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1.
Map-based cloning of a novel rice cytochrome P450 gene CYP81A6 that confers resistance to two different classes of herbicides 总被引:2,自引:0,他引:2
Development of hybrid rice has greatly contributed to increased yields during the past three decades. Two bentazon-lethal
mutants 8077S and Norin8m are being utilized in developing new hybrid rice systems. When the male sterile lines are developed
in such a mutant background, the problem of F1 seed contamination by self-seeds from the sterile lines can be solved by spraying
bentazon at seedling stage. We first determined the sensitivity of the mutant plants to bentazon. Both mutants showed symptoms
to bentazon starting from 100 mg/l, which was about 60-fold, lower than the sensitivity threshold of their wild-type controls.
In addition, both mutants were sensitive to sulfonylurea-type herbicides. The locus for the mutant phenotype is bel for 8077S and bsl for Norin8m. Tests showed that the two loci are allelic to each other. The two genes were cloned by map-based cloning. Interestingly,
both mutant alleles had a single-base deletion, which was confirmed by PCR-RFLP. The two loci are renamed bel
a
(for bel) and bel
b
(for bsl). The wild-type Bel gene encodes a novel cytochrome P450 monooxgenase, named CYP81A6. Analysis of the mutant protein sequence also revealed the reason for bel
a
being slightly tolerant than bel
b
. Introduction of the wild-type Bel gene rescued the bentazon- and sulfonylurea-sensitive phenotype of bel
a
mutant. On the other hand, expression of antisense Bel in W6154S induced a mutant phenotype. Based on these results we conclude that the novel cytochrome P450 monooxygenase CYP81A6 encoded by Bel confers resistance to two different classes of herbicides.
Gang Pan and Xianyin Zhang contributed equally to this work. 相似文献
2.
Chao Liu Shuangqing Liu Fang Wang Yaqin Wang Kede Liu 《Plant Cell, Tissue and Organ Culture》2012,109(3):419-428
Bentazon and sulfonylurea are two different classes of herbicides that have been widely used to kill broad-leaf weeds in rice
fields. A cytochrome P450 gene, CYP81A6, encoding a monooxygenase has been previously identified to confer resistance to these two classes of herbicides in wild-type
rice. In this study, we introduced the rice CYP81A6 gene into Arabidopsis and tobacco plants to test the possibility of engineering tolerance to these two types of herbicides in other susceptible
plants. Arabidopsis and tobacco plants expressing CYP81A6 showed tolerance to both bentazon and bensulfuron-methyl (BSM), a widely applied sulfonylurea herbicide. The optimal concentrations
of bentazon and BSM for the selection of CYP81A6 transgenic plants were also determined. In addition, we also demonstrated that CYP81A6 can be used as a selection marker to effectively screen for positive transgenic Arabidopsis plants. The selection efficiency of CYP81A6 was comparable to that of the bar gene in Arabidopsis. These results suggest that CYP81A6 can not only be used to produce transgenic plants tolerant to both bentazon and sulfonylureas, but that it can also be used
as a novel plant-derived selection marker in plant transformation. 相似文献
3.
Claire Domon Jean-Luc Evrard Datta T.N. Pillay André Steinmetz 《Molecular & general genetics : MGG》1991,229(2):238-244
Summary Metabolism of sulfonylurea herbicides by Streptomyces griseolus ATCC 11796 is carried out via two cytochromes P-450, P-450SU1 and P-450SU2. Mutants of S. griseolus, selected by their reduced ability to metabolize a fluorescent sulfonylurea, do not synthesize cytochrome P-450SU1 when grown in the presence of sulfonylureas. Genetic evidence indicated that this phenotype was the result of a deletion of > 15 kb of DNA, including the structural genes for cytochrome P-450SU1 and an associated ferredoxin Fd-1 (suaC and suaB, respectively). In the absence of this monooxygenase system, the mutants described here respond to the presence of sulfonylureas or phenobarbital in the growth medium with the expression of only the suhC,B gene products (cytochrome P-450SU2 and Fd-2), previously observed only as minor components in wild-type cells treated with sulfonylurea. These strains have enabled an analysis of sulfonylurea metabolism mediated by cytochrome P-450SU2 in the absence of P-450SU1, yielding an in vivo delineation of the roles of the two different cytochrome P-450 systems in herbicide metabolism by S. griseolus. 相似文献
4.
