Imprinted gene expression in maize starchy endosperm and aleurone tissues of reciprocal F1 hybrids at a defined developmental stage

Meishan Zhang,Ruili Lv,Wei Yang,Tiansi Fu,Bao Liu 한국유전학회 2018 Genes & Genomics Vol.40 No.1

Imprinted gene expression in flowering plants predominantly occurs in the triploid endosperm of developing seed. However, endosperm is composed of distinct tissue types. For example, the maize (Zea mays) endosperm is constituted by two major tissues, starchy endosperm and aleurone. Previous studies in imprinted gene expression have generally assumed that the different tissues constituting endosperm would behavior the same, and hence have not examined them separately. Here, to examine parentalspecific expression of imprinted genes in different parts of the seed, eight previously reported maize protein-coding imprinted genes were selected, and analyzed by cleaved amplified polymorphic sequence (CAPS) coupled with Sanger sequencing for transcripts from the various seed tissues collected at 18 days after pollination (DAP). The studied tissues included seed coat, embryo, starchy endosperm and aleurone, which were collected from a pair of reciprocal F1 hybrids produced by crossing inbred lines B73 and Mo17. Six of these eight analyzed imprinted genes showed the same imprinted expression pattern between the starchy endosperm and aleurone, but two showed imprinted expression only in the starchy endosperm. Comparison of the expression pattern of 20 selected imprinted genes in multiple seed tissues and vegetative tissues indicated that the majority (~ 75%) of these imprinted genes exhibited seedspecific or endosperm-specific expression. Our results also uncovered that imprinted genes have a high propensity to be alternatively spliced via intron retention in the developing embryo compared with the other tissues.

Genome-wide identification and extensive analysis of rice-endosperm preferred genes using reference expression database

홍우종,유요한,박선아,문선옥,김성렬,안진흥,정기홍 한국식물학회 2017 Journal of Plant Biology Vol.60 No.3

Studying endosperm development in crop species enables us to understand the molecular mechanisms for producing and metabolizing carbohydrates as a main energy source. A significant accumulation of genome-wide transcriptome data can enhance the performance of metaexpression data for diverse applications. Using a gene search tool in Genevestigator (https://genevestigator.com/) for anatomical samples, we first conducted a meta-anatomical expression analysis based on 2566 Affymetrix array data in rice (Oryza sativa) and intended to identify 400 endospermpreferred probes. Tissue-preferred expression patterns were confirmed by performing an additional meta-analysis of anatomical expression data comprising 219 spatial or temporal Agilent 44K array data. We then identified 299 genes that showed strong endosperm-preferred expression. The functional significance of 24 previously characterized genes was evaluated, and tissue-specific expression patterns of two genes were validated using a GUS reporter system. This second approach demonstrated that new tools are available for delivering agronomic traits in the endosperm. MapMan (mapman.gabipd.org/) analysis revealed the Metabolism and Regulation overviews associated with the process of endosperm development. In particular, the starch metabolism pathway is very closely related to that process in rice. We then constructed a regulatory network using both KEGG (www. genome.jp/ kegg/) pathway information for our candidate genes and a predicted protein–protein interaction network tool. Examination of an osbzip58-1 mutant with defects in endosperm development was combined with global transcriptome data in the network. Our results indicated that osbzip58-1 regulates the starch and sucrose metabolism pathways as well as the production of seed storage protein precursors. This new fundamental information adds to our understanding about the molecular mechanism for endosperm development in rice, and the resulting data will contribute to future studies that work to enhance the agronomic trait(s) associated with the endosperm.

Transgenic rice lines expressing maize <i>C1</i> and <i>R-S</i> regulatory genes produce various flavonoids in the endosperm

Shin, Young-Mi,Park, Hee-Jin,Yim, Sun-Duck,Baek, Nam-In,Lee, Choong-Hwan,An, Gynheung,Woo, Young-Min Blackwell Publishing Ltd 2006 PLANT BIOTECHNOLOGY JOURNAL Vol.4 No.3

