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2.
Front Plant Sci ; 11: 1034, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32733523

RESUMO

Brassinosteroids (BRs) as a class of steroid plant hormones participate in the regulation of numerous developmental processes, including root and shoot growth, vascular differentiation, fertility, flowering, and seed germination, as well as in responding to environmental stresses. During four decades of research, the BR biosynthetic pathways have been well studied with forward- and reverse genetics approaches. The free BRs contain 27, 28, and 29 carbons within their skeletal structure: (1): 5α-cholestane or 26-nor-24α-methyl-5α-cholestane for C27-BRs; (2) 24α-methyl-5α-cholestane, 24ß-methyl-5α-cholestane or 24-methylene-5α-cholestane for C28-BRs; (3) 24α-ethyl-5α-cholestane, 24(Z)-ethylidene-5α-cholestane, 25-methyl-5α-campestane or 24-methylene-25-methyl-5α-cholestane for C29-BRs, as well as different kinds and orientations of oxygenated functions in A- and B-ring. These alkyl substituents are also common structural features of sterols. BRs are derived from sterols carrying the same side chain. The C27-BRs without substituent at C-24 are biosynthesized from cholesterol. The C28-BRs carrying either an α-methyl, ß-methyl, or methylene group are derived from campesterol, 24-epicampesterol or 24-methylenecholesterol, respectively. The C29-BRs with an α-ethyl group are produced from sitosterol. Furthermore, the C29 BRs carrying methylene at C-24 and an additional methyl group at C-25 are derived from 24-methylene-25-methylcholesterol. Generally, BRs are biosynthesized via cycloartenol and cycloartanol dependent pathways. Till now, more than 17 compounds were characterized as inhibitors of the BR biosynthesis. For nine of the inhibitors (e.g., brassinazole and YCZ-18) a specific target reaction within the BR biosynthetic pathway has been identified. Therefore, the review highlights comprehensively recent advances in our understanding of the BR biosynthesis, sterol precursors, and dependencies between the C27-C28 and C28-C29 pathways.

3.
Int J Mol Sci ; 21(14)2020 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-32707671

RESUMO

The roles of endogenous brassinosteroids (BRs) in the modulation of reaction to drought and genetic regulation of this process are still obscure. In this study, a multidirectional analysis was performed on semi-dwarf barley (Hordeum vulgare) Near-Isogenic Lines (NILs) and the reference cultivar "Bowman" to get insights into various aspects of metabolic reaction to drought. The NILs are defective in BR biosynthesis or signaling and displayed an enhanced tolerance to drought. The BR metabolism perturbations affected the glucose and fructose accumulation under the control and stress conditions. The BR metabolism abnormalities negatively affected the sucrose accumulation as well. However, during drought, the BR-deficient NILs accumulated higher contents of sucrose than the "Bowman" cultivar. Under the control conditions, accumulation of transcripts encoding antioxidant enzymes ascorbate peroxidase (HvAPX) and superoxide dismutase (HvSOD) was BR-dependent. However, during drought, the accumulation of HvAPX transcript was BR-dependent, whereas accumulations of transcripts encoding catalase (HvCAT) and HvSOD were not affected by the BR metabolism perturbations. The obtained results reveal a significant role of BRs in regulation of the HvAPX and HvCAT enzymatic activities under control conditions and the HvAPX and HvSOD activities during physiological reactions to drought.


Assuntos
Brassinosteroides/metabolismo , Hordeum/genética , Hordeum/metabolismo , Ascorbato Peroxidases/genética , Ascorbato Peroxidases/metabolismo , Catalase/genética , Catalase/metabolismo , Secas , Frutose/metabolismo , Genes de Plantas , Glucose/metabolismo , Hordeum/crescimento & desenvolvimento , Peróxido de Hidrogênio/metabolismo , Mutação , Reguladores de Crescimento de Plantas/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Ribulose-Bifosfato Carboxilase/genética , Ribulose-Bifosfato Carboxilase/metabolismo , Estresse Fisiológico/genética , Estresse Fisiológico/fisiologia , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismo
4.
Cells ; 9(5)2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32370052

