Starch Accumulation In Endosperm Research Articles

Pericarp starch metabolism is associated with caryopsis development and endosperm starch accumulation in common wheat

The wheat pericarp is the main component of the caryopsis at the early development stage and ultimately converts into a tissue that covers the mature caryopsis. A large number of starch granules are accumulated in the pericarp, but the production of and the role of starch granules in caryopsis development remain- elusive. In the present study, the relationship between accumulated starch granules and starch metabolism-related genes in wheat pericarp was investigated using paraffin section observations, expression analysis, and mutant analysis. Starch synthesis is initiated before anthesis and is dependent on a sucrose uptake and conversion system similar to that in the endosperm. TaPTST2 is required to initiate the production of pericarp starch granules. Pericarp starch granules gradually disappeared at the filling stage with high expression levels of genes encoding β-amylase, sucrose-phosphate synthase, and sucrose-phosphate phosphatase. As a maternal tissue adjacent to the endosperm and embryo, the pericarp plays a temporary reservoir for excess nutrients delivered into the caryopsis during the early development stage and exported at the filling stage. The pericarp contributes to the development of the endosperm and embryo as well as the accumulation of endosperm starch. The metabolism of pericarp starch may affect the weight of the wheat caryopsis.

The novel ZmTCP7 transcription factor targets AGPase-encoding gene ZmBt2 to regulate storage starch accumulation in maize.

The process of starch biosynthesis is a major developmental event that affects the final grain yield and quality in maize (Zea mays L.), and transcriptional regulation plays a key role in modulating the expression of the main players in the pathway. ZmBt2, which encodes the small subunits of AGPase, is a rate-controlling gene of the pathway; however, much remains unknown about its transcriptional regulation. Our earlier study identifies a short functional fragment of ZmBt2 promoter (394-bp), and further shows it contains multiple putative cis-acting regulatory elements, demonstrating that several transcription factors may govern ZmBt2 expression. Here, we identified a novel TCP transcription factor (TF), ZmTCP7, that interacted with the functional fragment of the ZmBt2 promoter in a yeast one hybrid screening system. We further showed that ZmTCP7 is a non-autonomous TF targeted to the nucleus and predominantly expressed in maize endosperm. Using promoter deletion analyzes by transient expression in maize endosperm protoplasts combined with electrophoretic mobility shift assays, we found that ZmTCP7 bound to GAACCCCAC elements on the ZmBt2 promoter to suppress its expression. Transgenic overexpression of ZmTCP7 in maize caused a significant repression of ZmBt2 transcription by ~77.58%, resulting in a 21.51% decrease in AGPase activity and a 9.58% reduction in the endosperm starch content of transgenic maize. Moreover, the expressions of ZmBt1, ZmSSI, ZmSSIIa, and ZmSSIIIa were increased, while those of ZmSh2 and ZmSSIV reduced significantly in the endosperm of the transgenic maize. Overall, this study shows that ZmTCP7 functions as a transcriptional repressor of ZmBt2 and a negative regulator of endosperm starch accumulation, providing new insights into the regulatory networks that govern ZmBt2 expression and starch biosynthesis pathway in maize.

Metabolic factors restricting sink strength in superior and inferior spikelets in high-yielding rice cultivars

Many high-yielding rice cultivars with large sink size (total number of spikelet per unit area × mean grain weight) have been developed, but some japonica cultivars developed in Japan often fail to attain the expected high yield due to low sink strength of spikelets. Although there is natural variation in sink strength of spikelets among high-yielding cultivars, metabolic factors involved in the natural variation and relationships of sink strength in spikelets with final percentage of filled spikelets are not fully understood. In the present study, we examined cultivar differences in sink strength for superior and inferior spikelets (i.e. earlier fertilizing spikelets with faster growth and later fertilizing ones with slower growth, respectively) in a panicle, using each spikelet at 10 d after the onset of development (10 DAD) when starch accumulation in endosperm was actively proceeding. Nine high-yielding cultivars were used: five japonica-dominant and four indica-dominant cultivars. Cultivar differences were observed in starch contents at 10 DAD in each spikelet type, and indica cultivars had higher starch contents than japonica cultivars in both superior and inferior spikelets. In addition, starch contents at 10 DAD were closely related to percentage of filled grains at maturity in both spikelet types. The activities of sucrose synthase (SUS) and uridine diphosphoglucose pyrophosphorylase (UGP), and the protein levels of phosphorylase 1 (Pho1), were higher in indica than japonica cultivars, and were positively correlated with starch contents at 10 DAD for both superior and inferior spikelets; although metabolic states, revealed from relations between intermediate metabolites and starch contents, differed among spikelet types. Consequently, it was considered that SUS and UGP at the step from sucrose cleavage to adenosine diphosphoglucose synthesis, and Pho1 at the starch biosynthesis step, were key metabolic factors involved in cultivar differences of sink strength (ability to synthesize starch).

