.g., development of polyembryonic varieties, which agronomic performance could positively impact the grain yield per unit area, and nutritional quality. The objectives of this study were to (1) determine the phytochemicals present in the embryo and endosperm of grain from maize families with high, low, and null polyembryony frequency, which were planted at different locations, and (2) state the relationship between these compounds and seed germination. The extracted phytochemicals from corn were identified by HPLC-MS. The results showed that the genotype with the highest presence of phytochemicals was the brachytic population with high polyembryony called “BAP”, which also required less water during the germination process. The number of phytochemicals in both embryo and endosperm tissues was not related to the sowing location where they came from or the type of polyembryony. The number of different phytochemicals depended on the grain tissue from where they were extracted. The chemical compounds found in the different maize tissues were related to the development of the plant, either in roots or nibs because these are mainly associated with the lignin synthesis.     

Tau Protein Function in Axonal Formation

Tau protein is a predominantly neuronal microtubule-associated protein that is enriched in axons and is capable of promoting microtubule assembly and stabilization. In the present article we review some of the key experiments directed to obtain insights about tau protein function in developing neurons. Aspects related to whether or not tau has essential, unique, or complementary functions during axonal formation are discussed.     

Molecular Species in the Protein Body II (PB-II) of Developing Rice Endosperm

The molecular weight and subunit composition of glutelin, the major storage protein of rice, in the major type of protein bodies of developing rice seeds was examined by gradient and two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Glutelin in the protein body was the assembled from heterogeneous subunits, and the molecular weights were estimated to be 64, 140, 240, 320, 380, and 500 k by gradient SDS-PAGE. High molecular weight proteins (larger than 2,000 k) were also observed.

The two-dimensional SDS-PAGE under reduced conditions showed, that the glutelin in the protein body was composed of two groups of polypeptides, 22~23 and 37~39k, bound by disulfide linkages.     

Correlation between Protein Body Formation and Storage Protein Accumulation in Soybean Cotyledons

At 20 days after flowering (DAF), the 7S α' and α subunits began to accumulate. At 25 DAF, the 7Sβ, l1SA and llSB subunits appeared. Five days later, the 11SA-4 subunit was present During the period of 25–55DAF, the storage protein content continued to increase. From 55 to 63 DAF, there was a decrease in the synthetic rate of the storage proteins. Comparing these results with the two paths of protein body formation reported previously, we draw the conclusion that the protein bodies developed from vacuoles contained not only the 7S bm also the lis proteins in soybean cotyledon cells.     

Phosphorylases I and II of Maize Endosperm

Two phosphorylases have been found in the endosperm of Zea mays. Phosphorylase I is found through all stages of endosperm development and seed germination investigated. The other enzyme, phosphorylase II appears only at the stage of rapid starch biosynthesis and is not found during germination. At 22 days after pollination, the activity of phosphorylase II is 10 times that of phosphorylase I. These 2 phosphorylases are separable by column chromatography and behave differently in several respects.     

Endosperm Protein Synthesis and l-[S]Methionine Incorporation in Maize Kernels Cultured In Vitro

This study was conducted to examine protein synthesis and l-[³⁵S] methionine incorporation into the endosperm of Zea mays L. kernels developing in vitro. Two-day-old kernels of the inbred line W64A were placed in culture on a defined medium containing 10 microCuries l-[³⁵S] methionine per milliliter (13 milliCuries per millimole) and harvested at 10, 15, 20, 25, 30, 35, and 40 days after pollination. Cultured kernels attained a final endosperm mass of 120 milligrams compared to 175 milligrams for field-grown controls. Field and cultured kernels had similar concentrations (microgram per milligram endospern) for total protein, albumin plus globulin, zein, and glutelin fractions at most kernel ages.     

Effect of Gibberellin on Growth, Protein Secretion, and Starch Accumulation in Maize Endosperm Suspension Cells

The physiologic effect of gibberellins (GA) in seed development is poorly understood. We examined the effect of gibberellic acid (GA₃) on growth, protein secretion, and starch accumulation in cultured maize (Zea mays L.) endosperm suspension cells. GA₃ (5 and 30 μm) increased the fresh weight, dry weight, and protein content of the cultured cells, but the effect of GA₃ at 50 μm was not significantly different. However, the protein content in the culture medium was increased by these three concentrations of GA₃. The effect of GA₃ on the amount of cellular structural polysaccharides was not significant, but GA₃ had a dramatic effect on the starch content. At 5 μm, GA₃ caused an increase in the starch content, but at 50 μm the starch accumulation was reduced. Chlorocholine chloride (CCC), an inhibitor of GA biosynthesis, significantly increased the starch content and decreased the structural polysaccharide content of the cultured cells. The effects of CCC at 500 μm on the starch and polysaccharide content were partially reversed by 5 μm GA₃ applied exogenously. Based on these results we suggest that GA does not favor starch accumulation in the cell cultures and that the addition of lower concentrations of GA₃ in the medium may provide an improved balance among the endogenous GA in the cultured cells. Received October 31, 1995; accepted March 25, 1997     

Starch-synthesizing Enzymes in the Endosperm and Pollen of Maize

Two mutations, amylose-extender and waxy, which affect the proportion of amylose and amylopectin of starch synthesized in the endosperm of maize (Zea mays L.) seeds, are also expressed in the pollen. However, most mutations that affect starch synthesis in the maize endosperm are not expressed in the pollen. In an attempt to understand the nonconcordance between the endosperm and pollen, extracts of mature pollen grains were assayed for a number of the enzymes possibly implicated in starch synthesis in the endosperm. Sucrose synthetase (sucrose-UDP glucosyl transferase, EC 2.4.1.13) activity was not detectable in either mature or immature pollen grains of nonmutant maize, but both bound and soluble invertase (EC 3.2.1.26) exhibited much greater specific activity (per milligram protein) in pollen extracts than in 22-day-old endosperm extracts. Phosphorylase (EC 2.4.1.1) activity was also higher in pollen than in endosperm extracts. ADP-Glucose pyrophosphorylase (EC 2.7.7.27) activity was much lower in pollen than endosperm extracts, but mutations that drastically reduced ADP-glucose pyrophosphorylase activity in the endosperm (brittle-2 and shrunken-2) did not markedly affect enzymic activity in the pollen. Specific activities of other enzymes implicated in starch synthesis were similar in endosperm and pollen extracts.