RESUMO

(1) Background: The study characterized barley mutants with brassinosteroid (BR) biosynthesis and signaling disturbances in terms of the physicochemical/structural properties of membranes to enrich the knowledge about the role of brassinosteroids for lipid metabolism and membrane functioning. (2) Methods: The Langmuir method was used to investigate the properties of the physicochemical membranes. Langmuir monolayers were formed from the lipid fractions isolated from the plants growing at 20 °C and then acclimated at 5 °C or 27 °C. The fatty acid composition of the lipids was estimated using gas chromatography. (3) Results: The BR-biosynthesis and BR-signaling mutants of barley were characterized by a temperature-dependent altered molar percentage of fatty acids (from 14:0 to 20:1) in their galactolipid and phospholipid fractions in comparison to wild-type (WT). For example, the mutants had a lower molar percentage of 18:3 in the phospholipid (PL) fraction. The same regularity was observed at 5 °C. It resulted in altered physicochemical parameters of the membranes (Alim, πcoll, Cs-1). (4) Conclusions: BR may be involved in regulating fatty acid biosynthesis or their transport/incorporation into the cell membranes. Mutants had altered physicochemical parameters of their membranes, compared to the WT, which suggests that BR may have a multidirectional impact on the membrane-dependent physiological processes.


Assuntos
Aclimatação/fisiologia , Membrana Celular/metabolismo , Genes de Plantas , Hordeum/genética , Hordeum/fisiologia , Mutação/genética , Temperatura , Fenômenos Químicos , Ácidos Graxos/metabolismo , Hordeum/crescimento & desenvolvimento , Lipídeos de Membrana/metabolismo
5.
Int J Mol Sci ; 21(5)2020 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-32164259

RESUMO

In temperature stress, the main role of heat-shock proteins (HSP) is to act as molecular chaperones for other cellular proteins. However, knowledge about the hormonal regulation of the production of the HSP is quite limited. Specifically, little is known about the role of the plant steroid hormones-brassinosteroids (BR)-in regulating the HSP expression. The aim of our study was to answer the question of how a BR deficit or disturbances in its signaling affect the accumulation of the HSP90, HSP70, HSP18, and HSP17 transcripts and protein in barley growing at 20 °C (control) and during the acclimation of plants at 5 °C and 27 °C. In barley, the temperature of plant growth modified the expression of HSPs. Furthermore, the BR-deficient mutants (mutations in the HvDWARF or HvCPD genes) and BR-signaling mutants (mutation in the HvBRI1 gene) were characterized by altered levels of the transcripts and proteins of the HSP group compared to the wild type. The BR-signaling mutant was characterized by a decreased level of the HSP transcripts and heat-shock proteins. In the BR-deficient mutants, there were temperature-dependent cases when the decreased accumulation of the HSP70 and HSP90 transcripts was connected to an increased accumulation of these HSP. The significance of changes in the accumulation of HSPs during acclimation at 27 °C and 5 °C is discussed in the context of the altered tolerance to more extreme temperatures of the studied mutants (i.e., heat stress and frost, respectively).


Assuntos
Brassinosteroides/biossíntese , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Hordeum/crescimento & desenvolvimento , Aclimatação , Vias Biossintéticas , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Hordeum/genética , Hordeum/metabolismo , Mutação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Temperatura
6.
Int J Mol Sci ; 21(1)2020 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-31948086

RESUMO

Brassinosteroids (BRs) are a class of steroidal phytohormones which are key regulators of diverse processes during whole life cycle of plants. Studies conducted in the dicot model species Arabidopsis thaliana have allowed identification and characterization of various components of the BR signaling. It is currently known that the BR signaling is interconnected at various stages with other phytohormonal and stress signaling pathways. It enables a rapid and efficient adaptation of plant metabolism to constantly changing environmental conditions. However, our knowledge about mechanism of the BR signaling in the monocot species is rather limited. Thus, identification of new components of the BR signaling in monocots, including cereals, is an ongoing process and has already led to identification of some monocot-specific components of the BR signaling. It is of great importance as disturbances in the BR signaling influence architecture of mutant plants, and as a consequence, the reaction to environmental conditions. Currently, the modulation of the BR signaling is considered as a target to enhance yield and stress tolerance in cereals, which is of particular importance in the face of global climate change.