Engineering 6-phosphogluconate dehydrogenase improves grain yield in heat-stressed maize

Endosperm starch synthesis is a primary determinant of grain yield and is sensitive to high-temperature stress. The maize chloroplast-localized 6-phosphogluconate dehydrogenase (6PGDH), PGD3, is critical for endosperm starch accumulation. Maize also has two cytosolic isozymes, PGD1 and PGD2, that are not required for kernel development. We found that cytosolic PGD1 and PGD2 isozymes have heat-stable activity, while amyloplast-localized PGD3 activity is labile under heat stress conditions. We targeted heat-stable 6PGDH to endosperm amyloplasts by fusing the Waxy1 chloroplast targeting the peptide coding sequence to the Pgd1 and Pgd2 open reading frames (ORFs). These WPGD1 and WPGD2 fusion proteins import into isolated chloroplasts, demonstrating a functional targeting sequence. Transgenic maize plants expressing WPGD1 and WPGD2 with an endosperm-specific promoter increased 6PGDH activity with enhanced heat stability in vitro. WPGD1 and WPGD2 transgenes complement the pgd3-defective kernel phenotype, indicating the fusion proteins are targeted to the amyloplast. In the field, the WPGD1 and WPGD2 transgenes can mitigate grain yield losses in high-nighttime-temperature conditions by increasing kernel number. These results provide insight into the subcellular distribution of metabolic activities in the endosperm and suggest the amyloplast pentose phosphate pathway is a heat-sensitive step in maize kernel metabolism that contributes to yield loss during heat stress.

찰옥수수 2기작 재배시 등숙 중 이삭 및 종실 특성 변화

본 연구에서는 간식용 풋옥수수로 이용되는 찰옥수수 주요 품종인 찰옥 4호와 일미찰을 2기작 재배하여 품질 좋은 찰옥수수 생산을 위한 재배법을 확립하고자 이삭과 종실 특성 변화를 조사하였다. 2기작 재배시 출사일수는 후기작이 전기작 보다 약 20일 정도 빨라졌고, 출사에서 수확적기까지는 약 10일 정도 느려졌다. 파종에서 출사까지의 적산온도는 약 <TEX>$1,380{\sim}1,420^{\circ}C$</TEX>, 출사에서 수확까지는 약 <TEX>$590{\sim}630^{\circ}C$</TEX> 정도 범위였다. 이삭무게와 이삭직경은 출사 후 일수가 경과함에 따라 유의하게 증가하였으나 파종시기에 따른 차이는 없었고, 이삭길이와 착립길이는 후기작에서 유의하게 짧았다(p<0.05). 알곡 줄수는 파종시기에 따른 차이를 보이지 않았고, 줄당 립수는 찰옥 4호가 전기작보다 후기작에 재배한 것이 적었으며 변이폭이 컸다(p<0.05). 등숙 중 종실의 길이, 너비, 두께와 100립 생체중, 전분함량은 출사후 일수에 따라 유의하게 증가하였다(p<0.05). 후기작 재배시 저온 등숙으로 전기작 보다 100립 생체중과 전분함량의 증가속도가 느려졌다. 찰옥수수의 전분립은 과피와 가까운 배유의 상단부터 집적되기 시작하여 출사 후 일수가 경과함에 따라 배유 내부까지 집적되었으며, 후기작 재배한 찰옥수수는 전분립 축적속도가 느리고 조밀하지 않은 것이 관찰되었다. 100립 생체중은 springiness(<TEX>$0.80^{***}$</TEX>), cohesiveness(<TEX>$0.44^{***}$</TEX>), adhesiveness(<TEX>$-0.80^{***}$</TEX>), chewiness(<TEX>$0.55^{***}$</TEX>), 전분함량은 springiness(<TEX>$0.79^{***}$</TEX>), adhesiveness(<TEX>$-0.81^{***}$</TEX>), chewiness(<TEX>$0.49^{**}$</TEX>)와 고도로 유의한 정의 상관을 나타내었다. This study was conducted to investigate the ear and kernel characteristics of waxy corn during ripening by double cropping, and to understand the pattern of starch accumulation in endosperm of waxy corn kernels. Chalok4 and Ilmichal were sown at April 20 (first cropping) and July 20 (second cropping) in 2011~2012. The accumulated temperature from silking to harvesting was about <TEX>$590{\sim}630^{\circ}C$</TEX>. It takes 23~24 days when Chalok4 and Ilmichal were sown in April 20, but July 20 sowing takes 32~35 days. Ear length and kernel set length were significantly shorter in second cropping (p<0.05). Kernel length, kernel width, 100-kernel weight, and starch content of waxy corn were increased as ears matured (p<0.05). Growth temperature was getting decreased during the ripening stage of second cropping, the rate of ear and kernel development had slowed. Starch granules started to accumulate in the cells around the pericarp, then developed in the cells around the embryo. In the second cropping, starch granules in the kernel of waxy corn were less compact than the first cropping, harvesting time of waxy corns can be extended. These results will be helpful to farmers for double cropping of waxy corn cultivation and management.

