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Michela Landoni,
Francesco Cerino Badone,
Nabil Haman,
Alberto Schiraldi,
Dimitrios Fessas,
Valentina Cesari,
Ivan Toschi,
Roberta Cremona,
Chiara Delogu,
Daniela Villa,
Elena Cassani, Roberto Pilu
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ABSTRACT: Monogastric animals are unable to digest phytic acid, so it represents an antinutritional factor and also an environmental problem. One strategy to solve this problem is the utilization of low phytic acid (lpa) mutants that accumulate low levels of phytic P and high levels of free phosphate in the seeds; among the lpa maize mutants lpa1 exhibited the highest reduction of phytic acid in the seed. This study indicated that the low phytic acid mutations exerted pleiotropic effects not directly connected to the phytic acid pathway, such as on seed density, content of ions, and the antioxidant compounds present in the kernels. Furthermore some nutritional properties of the flour were altered by the lpa1 mutations, in particular lignin and protein content, while the starch does not seem to be modified as to the total amount and in the amylose/amylopectin ratio, but alterations were noticed in the structure and size of granules.
Journal of Agricultural and Food Chemistry 05/2013; · 2.82 Impact Factor
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ABSTRACT: Phytic acid (PA), myo-inositol 1,2,3,4,5,6-hexakisphosphate, is the main storage form of phosphorus in plants. It is localized in seeds, deposited as mixed salts of mineral cations in protein storage vacuoles; during germination, it is hydrolyzed by phytases. When seeds are used as food/feed, PA and the bound cations are poorly bioavailable for human and monogastric livestock due to their lack of phytase activity. Reducing the amount of PA is one strategy to solve these problems and is an objective of genetic improvement for improving the nutritional properties of major crops. In this work, we present data on the isolation of a new maize (Zea mays L.) low phytic acid 1 (lpa1) mutant allele obtained by chemical mutagenesis. This mutant, named lpa1-7, is able to accumulate less phytic phosphorus and a higher level of free inorganic phosphate in the seeds compared with wild type. It exhibits a monogenic recessive inheritance and lethality as homozygous. We demonstrate that in vitro cultivation can overcome lethality allowing the growth of adult plants, and we report data regarding embryo and leaf abnormalities and other defects caused by negative pleiotropic effects of this mutation.
The Journal of heredity 05/2012; 103(4):598-605. · 2.05 Impact Factor
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ABSTRACT: In maize there are three types of brachytic mutants (br1, br2 and br3) showing short stature and a gibberellins-insensitive phenotype. So far only the brachytic 2 gene has been cloned and it encodes for a putative protein of the Multidrug Resistant (MDR) class of P-glycoproteins (PGPs)
that could be involved in polar movement of auxins: in fact the br2 mutant is insensitive to treatment with auxins and gibberellins. We have isolated a new recessive mutation of br2 gene (named br2-23) and with the aim of study its interactions with the other brachytic mutations we produced a br2 br3 double mutant that showed an additive effect on the stature with respect to the single mutants br2 and br3 and abnormal growth. In the progeny of the selfed double mutant we observed various defective seedlings, mirroring an altered
embryo development and growth, which also suggested a role for the br3 gene in auxin transport. Expression analysis of the auxin efflux transporters codified by ZmPIN1 genes supports this finding, showing the up-regulation of the ZmPIN1a gene in the br3 mutant. To our knowledge this is the first report showing the involvement of Br2 and Br3 genes in embryo development. These single and double mutants appear to be useful tools to study the genetics of plant height
and to investigate auxin transport in plants.
KeywordsMaize–Embryo development–
Brachytic 2 gene–Polar auxin transport–
ZmPIN1 genes
Plant Growth Regulation 04/2012; 64(2):185-192. · 1.60 Impact Factor
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ABSTRACT: More than 40 monogenic dwarfing mutants have been described in maize; however, the majority of these lead to great reductions
in grain yield and, consequently, they have not been used to enhance crop yield in germplasm that is sensible to lodging.