<P>Summary</P><P>Flavonoids, compounds that possess diverse health-promoting benefits, are lacking in the endosperm of rice. Therefore, to develop transgenic lines that produce flavonoids, we transformed a white rice cultivar, <I>Oryza sativa japonica</I> cv. Hwa-Young, with maize <I>C1</I> and <I>R-S</I> regulatory genes. Expression of these transgenes was restricted to the endosperm using the promoter of a rice prolamin gene. The pericarp of the <I>C1</I>/<I>R-S</I> homozygous lines became dark brown in accordance with their maternal genotype, whereas the endosperm turned chalky, similar to the opaque kernel phenotype. Analysis via high-performance liquid chromatography (HPLC) revealed that numerous kinds of flavonoids were produced in these transgenic kernels. To identify individual flavonoids, the number of HPLC peaks was reduced through moderate acid hydrolysis, followed by ethyl acetate partitioning. Amongst the major flavonoids, dihydroquercetin (taxifolin), dihydroisorhamnetin (3′-<I>O</I>-methyl taxifolin) and 3′-<I>O</I>-methyl quercetin were identified through liquid chromatography/mass spectrometry/mass spectrometry and nuclear magnetic resonance analyses. Fluorescence labelling with diphenylboric acid showed that the flavonoids were highly concentrated in the cells of four to five outer endosperm layers. More importantly, a high fluorescence signal was present in the cytosol of the inner endosperm layers. However, the overall signal in the inner layers was significantly lower because starch granules and protein bodies occupied most of the cytosolic space. Our estimate of the total flavonoid content in the transgenic kernels suggests that <I>C1</I>/<I>R-S</I> rice has the potential to be developed further as a novel variety that can produce various flavonoids in its endosperm.</P>

Application of two bicistronic systems involving 2A and IRES sequences to the biosynthesis of carotenoids in rice endosperm

Ha, Sun-Hwa,Liang, Ying Shi,Jung, Harin,Ahn, Mi-Jeong,Suh, Seok-Cheol,Kweon, Soon-Jong,Kim, Dong-Hern,Kim, Young-Mi,Kim, Ju-Kon Blackwell Publishing Ltd 2010 PLANT BIOTECHNOLOGY JOURNAL Vol.8 No.8

<P>Summary</P><P>Coordination of multiple transgenes is essential for metabolic engineering of biosynthetic pathways. Here, we report the utilization of two bicistronic systems involving the 2A sequence from the foot-and-mouth disease virus and the internal ribosome entry site (IRES) sequence from the crucifer-infecting tobamovirus to the biosynthesis of carotenoids in rice endosperm. Two carotenoid biosynthetic genes, phytoene synthase (<I>Psy</I>) from <I>Capsicum</I> and carotene desaturase (<I>CrtI</I>) from <I>Pantoea,</I> were linked via either the synthetic 2A sequence that was optimized for rice codons or the IRES sequence under control of the rice globulin promoter, generating <I>PAC</I> (<I><U>P</U>sy-2<U>A</U>-<U>C</U>rtI</I>) and <I>PIC</I> (<I><U>P</U>sy-<U>I</U>RES-<U>C</U>rtI</I>) constructs, respectively. The transgenic endosperm of <I>PAC</I> rice had a more intense golden color than did <I>PIC</I> rice, demonstrating that 2A was more efficient than IRES in coordinating gene expression. The 2A and IRES constructs were equally effective in driving transgene transcription. However, immunoblot analysis of CRTI, a protein encoded by the downstream open reading frame of the bicistronic constructs, revealed that 2A was ninefold more effective than IRES in driving translation. The <I>PAC</I> endosperms accumulated an average of 1.3 &mgr;g/g of total carotenoids, which was ninefold higher than was observed for <I>PIC</I> endosperms. In particular, accumulation of &bgr;-carotene was much higher in <I>PAC</I> endosperms than in <I>PIC</I> endosperms. Collectively, these results demonstrate that both 2A and IRES systems can coordinate the expression of two biosynthetic genes, with the 2A system exhibiting greater efficiency. Thus, the 2A expression system described herein is an effective new tool for multigene stacking in crop biotechnology.</P>

FLOURY AND SHRUNKEN ENDOSPERM6 Encodes a Glycosyltransferase and is Essential for the Development of Rice Endosperm

Yang Hang 한국식물학회 2022 Journal of Plant Biology Vol.65 No.3

Composition and structure of endosperm starch are major determinant factors for rice quality. Glycosylinositol phosphorylceramides (GIPC) is a kind of sphingolipid and it accounts for ~ 25% of total plasma membrane lipids in plants. The relationship between synthesis of GIPC and development of endosperm starch, however, remains unclear. We here identified a mutant with a floury and opaque endosperm, named floury and shrunken endosperm 6 (fse6). The mutant seeds displayed a shrunken grain appearance. Physicochemical analysis showed that both total starch and amylose contents were decreased, while lipid and protein contents were increased in the mature mutant seeds, compared to their counterparts in the wild type. Further observation of semi-thin sections indicated the development of mutant amyloplasts was defective. The mutant seeds germinated normally but failed to survive in a later stage of seedling growth. Map-based cloning and genetic complementation revealed that FSE6 encodes a glycosyltransferase and is homologous to Arabidopsis GLUCOSAMINE INOSITOLPHOSPHORYLCERAMIDE TRANSFERASE1 (GINT1), an enzyme vital for GIPC synthesis. We further found that cellulose content and starch biosynthesis in the mutant were altered. This study connects a gap between a rice GINT1 and starch synthesis, which will be helpful for rice quality improvement.

