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An SC35-like protein and a novel serine/arginine-rich protein interact with Arabidopsis U1-70K protein

JOURNAL OF BIOLOGICAL CHEMISTRY 1999年第51期274卷 36428-36438页

作者： Golovkin, M Reddy, ASN Colorado State Univ Dept Biol Ft Collins CO 80523 USA Colorado State Univ Cell & Mol Biol Program Ft Collins CO 80523 USA

The U1 small nuclear ribonucleoprotein 70-kDa protein, a U1 small nuclear ribonucleoprotein-specific protein, has been shown to have multiple roles in nuclear precursor mRNA processing in animals. By using the C-terminal arginine-rich region of Arabidopsis U1-'IOII protein in the yeast two-hybrid system, we have identified an SC35-like (SR33) and a novel plant serine/arginine-rich (SR) protein (SR45) that interact with the plant U1-70K The SR33 and SR45 proteins share several features with SR proteins including modular domains typical of splicing factors in the SR family of proteins. However, both plant SR proteins are rich in proline, and SR45, unlike most animal SR proteins, has two distinct arginine/serine-rich domains separated by an RNA recognition motif. By using coprecipitation assays we confirmed the interaction of plant U1-70K with SR33 and SR45 proteins. Furthermore, in vivo and in vitro protein-protein interaction experiments have shown that SR33 protein interacts with itself and with SR45 protein but not with two other members (SRZ21 and SRZ22) of the SR family that are known to interact with the Arabidopsis full-length U-70H only. A Clk/Sty protein kinase (AFC-8) from Arabidopsis phosphorylated four SR proteins (SR33, SR45, SRZ21, and SRZ22). Coprecipitation studies have confirmed the interaction of SR proteins with AFC2 kinase, and the interaction between AFC2 and SR33 is modulated by the phosphorylation status of these proteins. These and our previous results suggest that the plant U1-70B: interacts with at least four distinct members of the SR family including SR45 with its two arginine/serine-rich domains, and the interaction between the SR proteins and AFC2 is modulated by phosphorylation. The interaction of plant U1-70K with a novel set of proteins suggests the early stages of spliceosome assembly, and intron recognition in plants is likely to be different from animals.

关键词：氨基酸序列拟南芥属拟南芥蛋白质类/遗传学拟南芥蛋白质类/代谢精氨酸碱基序列分子序列数据核蛋白质类/遗传学核蛋白质类/代谢 RNA剪接 RNA 信使/代谢核糖核蛋白 U1小核/代谢核糖核蛋白类酿酒酵母蛋白质类序列比对丝氨酸

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The GRAS gene family in Arabidopsis:: sequence characterization and basic expression analysis of the SCARECROW-LIKE genes

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PLANT JOURNAL 1999年第1期18卷 111-119页

作者： Pysh, LD Wysocka-Diller, JW Camilleri, C Bouchez, D Benfey, PN NYU Dept Biol New York NY 10003 USA INRA Biol Cellulaire Lab F-78026 Versailles France

Mutations at the SCARECROW(SCR) locus in Arabidopsis thaliana result in defective radial patterning in the root and shoot. The SCR gene product contains sequences which suggest that it is a transcription factor. A number of Arabidopsis Expressed Sequence Tags (ESTs) have been identified that encode gene products bearing remarkable similarity to SCR throughout their carboxyl-termini, indicating that SCR is the prototype of a novel gene family. These ESTs have been designated SCARECROW-LIKE (SCL). The gene products of the GIBBERELLIN-INSENSITIVE (GAI) and the REPRESSOR of ga1-3 (RGA) loci show high structural and sequence similarity to SCR and the SCLs. Sequence analysis of the products of the GRAS (GAI, RGA, SCR) gene family indicates that they share a variable amino-terminus and a highly conserved carboxyl-terminus that contains five recognizable motifs. The SCLs have distinct patterns of expression, but all of those analyzed show expression in the root. One of them, SCL3, has a tissue-specific pattern of expression in the root similar to SCR. The importance of the GRAS gene family in plant biology has been established by the functional analyses of SCR, GAI and RGA.