An exception in terms of commercial value is the maize mutant brachytic 2 (br2). Br2 gene was cloned in 2003, and it encodes for a protein most probably involved in auxin polar transport. We have isolated a
new brachytic mutation that is allelic to the br2 locus and denoted this novel mutant as br2–23. Characterization of this mutant revealed that the br2 mutation modified not only the length of the internodes, as previously reported, but the structure of the leaves as well.
Br2–23/br2–23 heterozygotes have a useful intermediate phenotype in terms of plant height, ear height and leaf angle, suggesting a possible
utilization of this effect in developing new hybrids. This mutant also appears to be an useful tool by which to study the
switch points of the complex developmental program determining maize plant height and architecture.
Molecular Breeding 04/2012; 20(2):83-91. · 2.85 Impact Factor
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ABSTRACT: The “green revolution” involving mainly wheat and rice was based on the use by breeders of semidominant mutations involved
in the signal transduction pathway of Gibberellin (GA). In particular, mutations in the Reduced height (Rht) gene of wheat have been used to reduce plant height and consequently to avoid storm damage and lodging. These genes have
been cloned and they encode for DELLA proteins which contain an N-terminal DELLA and a VHYNP domain essential for GA-dependent
degradation of these proteins. In maize several mutations have been isolated which affect gibberellin biosynthesis and perception
and in particular, mutations in Dwarf8 (D8) gene cause a severe dwarfing phenotype. D8 gene has been identified as an orthologue of Rht (Reduced height), Slr1(Slender rice 1) and Gibberellic Acid Insensitive (GAI) genes, this latter is a negative regulator of GA response in Arabidopsis. In this work, for the first time, we isolated and characterized a single amino acid insertion in the VHYNP domain of D8 maize gene causing the appearance of a dominant dwarf mutation. This spontaneous mutation, named D8-1023, showed a phenotype which is less severe in comparison with the other D8 mutants previously isolated which have modifications
in the DELLA domain. This mutant appears to be an useful tool either to study the mechanism of GA-modulated growth in plants
or to lower the height of maize tropical germplasm for breeding purposes.
Molecular Breeding 04/2012; 24(4):375-385. · 2.85 Impact Factor
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Roberto Pilu
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ABSTRACT: Gene silencing is associated with heritable changes in gene expression which occur without changes in DNA sequence. In eukaryotes these phenomena are common and control important processes, such as development, imprinting, viral and transposon sequence silencing, as well as transgene silencing. Among the epigenetic events, paramutation occurs when a silenced allele (named paramutagenic) is able to silence another allele (paramutable) in trans and this change is heritable. The silenced paramutable allele acquires paramutagenic capacity in the next generations. In the 1950s, Alexander Brink described for the first time the phenomenon of paramutation, occurring in maize at the colored1 (r1) gene, a complex locus (encoding myc-homologous transcription factors) that regulates the anthocyanin biosynthetic pathway. Since then, paramutation and paramutation-like interactions have been discovered in other plants and animals, suggesting that they may underlie important mechanisms for gene expression. The molecular bases of these phenomena are unknown. However in some cases, the event of paramutation has been correlated with changes in DNA methylation, chromatin structure and recently several studies suggest that RNA could play a fundamental role. This last consideration is greatly supported by genetic screening for mutants inhibiting paramutation, which allowed the identification of genes involved in RNA-directed transcriptional silencing, although it is possible that proteins are also required for paramutation.The meaning of paramutation in the life cycle and in evolution remains to be determined even though we might conjecture that this phenomenon could be involved in a fast heritability of favourable epigenetic states across generations in a non-Mendelian way.