Sugary Endosperm is Modulated by <i>Starch Branching Enzyme IIa</i> in Rice ( <i>Oryza sativa</i> L.)

Lee, Yunjoo,Choi, Min-Seon,Lee, Gileung,Jang, Su,Yoon, Mi-Ra,Kim, Backki,Piao, Rihua,Woo, Mi-Ok,Chin, Joong Hyoun,Koh, Hee-Jong Springer US 2017 Rice Vol.10 No.-

<P><B>Background</B></P><P>Starch biosynthesis is one of the most important pathways that determine both grain quality and yield in rice (<I>Oryza sativa</I> L.). Sugary endosperm, <I>sugary-1</I> (<I>sug-1</I>), is a mutant trait for starch biosynthesis. Rice plants carrying <I>sug-1</I> produce grains that accumulate water-soluble carbohydrates instead of starch, even after maturity. Although this trait enhances the diversity of grain quality, sugary endosperm rice has hardly been commercialized due to the severely wrinkled grains and subsequent problems in milling. This study was conducted to identify the genes responsible for the <I>sug-h</I> phenotype through a map-based cloning technology.</P><P><B>Results</B></P><P>We induced a mild sugary mutant, <I>sugary-h</I> (<I>sug-h</I>) through the chemical mutagenesis on the Korean <I>japonica</I> cultivar Hwacheong. Grains of the <I>sug-h</I> mutant were translucent and amber-colored, and the endosperm appeared less wrinkled than <I>sug-1</I>, whereas the soluble sugar content was fairly high. These characteristics confer greater marketability to the <I>sug-h</I> mutant. Genetic analyses indicated that the <I>sug-h</I> mutant phenotype was controlled by a complementary interaction of two recessive genes, <I>Isoamylase1</I> (<I>OsISA1</I>), which was reported previously, and <I>Starch branching enzyme IIa</I> (<I>OsBEIIa</I>), which was newly identified in this study. Complementation tests indicated that <I>OsBEIIa</I> regulated the properties of sugary endosperm<I>.</I></P><P><B>Conclusions</B></P><P>Complementary interactions between the starch biosynthesis genes <I>OsISA1</I> and <I>OsBEIIa</I> determine the mild sugary endosperm mutant, <I>sugary-h</I>, in rice. Our finding may facilitate the breeding of sugaryendosperm rice for commercial benefit.</P><P><B>Electronic supplementary material</B></P><P>The online version of this article (doi:10.1186/s12284-017-0172-3) contains supplementary material, which is available to authorized users.</P>

Formation of Protein Bodies and the Response to Nitrogen in Different Positions During Wheat Endosperm Development

Yang Yang,Xinyu Chen,Liping Ran,Yunfei Wu,Xurun Yu,Zhaodi Dong,Fei Xiong 한국식물학회 2019 Journal of Plant Biology Vol.62 No.4

The development of protein bodies (PBs) determinesthe processing properties of wheat (Triticum aestivum L.). Ithas been known that nitrogen uptake has a strong impact ongrain protein concentration. However, the differences of theformation of PB in different developmental stages anddifferent positions in wheat endosperm are still controversial. To solve these issues, PBs formation in different wheatendosperm parts and the response to nitrogen were investigatedusing light, transmission electron microscopes in presentstudy. The main results were as follows. (1) Proteins mainlyformed PBs via the Golgi apparatus in the vacuole at theearly stage of wheat endosperm development. After 7 dayspost anthesis, most proteins were derived from the roughendoplasmic reticulum. (2) The morphology of PBs was diverseamong different endosperm parts. Compared with the centralendosperm, the PBs in sub-aleurone cells were abundant andlarge. (3) More abundant endoplasmic reticulum, Golgi andmitochondrion were observed at the early stage after nitrogentreatment. Nitrogen also increased the accumulation of PBsat the later stage. (4) The sub-aleurone region cells of theendosperm presented more significant responses to nitrogenthan the central endosperm.

