Dataset Information

MITE insertion-dependent expression of CitRKD1 with a RWP-RK domain regulates somatic embryogenesis in citrus nucellar tissues.

ABSTRACT: BACKGROUND:Somatic embryogenesis in nucellar tissues is widely recognized to induce polyembryony in major citrus varieties such as sweet oranges, satsuma mandarins and lemons. This capability for apomixis is attractive in agricultural production systems using hybrid seeds, and many studies have been performed to elucidate the molecular mechanisms of various types of apomixis. To identify the gene responsible for somatic embryogenesis in citrus, a custom oligo-DNA microarray including predicted genes in the citrus polyembryonic locus was used to compare the expression profiles in reproductive tissues between monoembryonic and polyembryonic varieties. The full length of CitRKD1, which was identified as a candidate gene responsible for citrus somatic embryogenesis, was isolated from satsuma mandarin and its molecular function was investigated using transgenic 'Hamlin' sweet orange by antisense-overexpression. RESULTS:The candidate gene CitRKD1, predominantly transcribed in reproductive tissues of polyembryonic varieties, is a member of the plant RWP-RK domain-containing protein. CitRKD1 of satsuma mandarin comprised two alleles (CitRKD1-mg1 and CitRKD1-mg2) at the polyembryonic locus controlling embryonic type (mono/polyembryony) that were structurally divided into two types with or without a miniature inverted-repeat transposable element (MITE)-like insertion in the upstream region. CitRKD1-mg2 with the MITE insertion was the predominant transcript in flowers and young fruits where somatic embryogenesis of nucellar cells occurred. Loss of CitRKD1 function by antisense-overexpression abolished somatic embryogenesis in transgenic sweet orange and the transgenic T1 plants were confirmed to derive from zygotic embryos produced by self-pollination by DNA diagnosis. Genotyping PCR analysis of 95 citrus traditional and breeding varieties revealed that the CitRKD1 allele with the MITE insertion (polyembryonic allele) was dominant and major citrus varieties with the polyembryonic allele produced polyembryonic seeds. CONCLUSION:CitRKD1 at the polyembryonic locus plays a principal role in regulating citrus somatic embryogenesis. CitRKD1 comprised multiple alleles that were divided into two types, polyembryonic alleles with a MITE insertion in the upstream region and monoembryonic alleles without it. CitRKD1 was transcribed in reproductive tissues of polyembryonic varieties with the polyembryonic allele. The MITE insertion in the upstream region of CitRKD1 might be involved in regulating the transcription of CitRKD1.

SUBMITTER: Shimada T

PROVIDER: S-EPMC6090715 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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MITE insertion-dependent expression of CitRKD1 with a RWP-RK domain regulates somatic embryogenesis in citrus nucellar tissues.

Shimada Takehiko T Endo Tomoko T Fujii Hiroshi H Nakano Michiharu M Sugiyama Aiko A Daido Genya G Ohta Satoshi S Yoshioka Terutaka T Omura Mitsuo M

BMC plant biology 20180813 1

<h4>Background</h4>Somatic embryogenesis in nucellar tissues is widely recognized to induce polyembryony in major citrus varieties such as sweet oranges, satsuma mandarins and lemons. This capability for apomixis is attractive in agricultural production systems using hybrid seeds, and many studies have been performed to elucidate the molecular mechanisms of various types of apomixis. To identify the gene responsible for somatic embryogenesis in citrus, a custom oligo-DNA microarray including pre ...[more]

PMID: 30103701

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Project description:Somatic embryogenesis in nucellar tissues is widely recognized to induce polyembryony in major citrus varieties such as sweet oranges, satsuma mandarins and lemons. This capability for apomixis is attractive in agricultural production systems using hybrid seeds, and many studies have been performed to elucidate the molecular mechanisms of various types of apomixis. To identify the gene responsible for somatic embryogenesis in citrus, a custom oligo-DNA microarray including predicted genes in the citrus polyembryonic locus was used to compare the expression profiles in reproductive tissues between monoembryonic and polyembryonic varieties. The full length of CitRKD1, which was identified as a candidate gene responsible for citrus somatic embryogenesis, was isolated from satsuma mandarin and its molecular function was investigated using transgenic ‘Hamlin’ sweet orange by antisense-overexpression. The candidate gene CitRKD1, predominantly transcribed in reproductive tissues of polyembryonic varieties, is a member of the plant RWP-RK domain proteins. CitRKD1 of satsuma mandarin comprised two alleles (CitRKD1-mg1 and CitRKD1-mg2) at the polyembryonic locus controlling embryony type (mono/polyembryony) that were structurally divided into two types with or without a miniature inverted-repeat transposable element (MITE)-like insertion in the upstream region. CitRKD1-mg2 with the MITE insertion was the predominant transcript in flowers and young fruits where somatic embryogenesis of nucellar cells occurred. Loss of CitRKD1 function by antisense-overexpression abolished somatic embryogenesis in transgenic sweet orange and the transgenic T1 plants were confirmed to derive from zygotic embryos produced by self-pollination by DNA diagnosis. Genotyping PCR analysis of 95 citrus traditional and breeding varieties revealed that the CitRKD1 allele with the MITE insertion (polyembryonic allele) was dominant and major citrus varieties with the polyembryonic allele produced polyembryonic seeds.

Project description:RWP-RKs represent a small family of transcription factors (TFs) that are unique to plants and function particularly under conditions of nitrogen starvation. These RWP-RKs have been classified in two sub-families, NLPs (NIN-like proteins) and RKDs (RWP-RK domain proteins). NLPs regulate tissue-specific expression of genes involved in nitrogen use efficiency (NUE) and RKDs regulate expression of genes involved in gametogenesis/embryogenesis. During the present study, using in silico approach, 37 wheat RWP-RK genes were identified, which included 18 TaNLPs (2865 to 7340 bp with 4/5 exons), distributed on 15 chromosomes from 5 homoeologous groups (with two genes each on 4B,4D and 5A) and 19 TaRKDs (1064 to 5768 bp with 1 to 6 exons) distributed on 12 chromosomes from 4 homoeologous groups (except groups 1, 4 and 5); 2-3 splice variants were also available in 9 of the 37 genes. Sixteen (16) of these genes also carried 24 SSRs (simple sequence repeats), while 11 genes had targets for 13 different miRNAs. At the protein level, MD simulation analysis suggested their interaction with nitrate-ions. Significant differences were observed in the expression of only two (TaNLP1 and TaNLP2) of the nine representative genes that were used for in silico expression analysis under varying levels of N at post-anthesis stage (data for other genes was not available for in silico expression analysis). Differences in expression were also observed during qRT-PCR, when expression of four representative genes (TaNLP2, TaNLP7, TaRKD6 and TaRKD9) was examined in roots and shoots of seedlings (under different conditions of N supply) in two contrasting genotypes which differed in NUE (C306 with low NUE and HUW468 with high NUE). These four genes for qRT-PCR were selected on the basis of previous literature, level of homology and the level of expression (in silico study). In particular, the TaNLP7 gene showed significant up-regulation in the roots and shoots of HUW468 (with higher NUE) during N-starvation; this gene has already been characterized in Arabidopsis and tobacco, and is known to be involved in nitrate-signal transduction pathway.

Dataset Information

MITE insertion-dependent expression of CitRKD1 with a RWP-RK domain regulates somatic embryogenesis in citrus nucellar tissues.

Publications

MITE insertion-dependent expression of CitRKD1 with a RWP-RK domain regulates somatic embryogenesis in citrus nucellar tissues.

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