Assuntos
Arabidopsis/metabolismo , Brassinosteroides/metabolismo , Produtos Agrícolas/metabolismo , Regulação da Expressão Gênica de Plantas/genética , Reguladores de Crescimento de Plantas/metabolismo , Transdução de Sinais/genética , Aclimatação , Adaptação Fisiológica , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Produtos Agrícolas/genética , Melhoramento Vegetal , Reguladores de Crescimento de Plantas/fisiologia , /metabolismo
7.
Biomolecules ; 11(1)2020 12 29.
Artigo em Inglês | MEDLINE | ID: mdl-33383794

RESUMO

Plants have developed various acclimation strategies in order to counteract the negative effects of abiotic stresses (including temperature stress), and biological membranes are important elements in these strategies. Brassinosteroids (BR) are plant steroid hormones that regulate plant growth and development and modulate their reaction against many environmental stresses including temperature stress, but their role in modifying the properties of the biological membrane is poorly known. In this paper, we characterise the molecular dynamics of chloroplast membranes that had been isolated from wild-type and a BR-deficient barley mutant that had been acclimated to low and high temperatures in order to enrich the knowledge about the role of BR as regulators of the dynamics of the photosynthetic membranes. The molecular dynamics of the membranes was investigated using electron paramagnetic resonance (EPR) spectroscopy in both a hydrophilic and hydrophobic area of the membranes. The content of BR was determined, and other important membrane components that affect their molecular dynamics such as chlorophylls, carotenoids and fatty acids in these membranes were also determined. The chloroplast membranes of the BR-mutant had a higher degree of rigidification than the membranes of the wild type. In the hydrophilic area, the most visible differences were observed in plants that had been grown at 20 °C, whereas in the hydrophobic core, they were visible at both 20 and 5 °C. There were no differences in the molecular dynamics of the studied membranes in the chloroplast membranes that had been isolated from plants that had been grown at 27 °C. The role of BR in regulating the molecular dynamics of the photosynthetic membranes will be discussed against the background of an analysis of the photosynthetic pigments and fatty acid composition in the chloroplasts.


Assuntos
Brassinosteroides/metabolismo , Cloroplastos/metabolismo , Hordeum/fisiologia , Aclimatação , Cloroplastos/genética , Resposta ao Choque Frio , Resposta ao Choque Térmico , Hordeum/genética , Simulação de Dinâmica Molecular , Mutação , Fotossíntese
8.
Open Life Sci ; 15: 217-228, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33987478

RESUMO

Plant hormones play important roles in controlling how plants grow and develop. While metabolism provides the energy needed for plant survival, hormones regulate the pace of plant growth. Strigolactones (SLs) were recently defined as new phytohormones that regulate plant metabolism and, in turn, plant growth and development. This group of phytohormones is derived from carotenoids and has been implicated in a wide range of physiological functions including regulation of plant architecture (inhibition of bud outgrowth and shoot branching), photomorphogenesis, seed germination, nodulation, and physiological reactions to abiotic factors. SLs also induce hyphal branching in germinating spores of arbuscular mycorrhizal fungi (AMF), a process that is important for initiating the connection between host plant roots and AMF. This review outlines the physiological roles of SLs and discusses the significance of interactions between SLs and other phytohormones to plant metabolic responses.

9.
J Plant Physiol ; 244: 153090, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31841952

RESUMO

The integral parts of the cell membranes are the functional proteins, which are crucial for cell life. Among them, proton-pumping ATPase and aquaporins appear to be of particular importance. There is some knowledge about the effect of the temperature during plant growth, including stress-inducing temperatures, on the accumulation of the membrane proteins: plasma membrane H+-ATPase and aquaporins, but not much is known about the effect of the phytohormones (i.e. brassinosteroids (BR)) on control of accumulation of these proteins. The aim of our study was to answer the question of how a BR deficit and disturbances in the BR perception/signalling affect the accumulation of plasma membrane H+-ATPase (PM H+-ATPase), the aquaporin HvPIP1 transcript and protein in barley growing at 20 °C and during its acclimation at 5 °C and 27 °C. For the studies, the BR-deficient mutant 522DK (derived from the wild-type Delisa), the BR-deficient mutant BW084 and the BR-signalling mutant BW312 and their wild-type Bowman were used. Generally, temperature of growth was significant factor influencing on the level of the accumulation of the H+-ATPase and HvPIP1 transcript and the PM H+-ATPase and HvPIP1 protein in barley leaves. The level of the accumulation of the HvPIP1 transcript decreased at 5 °C (compared to 20 °C), but was higher at 27 °C than at 20 °C in the analyzed cultivars. In both cultivars the protein HvPIP1 was accumulated in the highest amounts at 27 °C. On the other hand, the barley mutants with a BR deficiency or with BR signalling disturbances were characterised by an altered accumulation level of PM H+-ATPase, the aquaporin HvPIP1 transcript and protein (compared to the wild types), which may suggest the involvement of brassinosteroids in regulating PM H+-ATPase and aquaporin HvPIP1 at the transcriptional and translational levels.