关键词：氨基酸序列拟南芥属/遗传学拟南芥蛋白质类/遗传学辅阻遏蛋白质类/遗传学保守序列进化分子基因表达基因植物原位杂交分子序列数据多基因族突变序列同源性氨基酸

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Functional cloning of an Arabidopsis thaliana cDNA encoding cycloeucalenol cycloisomerase

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JOURNAL OF BIOLOGICAL CHEMISTRY 2000年第18期275卷 13394-13397页

作者： Lovato, MA Hart, EA Segura, MJR Giner, JL Matsuda, SPT Rice Univ Dept Chem Houston TX 77005 USA Rice Univ Dept Biochem & Cell Biol Houston TX 77005 USA SUNY Dept Chem Syracuse NY 13210 USA

Plants and certain protists use cycloeucalenol cycloisomerase (EC 5.5.1.9) to convert pentacyclic cyclopropyl sterols to conventional tetracyclic sterols. We used a novel complementation strategy to clone a cycloeucalenol cycloisomerase cDNA. Expressing an Arabidopsis thaliana cycloartenol synthase cDNA in a yeast lanosterol synthase mutant provided a sterol auxotroph that could be genetically complemented with the isomerase, We transformed this yeast strain with an Arabidopsis yeast expression library and selected sterol prototrophs to obtain a strain that accumulated biosynthetic ergosterol. The novel phenotype was conferred by an Arabidopsis cDNA that potentially encodes a 36-kDa protein. We expressed this cDNA (CP11) in Escherichia coli and showed by gas chromatography-mass spectrometry that extracts from this strain isomerized cycloeucalenol to obtusifoliol in vitro. The cDNA will be useful for obtaining heterologously expressed protein for catalytic studies and elucidating the in vivo roles of cyclopropyl sterols.

关键词：氨基酸序列拟南芥属/遗传学拟南芥蛋白质类/遗传学碱基序列克隆分子 DNA 互补/遗传学 DNA 互补/分离和提纯大肠杆菌分子内裂合酶类/遗传学异构酶类/遗传学分子序列数据

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HRT gene function requires interaction between a NAC protein and viral capsid protein to confer resistance to turnip crinkle virus

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PLANT CELL 2000年第10期12卷 1917-1925页

作者： Ren, T Qu, F Morris, TJ Univ Nebraska Sch Biol Sci Lincoln NE 68588 USA

An Arabidopsis protein was found to interact specifically with the capsid protein (CP) of turnip crinkle virus (TCV) through yeast two-hybrid screening. This protein, designated TIP (for TCV-interacting protein), was found to be a member of the recently recognized NAC family of proteins. NAC proteins have been implicated in the regulation of development of plant embryos and flowers. TIP alone was able to activate expression of reporter genes in yeast if fused to a DNA binding domain, suggesting that it may be a transcriptional activator. The TIP binding region in the TCV CP has been mapped to the N-terminal 25 amino acids. Site-directed mutagenesis within this region revealed that loss of the TIP-CP interaction in the yeast two-hybrid assay correlated with loss of the ability of TCV to induce the hypersensitive response and resistance in the TCV-resistant Arabidopsis ecotype Dijon (Di-0 and its inbred line Di-17). These data suggest that TIP is an essential component in the TCV resistance response pathway.

关键词：氨基酸序列拟南芥属/遗传学拟南芥属/生理学拟南芥属/病毒学拟南芥蛋白质类/遗传学拟南芥蛋白质类/代谢细菌蛋白质类碱基序列病毒壳体/代谢 DNA结合蛋白质类/遗传学 DNA结合蛋白质类/代谢分子序列数据植物蛋白质类/遗传学植物蛋白质类/代谢聚合酶链反应阻遏蛋白质类/遗传学阻遏蛋白质类/代谢逆转录聚合酶链反应序列比对序列同源性氨基酸转录因子/遗传学转录因子/代谢

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Convergent pathways for lipochitooligosaccharide and auxin signaling in tobacco cells

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PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 1996年第23期93卷 13389-13392页

作者： Rohrig, H Schmidt, J Walden, R Czaja, I Lubenow, H Wieneke, U Schell, J John, M Max-Planck-Institut für Züchtungsforschung Carl-von-Linné-Weg 10 D-50829 Köln Germany.