Chloroplast-localized 6-phosphogluconate dehydrogenase is critical for maize endosperm starch accumulation

Plants have duplicate versions of the oxidative pentose phosphate pathway (oxPPP) enzymes with a subset localized to the chloroplast. The chloroplast oxPPP provides NADPH and pentose sugars for multiple metabolic pathways. This study identified two loss-of-function alleles of the Zea mays (maize) chloroplast-localized oxPPP enzyme 6-phosphogluconate dehydrogenase (6PGDH). These mutations caused a rough endosperm seed phenotype with reduced embryo oil and endosperm starch. Genetic translocation experiments showed that pgd3 has separate, essential roles in both endosperm and embryo development. Endosperm metabolite profiling experiments indicated that pgd3 shifts redox-related metabolites and increases reducing sugars similar to starch-biosynthetis mutants. Heavy isotope-labelling experiments indicates that carbon flux into starch is altered in pgd3 mutants. Labelling experiments with a loss of cytosolic 6PGDH did not affect flux into starch. These results support the known role for plastid-localized oxPPP in oil synthesis and argue that amyloplast-localized oxPPP reactions are integral to endosperm starch accumulation in maize kernels.

Культура ізольованих зернівок як біотехнологічна система дорощування зиготичних зародків кукурудзи in vitro

The investigation established the possibility of production of maize embryo in culture of isolated caryopsis on artificial nutrient medium in vitro conditions from a zygote / proembryo and globular stage up to the full maturity. It was determined that the cultivation of caryopsis on artificial nutrient medium provides the accumulation of starch in endosperm. The internal state of cultured caryopsis can be considered as a marker of embryo upgrowing and starch accumulation in endosperm, irrespective of the genotype. The genotype, the age of cultured caryopsis and concentration of sucrose influence on the development of endosperm, accumulation of starch in endosperm and embryo upgrowing from the zygote / proembryo and globular stage.

Культура ізольованих зародкових мішків як біотехнологічна система дорощування зиготичних зародків кукурудзи in vitro

Установлено можливість дорозвинення зародка кукурудзи в культурі ізольованих зародкових мішків на штучному живильному середовищі в умовах in vitro від ранньої перехідної стадії до початку органогенезу. З’ясовано, що культивування зародкових мішків з метою дорощування зародка та накопичення крохмалю в ендоспермі повинно проводитися понад 20 діб. Забарвлення культивованого зародкового мішка може слугувати маркерною ознакою дорозвинення зародка та накопичення крохмалю в ендоспермі.

Current status and strategies for improvement of rice grain chalkiness

This paper reviews the current status of correlation between rice chalkiness and other rice quality characters, formative mechanism, and classical and molecular genetics for rice chalkiness. The formation of rice chalkiness proves to be a complicated physiological process and tightly relate to "source-sink" of rice, dynamics of grain filing, biosynthesis and accumulation of starch in endosperm. Rice chalkiness is a complicated quantitative trait, which is controlled by maternal effects, endosperm effects, and cytoplasmic effects. There are some stable quantitative trait loci (QTL) for rice chalkiness on many rice chromosomes. Of them, three genes controlling rice chalkiness, which have an impact on starch synthesis, starch metabolism, and fruit development, have been cloned. But the manipulative network and formative mechanisms of rice chalkiness remain unclear. At present in breeding practice, decrease of rice chalkiness has become one of the main aims in rice quality breeding, especially for indica rice. The direction of genetic research and improvement strategy of rice chalkiness in future were discussed in this review.