Assuntos
Adenosina Trifosfatases/genética , Aquaporinas/genética , Hordeum/fisiologia , Proteínas de Plantas/genética , Aclimatação , Adenosina Trifosfatases/metabolismo , Aquaporinas/metabolismo , Brassinosteroides/metabolismo , Temperatura Baixa , Hordeum/genética , Temperatura Alta , Mutação , Proteínas de Plantas/metabolismo
10.
Front Plant Sci ; 10: 761, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31244877

RESUMO

Meiosis is a process of essential importance for sexual reproduction, as it leads to production of gametes. The recombination event (crossing-over) generates genetic variation by introducing new combination of alleles. The first step of crossing-over is introduction of a targeted double-strand break (DSB) in DNA. DMC1 (Disrupted Meiotic cDNA1) is a recombinase that is specific only for cells undergoing meiosis and takes part in repair of such DSBs by searching and invading homologous sequences that are subsequently used as a template for the repair process. Although role of the DMC1 gene has been validated in Arabidopsis thaliana, a functional analysis of its homolog in barley, a crop species of significant importance in agriculture, has never been performed. Here, we describe the identification of barley mutants carrying substitutions in the HvDMC1 gene. We performed mutational screening using TILLING (Targeting Induced Local Lesions IN Genomes) strategy and the barley TILLING population, HorTILLUS, developed after double-treatment of spring barley cultivar 'Sebastian' with sodium azide and N-methyl-N-nitrosourea. One of the identified alleles, dmc1.c, was found independently in two different M2 plants. The G2571A mutation identified in this allele leads to a substitution of the highly conserved amino acid (arginine-183 to lysine) in the DMC1 protein sequence. Two mutant lines carrying the same dmc1.c allele show similar disturbances during meiosis. The chromosomal aberrations included anaphase bridges and chromosome fragments in anaphase/telophase I and anaphase/telophase II, as well as micronuclei in tetrads. Moreover, atypical tetrads containing three or five cells were observed. A highly increased frequency of all chromosome aberrations during meiosis have been observed in the dmc1.c mutants compared to parental variety. The results indicated that DMC1 is required for the DSB repair, crossing-over and proper chromosome disjunction during meiosis in barley.

11.
Int J Mol Sci ; 19(9)2018 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-30205610

RESUMO

Brassinosteroids (BRs) are a class of phytohormones, which regulate various processes during plant life cycle. Intensive studies conducted with genetic, physiological and molecular approaches allowed identification of various components participating in the BR signaling-from the ligand perception, through cytoplasmic signal transduction, up to the BR-dependent gene expression, which is regulated by transcription factors and chromatin modifying enzymes. The identification of new components of the BR signaling is an ongoing process, however an emerging view of the BR signalosome indicates that this process is interconnected at various stages with other metabolic pathways. The signaling crosstalk is mediated by the BR signaling proteins, which function as components of the transmembrane BR receptor, by a cytoplasmic kinase playing a role of the major negative regulator of the BR signaling, and by the transcription factors, which regulate the BR-dependent gene expression and form a complicated regulatory system. This molecular network of interdependencies allows a balance in homeostasis of various phytohormones to be maintained. Moreover, the components of the BR signalosome interact with factors regulating plant reactions to environmental cues and stress conditions. This intricate network of interactions enables a rapid adaptation of plant metabolism to constantly changing environmental conditions.