Current Genomics 06/2011; 12(4):298-306. · 2.41 Impact Factor
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Dario Panzeri,
Elena Cassani,
Enrico Doria,
Giovanni Tagliabue,
Luca Forti,
Bruno Campion,
Roberto Bollini,
Charles A Brearley, Roberto Pilu,
Erik Nielsen,
Francesca Sparvoli
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ABSTRACT: • We previously identified the lpa1 (low phytic acid) 280-10 line that carries a mutation conferring a 90% reduction in phytic acid (InsP(6) ) content. In contrast to other lpa mutants, lpa1(280-10) does not display negative pleiotropic effects. In the present paper, we have identified the mutated gene and analysed its impact on the phytic acid pathway. • Here, we mapped the lpa1(280-10) mutation by bulk analysis on a segregating F(2) population, an then, by comparison with the soybean genome, we identified and sequenced a candidate gene. The InsP(6) pathway was analysed by gene expression and quantification of metabolites. • The mutated Pvmrp1(280-10) cosegregates with the lpa1(280-10) mutation, and the expression level of several genes of the InsP(6) pathway are reduced in the lpa1(280-10) mutant as well as the inositol and raffinosaccharide content. PvMrp2, a very similar paralogue of PvMrp1 was also mapped and sequenced. • The lpa1 mutation in beans is likely the result of a defective Mrp1 gene (orthologous to the lpa genes AtMRP5 and ZmMRP4), while its Mrp2 paralog is not able to complement the mutant phenotype in the seed. This mutation appears to down-regulate the InsP(6) pathway at the transcriptional level, as well as altering inositol-related metabolism and affecting ABA sensitivity.
New Phytologist 03/2011; 191(1):70-83. · 6.64 Impact Factor
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ABSTRACT: Mycotoxins are secondary metabolites with potential dangers for animal and human health. In particular, maize (Zea mays L.) infection caused by Fusarium and the consequent fumonisin contamination is widespread in several countries such as Italy. We developed six maize populations differing in their constitution of regulatory genes able to accumulate respectively anthocyanins in the aleurone layer (r1 gene), pericarp (b1 and pl1 genes) and phlobaphene in the pericarp (p1 gene). These coloured populations, with the related control colourless populations were analysed for mycotoxin content in the kernels during three field seasons with the aim of understanding if there were any correlations with their ability to accumulate flavonoids in kernel tissues. Our results indicate that accumulation of flavonoid pigments in the seeds, in particular phlobaphenes, is able to reduce the level of fumonisin B1. This finding could be used to minimize kernel mycotoxin contamination in this crop, in particular, the development of sweet, pop and polenta coloured corn varieties will help the farmer to keep the level of fumonisin under the threshold of contamination established for human corn consumption.
Journal of applied genetics 11/2010; 52(2):145-52. · 1.66 Impact Factor
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ABSTRACT: The lpa1 mutations in maize are caused by lesions in the ZmMRP4 (multidrug resistance-associated proteins 4) gene. In previous studies (Raboy et al. in Plant Physiol 124:355-368, 2000; Pilu et al. in Theor Appl Genet 107:980-987, 2003a; Shi et al. Nat Biotechnol 25:930-937, 2007), several mutations have been isolated in this locus causing a reduction of phytic acid (myo-inositol-1,2,3,4,5,6-hexakisphosphate, or InsP(6)) content and an equivalent increasing of free phosphate. In particular, the lpa1-241 mutation causes a reduction of up to 90% of phytic acid, associated with strong pleiotropic effects on the whole plant. In this work, we show, for the first time to our knowledge, an interaction between the accumulation of anthocyanin pigments in the kernel and the lpa mutations. In fact the lpa1-241 mutant accumulates a higher level of anthocyanins as compared to wild type either in the embryo (about 3.8-fold) or in the aleurone layer (about 0.3-fold) in a genotype able to accumulate anthocyanin. Furthermore, we demonstrate that these pigments are mislocalised in the cytoplasm, conferring a blue pigmentation of the scutellum, because of the neutral/basic pH of this cellular compartment. As a matter of fact, the propionate treatment, causing a specific acidification of the cytoplasm, restored the red pigmentation of the scutellum in the mutant and expression analysis showed a reduction of ZmMRP3 anthocyanins' transporter gene expression. On the whole, these data strongly suggest a possible interaction between the lpa mutation and anthocyanin accumulation and compartmentalisation in the kernel.