Lipochitooligosaccharides (LCOs) are a novel class of plant growth regulators that activate in tobacco protoplasts the expression of AXI1, a gene implicated in auxin signaling. Transient assays with a chimeric P-AXI-GUS expression plasmid revealed that the N-octadecenoylated monosaccharide GlcN has all structural requirements for a biological active glycolipid, whereas the inactive N-acylated GalN epimer inhibits LCO action. Specific inhibition of LCO and auxin action shows that both signals are transduced within the tobacco cell via separate pathways that converge at or before AXI1 transcription. Cytokinin is suggested to be a common effector of LCO and auxin signaling. We also show that activation of AXI1 correlates with growth factor-induced cell division.

关键词：拟南芥蛋白质类/遗传学印迹法 RNA 细胞分裂/药物作用细胞分裂/遗传学细胞培养的细胞激肽类/药理学剂量效应关系药物药物协同作用基因表达调控植物/药物作用葡糖醛酸糖苷酶/遗传学吲哚乙酸类/药理学脂多糖类/化学脂多糖类/药理学模型遗传学植物生长调节物/化学植物生长调节物/药理学启动区遗传/遗传学原生质体/细胞学原生质体/药物作用原生质体/代谢重组融合蛋白质类/遗传学重组融合蛋白质类/代谢信号传导/遗传学烟草/细胞学转录遗传/药物作用转染

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Five geranylgeranyl diphosphate synthases expressed in different organs are localized into three subcellular compartments in Arabidopsis

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PLANT PHYSIOLOGY 2000年第4期122卷 1045-1056页

作者： Okada, K Saito, T Nakagawa, T Kawamukai, M Kamiya, Y RIKEN Inst Phys & Chem Res Frontier Res Program Plant Hormone Funct Wako Saitama 3510198 Japan Shimane Univ Dept Appl Biosci & Biotechnol Matsue Shimane 6908504 Japan Shimane Univ Res Inst Mol Genet Matsue Shimane 6908504 Japan

Geranylgeranyl diphosphate (GGPP) is the precursor for the biosynthesis of gibberellins, carotenoids, chlorophylls, isoprenoid quinones, and geranylgeranylated proteins in plants. There is a small gene family for GGPP synthases encoding five isozymes and one related protein in Arabidopsis, and all homologs have a putative localization signal to translocate into specific subcellular compartments. Using a synthetic green fluorescent protein (sGFP), we studied the subcellular localization of these GGPP synthases. When these fusion proteins were expressed by the cauliflower mosaic virus 35S promoter in Arabidopsis, GGPS1-sGFP and GGPS3-sGFP proteins were translocated into the chloroplast, GGPS2-sGFP and GGPS4-sGFP proteins were localized in the endoplasmic reticulum, and the GGPS6-sGFP protein was localized in the mitochondria, Both GGPS1 and GGPS3 proteins synthesized in vitro were taken up into isolated intact pea chloroplasts and processed to the mature form. RNA-blot and promoter-beta-glucuronidase (GUS) analysis showed that these GGPP synthases genes are organ-specifically expressed in Arabidopsis. GGR and GGPS1 were ubiquitously expressed, while GGPS2, GGPS3, and GGPS4 were expressed specifically in the flower, root, and flower, respectively. These results suggest that each GGPP synthase gene is expressed in different tissues during plant development and GGPP is synthesized by the organelles themselves rather than being transported into the organelles. Therefore, we predict there will be specific pathways of GGPP production in each organelle.

关键词：烷基和芳基转移酶类/化学烷基和芳基转移酶类/遗传学烷基和芳基转移酶类/代谢氨基酸序列拟南芥蛋白质类/化学拟南芥蛋白质类/遗传学拟南芥蛋白质类/代谢碱基序列细胞区室化 DNA引物法尼基转转移酶同工酶类/化学同工酶类/遗传学同工酶类/代谢分子序列数据启动区遗传 RNA 信使/遗传学 RNA 信使/代谢序列同源性氨基酸

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