Fine Mapping and in silico Isolation of the EUI1 Gene Controlling Upper Internode Elongation in Rice 总被引:8,自引:0,他引:8
Upper internode elongation in rice is an important agronomic trait. Well-known mutants with an elongated uppermost internode
(eui) are important germplasms for developing unsheathed-panicle male-sterile lines in hybrid rice breeding. We finely mapped
the eui1 gene and identified its candidate gene using in silico analysis based on previous research work and rice genomic sequence data. The rice eui1 gene was mapped to two overlapping BAC clones, OSJNBa0095J22 and OSJNBb0099O15, between the markers AC40 and AC46, that were
0.64 cM apart and spanned approximately 152 kb. A simple sequence repeat (SSR) marker AC41 that cosegregated with eui1 was located in an intron of a putative cytochrome P450-related gene. In silico analysis suggested that this encoded the cytochrome CYP714D1. Allelic sequencing confirmed that EUI1 corresponded to this P450 gene. A gamma ray-induced eui1 mutant carried a deletion in exon II of the EUI1 gene, and resulted in a frame-shift deletion that produced a truncated polypeptide. We conclude that the EUI1 gene controlling the upper internode elongation in rice is 9804 bp long, and comprises two exons and one intron. The length
of the cDNA is 1931 bp containing a 1734 bp ORF, a 110 bp 5′-UTR and a 87 bp 3′-UTR. The ORF encodes an unknown 577 amino
acid functional protein, that appears to be a member of the cytochrome P450 family.
Hongli Ma, Shubiao Zhang: These authors contributed equally to this work 相似文献
5.
A bentazon and sulfonylurea sensitive mutant: breeding,genetics and potential application in seed production of hybrid rice 总被引:7,自引:0,他引:7
Zhang J Xu Y Wu X Zhu L 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2002,105(1):16-22
The use of a thermosensitive genic male sterility (TGMS) system in two-line hybrid rice breeding is affected greatly by the sterility instability of TGMS lines caused by temperature fluctuation beyond their critical temperatures for fertility reversion. To prevent seed production from self contamination, we have developed a system to secure seed purity using a herbicide-sensitive TGMS mutant, M8077S, obtained by radiation. Genetic analysis, using the F1, F2 and F3 populations derived from this mutant and other normal varieties, revealed that bentazon lethality/sensitivity was controlled by a single recessive gene, which was named bel. The mutant can be killed at the seedling stage by bentazon at 300 mg/l or higher, a dosage that is safe for its F1 hybrids and all other normal varieties. This mutant is also sensitive to all the tested sulfonylurea herbicides. Response of segregating plants to these two types of herbicide indicated that sulfonylurea sensitivity was also controlled by bel. By crossing this mutant with Pei-Ai 64S, an F2 population was developed for genetic mapping. Surveying the two DNA pools from sensitive and non-sensitive F2 plants identified four markers that were polymorphic between the pools. The putative linked markers were then confirmed with the F2 population. The bel locus was located on chromosome 3, 7.1 cM from the closest microsatellite marker RM168. Phenotypic analysis indicated that the bel gene had no negative effect on agronomic traits in either a homozygous or heterozygous status. The mutant M8077S is valuable in the development of a TGMS breeding system for preventing impurity resulting from temperature fluctuation of the TGMS. Several two-line hybrid rice crosses using this system are under development. 相似文献
6.