Planta 02/2010; 231(5):1189-99. · 3.00 Impact Factor
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ABSTRACT: Previous studies suggested that micropore surface area (MSA) of alkali-soluble bio-macromolecules of aerial plant residues of maize constitutes an important factor that explains their humification in soil, that is, preservation against biological degradation. On the other hand, root plant residue contributes to the soil humus balance, as well. Following the experimental design used in a previous paper published in this journal, this study shows that the biochemical recalcitrance of the alkali-soluble acid-insoluble fraction of the root plant material, contributed to the root maize humification of both Wild-type maize plants and its corresponding mutant brown midrib (bm3), this latter characterized by reduced lignin content. Humic acids (HAs) existed in root (root-HAs) were less degraded in soil than corresponding HAs existed in shoot (shoot-HAs): shoot-HAs bm3 (48%)>shoot-HAs Wild-type (37%)>root-HAs Wild-type (33%)>root-HAs bm3 (22%) (degradability shown in parenthesis). These differences were related to the MSA of HAs, that is, root-HAs having a higher MSA than shoot-HAs: shoot-HAs bm3 (41.43+/-1.2m(2)g(-1))<shoot-HAs Wild-type (43.43+/-1.7m(2)g(-1))<root-HAs Wild-type (51.7+/-3.6m(2)g(-1))<root-HAs bm3 (54.08+/-3.9m(2)g(-1)). Taking into account both the previous data obtained for maize shoots and the results of this study, it was possible to find a very good correlation between degradability of HAs and HA-MSAs (r=-0.88, P<0.08, n=4), confirming that MSA was able to explain bio-macromolecules recalcitrance in soil.
Chemosphere 12/2009; 78(8):1036-41. · 3.21 Impact Factor
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ABSTRACT: The quantity and quality of plant litter in the soil play an important role in the soil organic matter balance. Besides other pedo-climatic aspects, the content of recalcitrant molecules of plant residues and their chemical composition play a major role in the preservation of plant residues. In this study, we report that intrinsically resistant alkali-soluble bio-macromolecules extracted from maize plant (plant-humic acid) (plant-HA) contribute directly to the soil organic matter (OM) by its addition and conservation in the soil. Furthermore, we also observed that a high syringyl/guaiacyl (S/G) ratio in the lignin residues comprising the plant tissue, which modifies the microscopic structure of the alkali-soluble plant biopolymers, enhances their recalcitrance because of lower accessibility of molecules to degrading enzymes. These results are in agreement with a recent study, which showed that the humic substance of soil consists of a mixture of identifiable biopolymers obtained directly from plant tissues that are added annually by maize plant residues.
Chemosphere 04/2009; 76(4):523-8. · 3.21 Impact Factor
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ABSTRACT: A maize mutant defective in the synthesis of phytic acid during seed maturation was used as a tool to study the consequences of the lack of this important reserve substance on seed survival. Data on germinability, free iron level, free radical relative abundance, protein carbonylation level, damage to DNA, degree of lipid peroxidation, alpha- and gamma-tocopherol amount and antioxidant capacity were recorded on seeds of maize B73 and of an isogenic low phytic acid mutant (lpa1-241), either unaged or incubated for 7 d in accelerated ageing conditions (46 degrees C and 100% relative humidity). The lpa1-241 mutant, compared to wild type (wt), showed a lower germination capacity, which decreased further after accelerated ageing. Whole lpa1-241 mutant kernels contained about 50% more free or weakly bound iron than wt ones and showed a higher content of free radicals, mainly concentrated in embryos; in addition, upon accelerated ageing, lpa1-241 seed proteins were more carbonylated and DNA was more damaged, whereas lipids did not appear to be more peroxidated, but the gamma-tocopherol content was decreased by about 50%. These findings can be interpreted in terms of previously reported but never proven antioxidant activity of phytic acid through iron complexation. Therefore, a novel role in plant seed physiology can be assigned to phytic acid, that is, protection against oxidative stress during the seed's life span. As in maize kernels the greater part of phytic acid (and thus of metal ions) is concentrated in the embryo, its antioxidant action may be of particular relevance in this crop.