裁培種 감자 半數體와 近緣野生種間 交雜에서 endosperm balance number(EBN)의 영향

조현묵(Hyun Mook Cho),김혜영(Hei Young Kim-Lee),엄영현(Young Hyun Om),김정간(Jung Kan Kim) 한국육종학회 1997 한국육종학회지 Vol.29 No.2

The effect of Endosperm Balance Number (EBN) in interploidal and interspecific crosses between S. tuberosum dihaploids and tuber-bearing wild species was investigated. The crossability was evaluated by the percentage of berry set and the average number of seeds per berry. The percentages of berry set were 11.5%, 11.4%, 7.4% and 2.2% in 4x×4x, 4x×2x, 2x×2x and 2x×4x crosses, respectively. The number of seeds per berry was in the same order as the percentages of berry set. The crossability was greatly affected by the ratio of the maternal and parternal EBN in the endosperm. The crosses with the maternal and parternal EBN ratio of 1 : 1, 1 : 1/2 and 4 : 1 in the developing endosperm did not produce hybrid seeds. However, a lot of plump hybrid seeds were produced only in 2 : 1 of EBN ratio irrespective of differences of the species used or ploidy levels of the parents.

FLOURY ENDOSPERM12 Encoding Alanine Aminotransferase 1 Regulates Carbon and Nitrogen Metabolism in Rice

Mingsheng Zhong,Xi Liu,Feng Liu,Yulong Ren,Yunlong Wang,Jianping Zhu,Xuan Teng,Erchao Duan,Fan Wang,Huan Zhang,Mingming Wu,Yuanyuan Hao,Xiaopin Zhu,Ruonan Jing,Xiuping Guo,Ling Jiang,Yihua Wang,Jianmi 한국식물학회 2019 Journal of Plant Biology Vol.62 No.1

Starch is a major storage substance in cerealgrains, and starch biosynthesis is a complex process. In orderto elucidate regulation of the starch biosynthesis pathway, wescreened a series of rice (Oryza sativa L.) endospermmutants. In this study, we identified a floury white-coreendosperm mutant named floury endosperm12 (flo12). Theflo12 mutant exhibited loosely packed starch granules and alower thousand kernel weight compared to wild type. Semithinsections revealed that compound starch grains (SG) inflo12 interior endosperm cells were developed abnormally. Furthermore, amylose content was decreased, while totalprotein content was significantly increased in flo12 grains. Map-based cloning showed that FLO12 encodes rice alanineaminotransferase 1 (OsAlaAT1). OsAlaAT1 is highly expressedin developing endosperm. Subcellular localization showedthat OsAlaAT1 is localized in the cytosol. Moreover, theexpression of most starch synthesis-related genes wasdecreased, while most of the storage protein coding geneshad elevated expression levels in the flo12 mutant. Inaddition, overexpression of the OsAlaAT1 gene increasedgrain weight. In brief, we demonstrated that OsAlaAT1regulates carbon and nitrogen metabolism, which provides anew insight for the improvement of rice quality and yield.

Characterization of a New High-Lysine Mutant in Barley (Hordeum vulgare L.)

Hong Sik Kim,Dea Wook Kim,Sun Lim Kim,Seong Bum Baek,Hyoung Ho Park,Jong Jin Hwang,Si Ju Kim 한국육종학회 2011 한국육종학회지 Vol.43 No.5

A chemical, MNU-induced hulless barley mutant line designated as 'Mutant 98 (M98)' was developed from a Korean hulless waxy barley cultivar, 'Chalssalbori'. The objective of the study was to determine the genetic basis of 'M98' and the possibility of using 'M98' as breeding parent to improve lysine level. Compared to 'Chalssalbori', 'M98' had large embryo and higher lysine content in both the embryo and endosperm. Significantly different lysine content in 'M98' and the other high-lysine barley mutant stocks was observed for two years. However, the genotype by year interaction was not significant. 'M98' was higher than the other high-lysine barley mutant stocks in the percentage of lysine of total amino acid composition (0.75%). The trait of shrunken endosperm of 'M98', which was typical in the high-lysine mutants, was inherited by a single recessive gene. Based on seed morphology and lysine content of F1 seeds, 'M98' had a genetically different gene from the other high-lysine mutants for shrunken endosperm. Segregation of F2 for plump/shrunken endosperm did not fit the expected ratio of Mendelian inheritance except for only one cross combination (GSHO1784 (lys1)/M98). The amino acid analysis of F5 and F6 progenies from the cross between 'M98' and 'Chalssalbori' revealed that the attempt to increase the range of lysine content of plump lines did not go beyond the limit of the average high-lysine barley germplasm.