Kawahigashi H Hirose S Ozawa K Ido Y Kojima M Ohkawa H Ohkawa Y 《Transgenic research》2005,14(6):907-917
We introduced two novel types of pig (Sus scrofa) cytochrome P450, CYP2B22 and CYP2C49, into rice plants (Oryza sativa L. cv. ‘Nipponbare’) to produce herbicide-tolerant plants and to confirm the metabolic activities of the cytochrome P450
species. In germination tests, both types of transgenic plants showed tolerance to various herbicides with different modes
of action. CYP2B22 rice plants showed tolerance towards 12 herbicides including chlortoluron (100 μM), amiprofos-methyl (2.5 μM),
pendimethalin (10 μM), metolachlor (2.5 μM), and esprocarb (20 μM). CYP2C49 rice plants showed tolerance towards 13 herbicides,
including chlortoluron (100 μM), norflurazon (0.5 μM), amiprofos-methyl (2.5 μM), alachlor (0.8 μM), and isoxaben (1 μM).
The herbicide tolerance was considered to reflect the substrate specificity of the introduced P450 species. We used 14C-labeled metolachlor and norflurazon to confirm the P450 activity in the transgenic rice plants. The herbicides were metabolized
more quickly in the transgenic rice plants than in the nontransgenic rice plants. Therefore, CYP2B22 and CYP2C49 rice plants
became more tolerant to various herbicides than nontransgenic control plants because of accelerated metabolism of the herbicides
by the introduced P450 species. Assuming that public and commercial acceptance is forthcoming, these transgenic rice plants
may become useful tools for the breeding of herbicide-tolerant crops. 相似文献
7.
To enhance our understanding of brassinosteroid (BR) biosynthesis in rice, we attempted to identify putative rice homologs
of Arabidopsis CYP90A1/ CPD and related mutants. Two candidate genes, designated CYP90A3/OsCPD1 and CYP90A4/OsCPD2, are located on chromosomes 11 (2.0 cM) and 12 (1.9 cM), respectively. Based on sequence similarity with the Arabidopsis CYP90A1/CPD gene, we predict that the CYP90A3/OsCPD1 and CYP90A4/OsCPD2 gene products function as C-23α hydroxylases in the BR biosynthesis pathway. Both are broadly expressed in wild-type rice,
and their expression is regulated by a feedback mechanism. A retrotransposon insertion mutant of CYP90A3/OsCPD1, oscpd1-1, did not produce any BR-deficient phenotype or feedback upregulation of genes for BR biosynthesis enzymes. These results
indicate that if, as predicted, the CYP90A3/OsCPD1 and CYP90A4/OsCPD2 genes do function in the BR biosynthesis pathway, they may each have enough capacity to catalyze BR biosynthesis on their
own. As a consequence, the oscpd1-1 mutant may not be deficient in endogenous BRs. Interestingly, BR biosynthesis enzymes except C-6 oxidase are encoded by plural
genes in rice but by single genes in Arabidopsis (again, except C-6 oxidase). On the basis of these findings, we discuss the differences in BR biosynthesis between rice and
Arabidopsis. 相似文献
8.
9.
Cotton plants engineered for resistance to the herbicides, glyphosate or glufosinate have made a considerable impact on the production of the crop worldwide. In this work, embryogenic cell cultures derived from Gossypium hirsutum L. cv Coker 312 hypocotyl callus were transformed via Agrobacterium tumefaciens with the rice cytochrome P450 gene, CYP81A6 (bel). In rice, bel has been shown to confer resistance to both bentazon and sulfanylurea herbicides. Transformed cells were selected on a liquid medium supplemented alternately or simultaneously with kanamycin (50mg/L) and bentazon (4.2 μmol). A total of 17 transgenic cotton lines were recovered, based on the initial resistance to bentazon and on PCR detection of the bel transgene. Bel integration into the cotton genome was confirmed by Southern blot and expression of the transgene was verified by RT-PCR. In greenhouse and experimental plot trials, herbicide (bentazon) tolerance of up to 1250 mg/L was demonstrated in the transgenic plants. Transgenic lines with a single copy of the bel gene showed normal Mendelian inheritance of the characteristic. Importantly, resistance to bentazon was shown to be stably incorporated in the T1, T2 and T3 generations of self-fertilised descendents and in plants outcrossed to another upland cotton cultivar. Engineering resistance to bentazon in cotton through the heterologous expression of bel opens the possibility of incorporating this trait into elite cultivars, a strategy that would give growers a more flexible alternative to weed management in cotton crops. 相似文献
10.