Assuntos
Adaptação Fisiológica , Brassinosteroides/metabolismo , Desenvolvimento Vegetal , Reguladores de Crescimento de Plantas/metabolismo , Transdução de Sinais , Estresse Fisiológico , Meio Ambiente , Regulação da Expressão Gênica de Plantas , Homeostase , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Fatores de Transcrição/metabolismo
12.
Front Plant Sci ; 9: 216, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29515615

RESUMO

TILLING (Targeting Induced Local Lesions IN Genomes) is a strategy used for functional analysis of genes that combines the classical mutagenesis and a rapid, high-throughput identification of mutations within a gene of interest. TILLING has been initially developed as a discovery platform for functional genomics, but soon it has become a valuable tool in development of desired alleles for crop breeding, alternative to transgenic approach. Here we present the HorTILLUS ( Hordeum-TILLING-University of Silesia) population created for spring barley cultivar "Sebastian" after double-treatment of seeds with two chemical mutagens: sodium azide (NaN3) and N-methyl-N-nitrosourea (MNU). The population comprises more than 9,600 M2 plants from which DNA was isolated, seeds harvested, vacuum-packed, and deposited in seed bank. M3 progeny of 3,481 M2 individuals was grown in the field and phenotyped. The screening for mutations was performed for 32 genes related to different aspects of plant growth and development. For each gene fragment, 3,072-6,912 M2 plants were used for mutation identification using LI-COR sequencer. In total, 382 mutations were found in 182.2 Mb screened. The average mutation density in the HorTILLUS, estimated as 1 mutation per 477 kb, is among the highest mutation densities reported for barley. The majority of mutations were G/C to A/T transitions, however about 8% transversions were also detected. Sixty-one percent of mutations found in coding regions were missense, 37.5% silent and 1.1% nonsense. In each gene, the missense mutations with a potential effect on protein function were identified. The HorTILLUS platform is the largest of the TILLING populations reported for barley and best characterized. The population proved to be a useful tool, both in functional genomic studies and in forward selection of barley mutants with required phenotypic changes. We are constantly renewing the HorTILLUS population, which makes it a permanent source of new mutations. We offer the usage of this valuable resource to the interested barley researchers on cooperative basis.

13.
Physiol Plant ; 163(2): 155-169, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29215730

RESUMO

Drought is one of the most adverse stresses that affect plant growth and yield. Disturbances in metabolic activity resulting from drought cause overproduction of reactive oxygen species. It is postulated that brassinosteroids (BRs) regulate plant tolerance to the stress conditions, but the underlying mechanisms remain largely unknown. An involvement of endogenous BRs in regulation of the antioxidant homeostasis is not fully clarified either. Therefore, the aim of this study was to elucidate the role of endogenous BRs in regulation of non-enzymatic antioxidants in barley (Hordeum vulgare) under control and drought conditions. The plant material included the 'Bowman' cultivar and a group of semi-dwarf near-isogenic lines (NILs), representing mutants deficient in BR biosynthesis or signaling. In general, accumulations of 11 compounds representing various types of non-enzymatic antioxidants were analyzed under both conditions. The analyses of accumulations of reduced and oxidized forms of ascorbate indicated that the BR mutants contain significantly higher contents of dehydroascorbic acid under drought conditions when compared with the 'Bowman' cultivar. The analysis of glutathione accumulation indicated that under the control conditions the BR-insensitive NILs contained significantly lower concentrations of this antioxidant when compared with the rest of genotypes. Therefore, we postulate that BR sensitivity is required for normal accumulation of glutathione. A complete accumulation profile of various tocopherols indicated that functional BR biosynthesis and signaling are required for their normal accumulation under both conditions. Results of this study provided an insight into the role of endogenous BRs in regulation of the non-enzymatic antioxidant homeostasis.