Journal of Experimental Botany 03/2009; 60(3):967-78. · 5.36 Impact Factor
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Marie-Claire Toufektsian,
Michel de Lorgeril,
Norbert Nagy,
Patricia Salen,
Maria Benedetta Donati,
Lucia Giordano,
Hans-Peter Mock,
Silke Peterek,
Andrea Matros,
Katia Petroni, Roberto Pilu,
Domenico Rotilio,
Chiara Tonelli,
Joel de Leiris,
François Boucher,
Cathie Martin
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ABSTRACT: Consumption of flavonoid-rich foods and beverages is thought to reduce the risk of cardiovascular diseases. Whereas the biological activities of flavonoids have been characterized in vitro, there are no clear experimental data demonstrating that chronic dietary intake and intestinal absorption of flavonoids actually protects the heart against ischemia-reperfusion injury. We tested whether long-term consumption of specific flavonoids (anthocyanins) included in normal food could render the heart of rats more resistant to myocardial infarction. Maize kernels that differed specifically in their accumulation of anthocyanins were used to prepare rodent food in which anthocyanins were either present or absent. Male Wistar rats were fed the anthocyanin-rich (ACN-rich) or the anthocyanin-free (ACN-free) diet for a period of 8 wk. Anthocyanins were significantly absorbed and detected in the blood and urine of only rats fed the ACN-rich diet. In Langendorff preparations, the hearts of rats fed the ACN-rich diet were more resistant to regional ischemia and reperfusion insult. Moreover, on an in vivo model of coronary occlusion and reperfusion, infarct size was reduced in rats that ate the ACN-rich diet than in those that consumed the ACN-free diet (P < 0.01). Cardioprotection was associated with increased myocardial glutathione levels, suggesting that dietary anthocyanins might modulate cardiac antioxidant defenses. Our findings suggest important potential health benefits of foods rich in anthocyanins and emphasize the need to develop anthocyanin-rich functional foods with protective activities for promoting human health.
Journal of Nutrition 04/2008; 138(4):747-52. · 3.92 Impact Factor
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ABSTRACT: The structure of the plant inflorescence and flower is an important agronomic and ornamental trait studied for its potential
economic applications. In particular, the capacity to modify flower size has always been a breeder’s goal. Genetic and molecular
studies have shown that the Zea mays gene Ramosa1 (Ra1) is involved in inflorescence branching regulation. In fact the ra1 loss of function mutation causes extra branching of the inflorescence. In this work we suggest a possible utilization of
the Ramosa1 maize gene as a tool to modify inflorescence architecture and flower size in transgenic plants. In fact overexpression of
this gene in Arabidopsis plants promotes an increase in reproductive organ size. Pollen, seeds, cotyledons, leaves and roots are also larger than
those of the wild type. Analysis of organs from transformants showed that cell expansion was increased without apparently
affecting cell division. These results suggest that the RA1 protein is able to up-regulate cell expansion in all organs of
Arabidopsis plants.