11.
Sitosterol and stigmasterol are major sterols in vascular plants. An altered stigmasterol:sitosterol ratio has been proposed
to influence the properties of cell membranes, particularly in relation to various stresses, but biosynthesis of stigmasterol
is poorly understood. Recently, however, Morikawa et al. (Plant Cell 18:1008–1022, 2006) showed in Arabidopsis thaliana that synthesis of stigmasterol and brassicasterol is catalyzed by two separate sterol C-22 desaturases, encoded by the genes
CYP710A1 and CYP710A2, respectively. The proteins belong to a small cytochrome P450 subfamily having four members, denoted by CYP710A1-A4, and
are related to the yeast sterol C-22 desaturase Erg5p acting in ergosterol synthesis. Here, we report on our parallel investigation
of the Arabidopsis CYP710A family. To elucidate the function of CYP710A proteins, transgenic Arabidopsis plants were generated overexpressing CYP710A1 and CYP710A4. Compared to wild-type plants, both types of transformant displayed a normal phenotype, but contained increased levels of
free stigmasterol and a concomitant decrease in the level of free sitosterol. CYP710A1 transformants also displayed higher
levels of esterified forms of stigmasterol, cholesterol, 24-methylcholesterol and isofucosterol. The results confirm the findings
of Morikawa et al. (Plant Cell 18:1008–1022, 2006) regarding the function of CYP710A1 in stigmasterol synthesis, and show that CYP710A4 also has this capacity. Furthermore,
our results suggest that an increased stigmasterol level alone is sufficient to stimulate esterification of other major sterols. 相似文献
12.
13.
Ohtsu K Hirano HY Tsutsumi N Hirai A Nakazono M 《Molecular genetics and genomics : MGG》2005,274(6):606-615
A new type of transposon, named Anaconda (Anac) has been found in rice (Oryza sativa). In this paper, we demonstrate that Anaconda elements have diversified by acquisition of host cellular genes, amplification of the elements, and substitution and deletion
of short segments. We identified four Anaconda elements in studies of rice alternative oxidase (AOX) genes, and subsequently isolated an additional 23 elements based on the identity of their terminal inverted repeats (TIRs).
The Anaconda elements have long TIRs (114–458 bp). They also have direct repeats of 9 or 10 bp in their flanking regions that are thought
to have been generated upon transposition. These structural features reveal that the Anaconda elements belong to the Mu superfamily. The most prominent feature of the Anaconda elements is the high frequency with which they have acquired host cellular genes. Of the 27 elements found here, 19 appear
to have sequences presumably derived from rice genes, for example, the genes for AOX1c (four elements), cytochrome P450 (five
elements), l-asparaginase (five elements), and PCF8 (two elements). Four elements, AnacA1–A4, have both the AOX1c and P450 genes. One element, AnacB14, involves a gene similar to mudrA of maize MuDR. Database analyses revealed that the loci of 26 of the 27 Anaconda elements in the subspecies japonica are the same as those in the subspecies indica. This suggests that these elements were incorporated before the divergence of these two subspecies. 相似文献
14.