Assuntos
Antioxidantes/metabolismo , Brassinosteroides/metabolismo , Hordeum/fisiologia , Transdução de Sinais , Secas , Genótipo , Homeostase , Hordeum/genética , Mutação , Estresse Fisiológico
14.
Front Plant Sci ; 7: 1824, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27994612

RESUMO

Brassinosteroids (BRs) are a class of steroid phytohormones, which regulate various processes of morphogenesis and physiology-from seed development to regulation of flowering and senescence. An accumulating body of evidence indicates that BRs take part in regulation of physiological reactions to various stress conditions, including drought. Many of the physiological functions of BRs are regulated by a complicated, and not fully elucidated network of interactions with metabolic pathways of other phytohormones. Therefore, the aim of this study was to characterize phytohormonal homeostasis in barley (Hordeum vulgare) in reaction to drought and validate role of BRs in regulation of this process. Material of this study included the barley cultivar "Bowman" and five Near-Isogenic Lines (NILs) representing characterized semi-dwarf mutants of several genes encoding enzymes participating in BR biosynthesis and signaling. Analysis of endogenous BRs concentrations in these NILs confirmed that their phenotypes result from abnormalities in BR metabolism. In general, concentrations of 18 compounds, representing various classes of phytohormones, including brassinosteroids, auxins, cytokinins, gibberellins, abscisic acid, salicylic acid and jasmonic acid were analyzed under control and drought conditions in the "Bowman" cultivar and the BR-deficient NILs. Drought induced a significant increase in accumulation of the biologically active form of BRs-castasterone in all analyzed genotypes. Another biologically active form of BRs-24-epi-brassinolide-was identified in one, BR-insensitive NIL under normal condition, but its accumulation was drought-induced in all analyzed genotypes. Analysis of concentration profiles of several compounds representing gibberellins allowed an insight into the BR-dependent regulation of gibberellin biosynthesis. The concentration of the gibberellic acid GA7 was significantly lower in all NILs when compared with the "Bowman" cultivar, indicating that GA7 biosynthesis represents an enzymatic step at which the stimulating effect of BRs on gibberellin biosynthesis occurs. Moreover, the accumulation of GA7 is significantly induced by drought in all the genotypes. Biosynthesis of jasmonic acid is also a BR-dependent process, as all the NILs accumulated much lower concentrations of this hormone when compared with the "Bowman" cultivar under normal condition, however the accumulation of jasmonic acid, abscisic acid and salicylic acid were significantly stimulated by drought.

15.
Int J Mol Sci ; 17(4)2016 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-27110778

RESUMO

Brassinosteroids (BRs) are plant steroid hormones, regulating a broad range of physiological processes. The largest amount of data related with BR biosynthesis has been gathered in Arabidopsis thaliana, however understanding of this process is far less elucidated in monocot crops. Up to now, only four barley genes implicated in BR biosynthesis have been identified. Two of them, HvDWARF and HvBRD, encode BR-6-oxidases catalyzing biosynthesis of castasterone, but their relation is not yet understood. In the present study, the identification of the HvDWARF genomic sequence, its mutational and functional analysis and characterization of new mutants are reported. Various types of mutations located in different positions within functional domains were identified and characterized. Analysis of their impact on phenotype of the mutants was performed. The identified homozygous mutants show reduced height of various degree and disrupted skotomorphogenesis. Mutational analysis of the HvDWARF gene with the "reverse genetics" approach allowed for its detailed functional analysis at the level of protein functional domains. The HvDWARF gene function and mutants' phenotypes were also validated by measurement of endogenous BR concentration. These results allowed a new insight into the BR biosynthesis in barley.


Assuntos
Brassinosteroides/biossíntese , Hordeum/genética , Reguladores de Crescimento de Plantas/biossíntese , Alelos , Sequência de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Colestanóis/metabolismo , Éxons , Homozigoto , Hordeum/fisiologia , Íntrons , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Fenótipo , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de Sequência
16.
Physiol Plant ; 158(3): 341-355, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27121184

RESUMO

In this study, the barley HvD14 gene encoding α/ß hydrolase, which is involved in strigolactone (SL) signaling, was identified. Bioinformatics analysis revealed that the identified gene is an orthologue of the D14, AtD14 and PhDAD2 genes that have been described in rice, Arabidopsis thaliana and petunia, respectively. Using TILLING strategy, an hvd14.d mutant that carried the G725A transition, located in the second exon, was identified. This mutation led to the substitution of a highly conserved glycine-193 to glutamic acid in the conserved fragment of the α/ß hydrolase domain of the HvD14 protein. The plants that carry the hvd14.d allele were semi-dwarf and produced a higher number of tillers in comparison to the wild-type (WT) parent cultivar. Additionally, the root architecture of mutant plants was affected: the total length of the seminal roots was significantly reduced, and the density of the lateral roots was higher than in the WT. Plants with the hvd14.d allele were insensitive to treatment with GR24, which is the synthetic analogue of SL. Analysis of the indole-3-acetic acid (IAA) concentration in the lateral buds showed no differences between the WT and mutant plants. By contrast, the WT seedlings treated with GR24 developed a lower number of tillers, longer primary roots with a reduced number of lateral roots and had an increased concentration of IAA in lateral buds. This paper describes the first barley SL mutant and shows the potential functions of SLs in barley growth and development.