The Allosterically Unregulated Isoform of ADP-Glucose Pyrophosphorylase from Barley Endosperm Is the Most Likely Source of ADP-Glucose Incorporated into Endosperm Starch.

We present the results of studies of an unmodified version of the recombinant major barley (Hordeum vulgare) endosperm ADP-glucose pyrophoshorylase (AGPase) expressed in insect cells, which corroborate previous data that this isoform of the enzyme acts independently of the allosteric regulators 3-phosphoglycerate and inorganic phosphate. We also present a characterization of the individual subunits expressed separately in insect cells, showing that the SS AGPase is active in the presence of 3-phosphoglycerate and is inhibited by inorganic phosphate. As a step toward the elucidation of the role of the two AGPase isoforms in barley, the temporal and spatial expression profile of the four barley AGPase transcripts encoding these isoforms were studied. The results show that the steady-state level of beps and bepl, the transcripts encoding the major endosperm isoform, correlated positively with the rate of endosperm starch accumulation. In contrast, blps and blpl, the transcripts encoding the major leaf isoform, were constitutively expressed at a very low steady-state level throughout the barley plant. The implications of these findings for the evolution of plant AGPases are discussed.

Nitrogen-Induced Changes in the Growth and Metabolism of Developing Maize Kernels Grown in Vitro

Cereal kernel growth and grain yield are functions of endosperm starch accumulation. The objective of this study was to examine how various metabolic factors in developing maize (Zea mays L.) endosperm influence starch deposition. Kernels were grown in vitro on medium with: (a) zero N (-N), (b) optimum N (+N), or (c) -N from 3 to 20 days after pollination followed by +N until maturity (+/-N) to produce different degrees of endosperm growth and to promote an enhancement of starch synthesis midway through development. At intervals, kernels were harvested and levels of enzyme activities and carbohydrate and N constituents examined. Endosperm starch and protein accumulation were decreased in -N compared to +N kernels, but relief of N starvation increased both constituents. With greater movement of N into +/-N kernels, endosperm sugar concentrations declined suggesting an inverse relationship between C and N transport. Unusually high concentrations of sugar in N stressed kernels did not appear to limit or enhance starch production. Rather, increased accumulation of starch in +/-N endosperm was correlated with significant increases in the enzymatic activities of sucrose synthase and PPi-linked phosphofructokinase, and to a lessor extent hexokinase. In addition, the occurrence of specific proteins of the albumin/globulin fraction either increased, decreased, or remained unchanged in relation to starch synthesis. These data suggest that lack of N limits starch deposition in maize endosperm primarily through an influence on synthesis of key proteins.

Dry Matter Accumulation and Carbohydrate Concentration Patterns of Field‐Grown and in Vitro Cultured Maize Kernels from the Tip and Middle Ear Positions1

Tip and middle position kernels from field‐grown ears and in vitro cultured maize (Zea mays L.) were compared for dry matter accumulation and pedicel and endosperm carbohydrate concentration patterns. The experiment was designed to determine whether the growth of tip kernels is limited by their inherent potential to accumulate dry matter or by the supply of assimilate entering the kernel. Tip kernels from field‐grown ears had the lowest linear dry matter accumulation rate, the shortest duration of kernel growth, and therefore, the lowest maximum mass. The mass of tip kernels cultured in vitro was three times greater than tip kernels from field‐grown ears, which indicated that tip kernels on field‐grown ears fail to attain maximum potential mass. The mass of tip kernels cultured in vitro was lower than the mass of middle kernels cultured in vitro or from field‐grown ears. Tip kernels that developed on field‐grown ears had a much lower pedicel fructose, glucose, and sucrose concentration throughout kernel growth than did tip kernels cultured in vitro and middle kernels of both field grown and in vitro culture. Tip kernels from the field also had a reduced maximum endosperm sucrose concentration as compared to tip and middle kernels cultured in vitro and middle kernels from field‐grown ears. The relative endosperm starch accumulation of field‐grown tip kernels was comparable to that of tip kernels cultured in vitro. Starch synthesis in the endosperm of tip kernels from the field was initiated 4 days later than in tip and middle kernels cultured in vitro and middle kernels from field‐grown ears. The low sugar concentrations in the pedicel and the decreased endosperm sucrose concentration in tip kernels from field‐grown ears indicate that an inadequate assimilate supply reaching these kernels may be limiting their growth more than their inherent potential to accumulate dry matter.