Whole genome sequencing of the model white rot basidiomycete Phanerochaete chrysosporium has revealed the largest P450 contingent known to date in fungi, along with related phase I and phase II metabolic genes
and signaling cascade genes. As a part of their functional characterization, genome-wide expression profiling under physiologically
distinct conditions, nutrient-limited (ligninolytic) and nutrient-rich (non-ligninolytic), was investigated using a custom-designed
70-mer oligonucleotide microarray developed based on 190 target genes and 23 control genes. All 150 P450 genes were found
to be expressible under the test conditions, with 27 genes showing differential expression based on a >twofold arbitrary cut-off
limit. Of these, 23 P450 genes were upregulated (twofold to ninefold) in defined high-nitrogen cultures whereas four genes
were upregulated (twofold to twentyfold) in defined low-nitrogen cultures. Furthermore, tandem P450 member genes in ten of
the 16 P450 genomic clusters showed nonassortative regulation of expression reflecting their functional diversity. Full-length
cDNAs for two of the high-nitrogen upregulated genes pc-hn1 (CYP5035A1) and pc-hn2 (CYP5036A1) and partial cDNA for a low-nitrogen upregulated gene pc-ln1 (CYP5037A1) were cloned and characterized. The study provided first molecular evidence for the presence of active components
of the cAMP- and MAP kinase-signaling pathways in a white rot fungus; four of these components (cpka and ste-12 of cAMP pathway and two MAP kinases, mps1 and sps1) were significantly upregulated (fourfold to eightfold) under nutrient-limited conditions, implying their likely role in
the regulation of gene expression involved in secondary metabolism and biodegradation processes under these conditions. 相似文献
15.
Plant cytochrome P450s interact with a flavoprotein, NADPH-cytochrome P450 reductase (CPR), to transfer electrons from NADPH.
The gene for rice P450 reductase (RCPR) was cloned and expressed in Saccaromyces cerevisiae, where the specific activity of the expressed RPCR was 0.91 U/mg protein. When isoflavone synthase gene (IFS) from red clover, used as a model system of plant cytochrome P450, was co-expressed with RCPR in yeast, the production of
genistein from naringein increased about 4.3-fold, indicating that the RCPR efficiently interacts with cytochrome P450 to
transfer electrons from NADPH. 相似文献
16.
Tributyl phosphate (TBP) is a toxic organophosphorous compound widely used in nuclear fuel processing and chemical industries.
Rhodopseudomonas palustris, one of the most metabolically versatile photosynthetic bacteria, is shown here to degrade TBP efficiently under photosynthetic
conditions. This study shows that this O2- and NADPH/FMNH2-dependent process was also catalyzed when TBP was incubated with membrane-associated proteins extracted from this strain.
The effects of several regulators of cytochrome P450 activity on the TBP consumption suggest a key role for a cytochrome P450 in this process. Disruption of the rpa0241 gene encoding a putative cytochrome P450 led to a 60% decrease of the TBP catabolism, whereas reintroducing the gene in the mutant restored the wild-type phenotype.
The rpa0241 gene was expressed and purified in Escherichia coli. Characterization by UV-visible spectroscopy of the purified recombinant membrane-bound protein (CYP201A2) encoded by the
rpa0241 gene revealed typical spectral characteristics of cytochrome P450 with a large spin state change of the heme iron associated with binding of TBP (K
d ≈ 65 μM). It is proposed that CYP201A2 catalyzes the initial step of the biodegradation process of TBP. 相似文献
17.
Heping Han Qin Yu Roland Beffa Susana González Frank Maiwald Jian Wang Stephen B. Powles 《The Plant journal : for cell and molecular biology》2021,105(1):79-92
Rapid and widespread evolution of multiple herbicide resistance in global weed species endowed by increased capacity to metabolize (degrade) herbicides (metabolic resistance) is a great threat to herbicide sustainability and global food production. Metabolic resistance in the economically damaging crop weed species Lolium rigidum is well known but a molecular understanding has been lacking. We purified a metabolic resistant (R) subset from a field evolved R L. rigidum population. The R, the herbicide susceptible (S) and derived F2 populations were used for candidate herbicide resistance gene discovery by RNA sequencing. A P450 gene CYP81A10v7 was identified with higher expression in R vs. S plants. Transgenic rice overexpressing this Lolium CYP81A10v7 gene became highly resistant to acetyl-coenzyme A carboxylase- and acetolactate synthase-inhibiting herbicides (diclofop-methyl, tralkoxydim, chlorsulfuron) and moderately resistant to hydroxyphenylpyruvate dioxygenase-inhibiting herbicide (mesotrione), photosystem II-inhibiting herbicides (atrazine and chlorotoluron) and the tubulin-inhibiting herbicide trifluralin. This wide cross-resistance profile to many dissimilar herbicides in CYP81A10v7 transgenic rice generally reflects what is evident in the R L. rigidum. This report clearly showed that a single P450 gene in a cross-pollinated weed species L. rigidum confers resistance to herbicides of at least five modes of action across seven herbicide chemistries. 相似文献
18.
