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Differential expression of certain genes during somatic embryogenesis in leaf explants of Cichorium intybus

ABSTRACT: In Cichorium intybus, macroscopic and histological modifications were identified during in vitro cell reactivation and morphogenesis events. The relationships between these modifications and transcript profiles of genes were investigated. Two chicory genotypes derived from the Hungarian landrace Koospol, were tested: K59 is a responsive genotype (embryogenic), as it undergoes cell de- and re-differentiation that leads to somatic embryogenesis (SE). C15 is an unresponsive genotype (nonembryogenic), unable to express this morphogenetic pattern. Previously studies showed that addition of beta-D-glucosyl Yariv reagent (BGlcY), a synthetic phenyl-glycoside that specifically binds arabinogalactan-proteins (AGPs), to the culture medium blocked somatic embryogenesis in chicory root explants in a concentration-dependent manner with complete inhibition of induction occurring at 250 µM of BGlcY (Chapmann et al. 2000a). A putative AGP (DT212818) encoding gene was previously found significantly over-expressed in the K59 as compared to the C15 and was suggested to be involved in chicory re-differentiation. To better understand the cell reactivation events, a treatment with BGlcY was applied in order to precipitate AGPs in muro and to block the in vitro cell re-differentiation processes. The two different genotypes together with BGlcY represented an original tool to discriminate cell reactivation events from cell fate determination. Microscopy, biochemical and transcriptome analyses showed that in vitro SE in chicory was blocked during BGlcY-treatment but cell reactivation still occurred. Consequently, AGP (DT212818) seems to be only important in cell fate determination leading to SE commitment. Gene expression profiles showed in addition to the AGP, other proteins could play a role in SE determination: 26S proteasome AAA ATPase subunit 6 (RPT6), remorin (REM), metallothionein 1 (MT1), 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase). Cell wall events during BGlcY-treatment were observed and discussed.

ORGANISM(S): Cichorium intybus

PROVIDER: GSE15502 | GEO | 2009/04/06

SECONDARY ACCESSION(S): PRJNA115669

REPOSITORIES: GEO

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Project description:In Cichorium intybus, macroscopic and histological modifications were identified during in vitro cell reactivation and morphogenesis events. The relationships between these modifications and transcript profiles of genes were investigated. Two chicory genotypes derived from the Hungarian landrace Koospol, were tested: K59 is a responsive genotype (embryogenic), as it undergoes cell de- and re-differentiation that leads to somatic embryogenesis (SE). C15 is an unresponsive genotype (nonembryogenic), unable to express this morphogenetic pattern. Previously studies showed that addition of beta-D-glucosyl Yariv reagent (BGlcY), a synthetic phenyl-glycoside that specifically binds arabinogalactan-proteins (AGPs), to the culture medium blocked somatic embryogenesis in chicory root explants in a concentration-dependent manner with complete inhibition of induction occurring at 250 ÂµM of BGlcY (Chapmann et al. 2000a). A putative AGP (DT212818) encoding gene was previously found significantly over-expressed in the K59 as compared to the C15 and was suggested to be involved in chicory re-differentiation. To better understand the cell reactivation events, a treatment with BGlcY was applied in order to precipitate AGPs in muro and to block the in vitro cell re-differentiation processes. The two different genotypes together with BGlcY represented an original tool to discriminate cell reactivation events from cell fate determination. Microscopy, biochemical and transcriptome analyses showed that in vitro SE in chicory was blocked during BGlcY-treatment but cell reactivation still occurred. Consequently, AGP (DT212818) seems to be only important in cell fate determination leading to SE commitment. Gene expression profiles showed in addition to the AGP, other proteins could play a role in SE determination: 26S proteasome AAA ATPase subunit 6 (RPT6), remorin (REM), metallothionein 1 (MT1), 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase). Cell wall events during BGlcY-treatment were observed and discussed. Gene expression of K59 explants induced (d4) was compared with gene expression of K59 explants without induction (d0). Gene expression of K59 explants induced in the presence of BGlcY (Y+; d4) was next compared with gene expression of K59 explants induced in the absence of BGlcY (Y-; d4). Similar comparisons were made for C15 leaf explants. To link up both genotypes, direct comparison was done between gene expression of K59 explants without induction (d0) and C15 explants without induction (d0). Three biological replicates were used for each analysed condition. Dye labelling for each paired sample was reversed in two subsequent individual hybridizations giving rise to a total of six hybridizations per condition except for the K59_D0_vs_C15_D0 condition.

Project description:The Mongolian horse has the characteristics of cold resistance, rough feeding resistance, good endurance, strong disease resistance and strong gregariousness. The normal production of sperm in male mammals is the basis of reproduction, and spermatogenesis includes a mitosis, meiosis twice and metamorphosis of sperm cells. The MEI1 gene is involved in the meiosis cycle and is required for normal meiosis chromosome association. Previous studies have shown that alternative splicing of MEI1 gene can promote spermatogenesis in Mongolian horses. In this study, the expression of MEI1 gene was determined by immunofluorescence in the third generation Mongolian horse testicular sertoli cells. Two overexpressed lentiviral vectors with MXE and SE events of MEI1 gene were constructed and successfully infected into sertoli cells. CCK-8 was used to detect the proliferation and activity of infected cells, transcripomics and metabolomics were used to screen and annotate differential genes and differential metabolites in the two sertoli cells with MXE and SE events. After 72 h of virus infection, 193 differentially expressed genes and 11360 differentially expressed metabolites were screened, 109 up-regulated genes including MEI1 and 84 down-regulated genes, 7494 up-regulated metabolites and 3866 down-regulated metabolites. Differential genes and differential metabolites were mainly enriched in several pathways related to spermatogenesis. Differential genes such as IL31RA, ATP2B3, CASQ2 were highly expressed in SE events, IL11, PRLR, CCR7 were highly expressed in MXE events. Metabolites such as folic acid and spermine were highly expressed in SE events, citric acid and glutathione were highly expressed in MXE events. This suggested that both MXE and SE events of MEI1 gene can promote the activity of spermatogenesis signaling pathway, and the gene influences spermatogenesis by regulating the expression of metabolites. In summary, MXE and SE events of MEI1 gene can promote the expression of spermatogen-related genes and metabolites, and genes affect spermatogenesis by regulating the expression of metabolites. The results of this study laid a theoretical foundation for exploring the regulatory mechanism of different alternative splicing events on spermatogenesis in Mongolian horse.

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Differential expression of certain genes during somatic embryogenesis in leaf explants of Cichorium intybus

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