Sexual Plant Reproduction 11/2007; 20(4):191-198. · 1.87 Impact Factor
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ABSTRACT: The maize seed comprises two major compartments, the embryo and the endosperm, both originating from the double fertilization event. The embryogenetic process allows the formation of a well-differentiated embryonic axis, surrounded by a single massive cotyledon, the scutellum. The mature endosperm constitutes the bulk of the seed and comprises specific regions containing reserve proteins, complex carbohydrates, and oils. To gain more insight into molecular events that underlie seed development, three monogenic mutants were characterized, referred to as emp (empty pericarp) on the basis of their extreme endosperm reduction, first recognizable at about 12 d after pollination. Their histological analysis reveals a partial development of the endosperm domains as well as loss of adhesion between pedicel tissues and the basal transfer layer. In the endosperm, programmed cell death (PCD) is delayed. The embryo appears retarded in its growth, but not impaired in its morphogenesis. The mutants can be rescued by culturing immature embryos, even though the seedlings appear retarded in their growth. The analysis of seeds with discordant embryo-endosperm phenotype (mutant embryo, normal endosperm and vice-versa), obtained using B-A translocations, suggests that emp expression in the embryo is necessary, but not sufficient, for proper seed development. In all three mutants the picture emerging is one of a general delay in processes related to growth, as a result of a mutation affecting endosperm development as a primary event.
Journal of Experimental Botany 02/2007; 58(5):1197-205. · 5.36 Impact Factor
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ABSTRACT: The genetic and molecular control of inflorescence and flower development has been extensively studied in model dicotyledonous plants such as Arabidopsis but even now little is known about monocotyledonous species. In maize several mutants have been isolated that perturbed normal inflorescence development. In particular, the Ramosa1 (Ra1) gene, coding for a zinc finger transcription factor, plays a role in inflorescence structure by determining the number of branches. Although the mechanism by which Ra1 acts is unclear, inflorescence meristems in these regions assume a branch meristem identity rather than becoming spikelet pairs. In this work we characterize a new mutation of Ra1 gene that originated spontaneously from a B73 inbred line. This loss-of-function mutation is caused by the deletion of the lysine residue at position 53 in the RA1 putative zinc-finger domain. This is the first evidence for a single amino acid deletion in the zinc finger domain that knocks out the function of the RA1 protein. This result strongly suggests that the RA1 protein functions by acting as a DNA-binding protein, probably involved in transcriptional regulation. Furthermore, Ra1 overexpression in the Arabidopsis ortholog superman (sup) mutant, whose flowers are characterized by the presence of additional stamens, was not able to restore the correct stamen number, indicating that SUP and Ra1 genes do not share an identical function.
Sexual Plant Reproduction 08/2006; 19(3):145-150. · 1.87 Impact Factor
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ABSTRACT: The pl1 gene encodes a MYB-related transcriptional activator committed to the regulation of anthocyanin biosynthesis in maize. Here, we report the genetic and molecular characterisation of pl-bol3, an Andean allele displaying features that make it different from all the known pl1 alleles. pl-bol3 has partial, light-independent expression, and it is active mainly in the juvenile phase of growth. It has a complex molecular structure, containing multiple pl1 gene copies, thus being the first complex locus discovered in the c1/pl1 family. Although the composite genes of the complex locus encode proteins identical to other functional PL1 proteins, the putative promoters of the pl-bol3 gene are different from the promoters of Pl-Rhoades (Pl-Rh) and pl1 sun-red alleles. The intensity and the tissue specificity of anthocyanin production directed by pl-bol3 differ significantly from that of Pl-Rh and the original pl-W22, and are specified by the interaction of pl-bol3 with the different r1/b1 gene family members and the competence of pl-bol3 to different pigment tissues. This allele represents a natural example of gene duplication and diversification of expression, giving rise to a significant change in phenotype and, in this way, is analogous to the complex r1 locus in maize. Analysis of the pl-bol3 allele contributes to understanding the generation of diversity associated with multiple-copy genes and the molecular basis of allele-specific gene expression.