Cytochrome P450 (P450) open reading frames (ORFs) identified in genome sequences of Bacillus species are potential resources for new oxidation biocatalysts. Phylogenetic analysis of 29 Bacillus P450 ORFs revealed that the P450s consist of a limited number of P450 families, CYP102, CYP106, CYP107, CYP109, CYP134, CYP152, and CYP197. Previously, we identified the catalytic activities of three P450s of Bacillus subtilis towards steroids by rapid substrate screening using Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR/MS). Here, we further applied this method to evaluate the activity of Bacillus cereus P450s towards steroids. Five P450 genes were cloned from B. cereus ATCC 10987 based on its genomic sequence and were expressed in Escherichia coli. These P450s were reacted with a mixture of 30 compounds that mainly included steroids, and the reaction mixtures were analyzed using FT-ICR/MS. We found that BCE_2659 (CYP106) catalyzed the monooxygenation of methyltestosterone, progesterone, 11-ketoprogesterone, medroxyprogesterone acetate, and chlormadinone acetate. BCE_2654 (CYP107) monooxygenated testosterone enanthate, and BCE_3250 (CYP109) monooxygenated testosterone and compactin. Based on the phylogenetic relationship and the known substrate specificities including ones identified in this study, we discuss the catalytic potential of Bacillus P450s towards steroids. 相似文献
19.
CHARACTERIZATION AND EXPRESSION PROFILES OF FIVE POSSIBLE CYTOCHROME P450 GENES FROM Liposcelis entomophila (ENDERLEIN) (PSOCOPTERA: LIPOSCELIDIDAE)
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Ting Li Yan Liu Dan‐Dan Wei Feng Shang Guy Smagghe Wei Dou Jin‐Jun Wang Guy Smagghe 《Archives of insect biochemistry and physiology》2016,92(4):259-273
20.
The final reactions of rosmarinic acid biosynthesis, the introduction of the aromatic 3- and 3′-hydroxyl groups, are catalysed
by cytochrome P450-dependent hydroxylases. The cDNAs encoding CYP98A14 as well as a NADPH:cytochrome P450 reductase (CPR)
were isolated from Coleus blumei and actively expressed in Saccharomyces cerevisiae. The CYP98A14-cDNA showed an open reading frame of 1521 nucleotides with high similarities to 4-coumaroylshikimate/quinate
3-hydroxylases. Yeast microsomes harbouring the CYP98A14 protein catalysed the 3-hydroxylation of 4-coumaroyl-3′,4′-dihydroxyphenyllactate
and the 3′-hydroxylation of caffeoyl-4′-hydroxyphenyllactate, in both cases forming rosmarinic acid. Apparent K
m-values for 4-coumaroyl-3′,4′-dihydroxyphenyllactate and caffeoyl-4′-hydroxyphenyllactate were determined to be at 5 μM and
40 μM, respectively. CYP98A14 differs from CYP98s from other plants, since 4-coumaroylshikimate or -quinate were not accepted
as substrates. Coexpression of the Coleus blumei CPR and CYP98A14 in the same yeast cells increased the hydroxylation activity up to sevenfold. CYP98A14 from Coleus blumei is a novel bifunctional cytochrome P450 specialised for rosmarinic acid biosynthesis. 相似文献