Assuntos
Genes de Plantas/fisiologia , Hordeum/fisiologia , Lactonas/metabolismo , Reguladores de Crescimento de Plantas/fisiologia , Proteínas de Plantas/fisiologia , Regulação da Expressão Gênica de Plantas/fisiologia , Genes de Plantas/genética , Hordeum/crescimento & desenvolvimento , Hordeum/metabolismo , Ácidos Indolacéticos/metabolismo , Lactonas/farmacologia , Mutação , Proteínas de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/fisiologia , Estrutura Terciária de Proteína , Análise de Sequência de Proteína , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
17.
Plant Physiol Biochem ; 99: 126-41, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26752435

RESUMO

Brassinosteroids (BR) are plant steroid hormones that were discovered more than thirty years ago, but their physiological function has yet to be fully explained. The aim of the study was to answer the question of whether/how disturbances in the production of BR in barley affects the plant's metabolism and development under conditions of optimal watering and drought. Mutants with an impaired production of BR are one of the best tools in research aimed at understanding the mechanisms of action of these hormones. The study used barley cultivars with a normal BR synthesis (wild type) and semi-dwarf allelic mutants with an impaired activity of C6-oxidase (mutation in HvDWARF), which resulted in a decreased BR synthesis. Half of the plants were subjected to drought stress in the seedling stage and the other half were watered optimally. Plants with impaired BR production were characterised by a lower height and developmental retardation. Under both optimal watering and drought, BR synthesis disorders caused the reduced production of ABA and cytokinins, but not auxins. The BR mutants also produced less osmoprotectant (proline). The optimally watered and drought-stressed mutants accumulated less sucrose, which was accompanied by changes in the production of other soluble sugars. The increased content of fructooligosaccharide (kestose) in optimally watered mutants would suggest that BR is a negative regulator of kestose production. The decreased level of nystose in the drought-stressed mutants also suggests BR involvement in the regulation of the production of this fructooligosaccharide. The accumulation of the transcripts of genes associated with stress response (hsp90) was lower in the watered and drought-stressed BR-deficient mutants. In turn, the lower efficiency of photosystem II and the net photosynthetic rate in mutants was revealed only under drought conditions. The presented research allows for the physiological and biochemical traits of two BR-barley mutants to be characterised, which helps BR function to be understood. The knowledge can also be a good starting point for some breeding companies that are interested in introducing new semi-dwarf barley cultivars.


Assuntos
Brassinosteroides/biossíntese , Secas , Hordeum/fisiologia , Água/metabolismo , Citocininas/metabolismo , Hordeum/enzimologia , Hordeum/genética , Fotossíntese
18.
Front Plant Sci ; 6: 885, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26557130

RESUMO

The genomic integrity of every organism is constantly challenged by endogenous and exogenous DNA-damaging factors. Mutagenic agents cause reduced stability of plant genome and have a deleterious effect on development, and in the case of crop species lead to yield reduction. It is crucial for all organisms, including plants, to develop efficient mechanisms for maintenance of the genome integrity. DNA repair processes have been characterized in bacterial, fungal, and mammalian model systems. The description of these processes in plants, in contrast, was initiated relatively recently and has been focused largely on the model plant Arabidopsis thaliana. Consequently, our knowledge about DNA repair in plant genomes - particularly in the genomes of crop plants - is by far more limited. However, the relatively small size of the Arabidopsis genome, its rapid life cycle and availability of various transformation methods make this species an attractive model for the study of eukaryotic DNA repair mechanisms and mutagenesis. Moreover, abnormalities in DNA repair which proved to be lethal for animal models are tolerated in plant genomes, although sensitivity to DNA damaging agents is retained. Due to the high conservation of DNA repair processes and factors mediating them among eukaryotes, genes and proteins that have been identified in model species may serve to identify homologous sequences in other species, including crop plants, in which these mechanisms are poorly understood. Crop breeding programs have provided remarkable advances in food quality and yield over the last century. Although the human population is predicted to "peak" by 2050, further advances in yield will be required to feed this population. Breeding requires genetic diversity. The biological impact of any mutagenic agent used for the creation of genetic diversity depends on the chemical nature of the induced lesions and on the efficiency and accuracy of their repair. More recent targeted mutagenesis procedures also depend on host repair processes, with different pathways yielding different products. Enhanced understanding of DNA repair processes in plants will inform and accelerate the engineering of crop genomes via both traditional and targeted approaches.

19.
Mutagenesis ; 30(6): 785-97, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25958390

RESUMO

Genotoxic stress causes a reduced stability of the plant genome and has a detrimental effect on plant growth and productivity. Double-strand breaks (DSBs) are the most harmful of all DNA lesions because they cause the loss of genetic information on both strands of the DNA helix. In the presented study the coding and genomic sequences of the HvKu80 gene were determined. A mutational analysis of two fragments of HvKu80 using TILLING (Targeting Induced Local Lesions IN Genomes) allowed 12 mutations to be detected, which resulted in identification of 11 alleles. Multidirectional analyses demonstrated that the HvKu80 gene is involved in the elimination of DSBs in Hordeum vulgare. The barley mutants carrying the identified ku80.c and ku80.j alleles accumulated bleomycin-induced DSBs to a much greater extent than the parent cultivar 'Sebastian'. The altered reaction of the mutants to DSB-inducing agent and the kinetics of DNA repair in these genotypes are associated with a lower expression level of the mutated gene. The study also demonstrated the significant role of the HvKu80 gene in the regulation of telomere length in barley.


Assuntos
Quebras de DNA de Cadeia Dupla , DNA Helicases/genética , Reparo do DNA , Genes de Plantas , Hordeum/genética , Homeostase do Telômero/genética , Alelos , Sequência de Aminoácidos , DNA Helicases/química , DNA Helicases/metabolismo , Análise Mutacional de DNA , Regulação da Expressão Gênica de Plantas , Ordem dos Genes , Aptidão Genética , Homozigoto , Hordeum/crescimento & desenvolvimento , Hordeum/metabolismo , Dados de Sequência Molecular , Mutação , Fases de Leitura Aberta , Alinhamento de Sequência
20.
DNA Repair (Amst) ; 28: 116-30, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25753810

RESUMO

Genome integrity is constantly challenged by endo- and exogenous DNA-damaging factors. The influence of genotoxic agents causes an accumulation of DNA lesions, which if not repaired, become mutations that can cause various abnormalities in a cell metabolism. The main pathway of DSB repair, which is based on non-homologous recombination, is canonical non-homologous end joining (C-NHEJ). It has been shown that this mechanism is highly conserved in both Pro- and Eukaryotes. The mechanisms that underlie DSB repair through C-NHEJ have mainly been investigated in mammalian systems, and therefore our knowledge about this process is much more limited as far as plants, and crop plants in particular, are concerned. Recent studies have demonstrated that PARP3 is an important response factor to the presence of DSB in a genome. The aims of this study were to identify the sequence of the barley PARP3 gene, to perform a mutational analysis of the sequence that was identified using the TILLING (Targeting Induced Local Lesions IN Genomes) method and to phenotype the mutants that were identified through their exposure to mutagenic treatment with the DSB-inducing chemical--bleomycin. A functional analysis led to the identification of a series of parp3 alleles. The mutants were characterized using several different approaches, including quantifying the DSB and γH2AX foci, which validated the function of the HvPARP3 gene in DSB repair in barley. The potential involvement of the HvPARP3 gene in the regulation of telomere length in barley was also analyzed.


Assuntos
Alelos , Reparo do DNA , Genes de Plantas , Hordeum/genética , Poli(ADP-Ribose) Polimerases/genética , Sequência de Aminoácidos , DNA/metabolismo , Quebras de DNA de Cadeia Dupla , Hordeum/metabolismo , Dados de Sequência Molecular
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