The Plant Journal 12/2003; 36(4):510-21. · 6.16 Impact Factor
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ABSTRACT: The shoot apical meristem (SAM), initially formed during embryogenesis, gives rise to the aboveground portion of the maize (Zea mays) plant. The shootless phenotype (sml) described here is caused by disruption of SAM formation due to the synergistic interaction of mutations at two genetic loci. Seedlings must be homozygous for both sml (shootmeristemless), and the unlinked dgr (distorted growth) loci for a SAM-less phenotype to occur. Seedlings mutant only for sml are impaired in their morphogenesis to different extents, whereas the dgr mutation alone does not have a recognisable phenotype. Thus, dgr can be envisaged as being a dominant modifier of sml and the 12 (normal):3 (distorted growth):1 (shoot meristemless) segregation observed in the F(2) of the double heterozygote is the result of the interaction between the sml and dgr genes. Other segregation patterns were also observed in the F(2), suggesting instability of the dgr gene. Efforts to rescue mutant embryos by growth on media enriched with hormones have been unsuccessful so far. However, mutant roots grow normally on medium supplemented with kinetin at a concentration that suppresses wild-type root elongation, suggesting possible involvement of the mutant in the reception or transduction of the kinetin signal or transport of the hormone. The shootless mutant appears to be a valuable tool with which to investigate the organization of the shoot meristem in monocots as well as a means to assay the origins and relationships between organs such as the scutellum, the coleoptile, and leaves that are initiated during the embryogenic process.
Plant physiology 03/2002; 128(2):502-11. · 6.53 Impact Factor
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ABSTRACT: Sharpley et al., 1994). These lpa mutants produced seeds in which the chemistry of seed P, but not the total Phytic acid is a nearly ubiquitous component of plant seeds, supply-amount of P, was greatly altered (Raboy and Gerbasi, ing both phosphate (P) and cations during germination. However, during digestion, the phytic acid form of P is not bioavailable for 1996; Larson et al., 1998, 2000; Raboy et al., 2000; Sebas-monogastric animals. A possible solution to this problem is the isola-tian et al., 2000; Pilu et al., 2003). tion of cereal mutants accumulating less phytic P and more free P These mutants are quite interesting, as they may be and cations in the seed (low phytic acid, lpa). In a previously published used to obtain information on the role and mechanisms study, we described a single, recessive lpa mutation (named lpa241) of action of myo-inositol and its derivatives in both plant in maize (Zea mays L.) mapping on chromosome 1S in the same and seed physiology. For instance, it has been shown location of the lpa1-1 mutant, showing a decrease in the expression that the genetic lesion causing the LR33 soybean muta-of the first enzyme of phytic acid pathway, myo-inositol-3-phosphate tion is a single base change in the third base of the synthase (MIPS). The goal of this study was to gain further information codon for the amino acid residue 396, which decreases about the nature of lpa241 mutation as well as its effects on seed the specific activity of the seed-expressed myo-inositol physiology and plant growth. We present new results of genetic, molec-ular, and histological analyses demonstrating that lpa241 is indeed 1-phosphate synthase by about 90% (Hitz et al., 2002). allelic to lpa1-1, its MIPS1S coding region does not show a relevant The LR33 mutation also confers a seed phenotype char-molecular lesion, is accompanied by a strong reduction in globoid acterized by increased P and decreased phytic acid and number and size, and shows highly variable expression causing some induces a decrease in the level of raffinosaccharides, negative pleiotropic effects related to embryo development and size, which are synthesized from myo-inositol. In cereals, al-germination rate, seedling growth rate, and ear size. The origin and though the molecular lesions have not been discovered, the consequences of the lpa241 mutation led us to advance the hypoth-the lpa-2 mutations affect some of the six phosphoryla-esis of an epigenetic nature of the mutation and to emphasize the tion steps involved in the synthesis of phytic acid from role of myo-inositol and its derivatives during seed maturation, germi-myo-inositol. Hatzack et al. (2001) reported that D/L-nation, and seedling growth. Ins(1,3,4,5)P4 accumulates in lpa-2 barley mutant seeds without affecting their germination vigor. All lpa-1 mu-tations affect the first committed step in inositol biosyn-P hytic acid is the most abundant form of P in cereals thesis, i.e., the production of myo-inositol-3-phosphate (O'Dell et al., 1972; Raboy et al., 1990, 2001; Brinch-(Ins3P) from glucose-6-phosphate (G6P), catalyzed by Pedersen et al., 2002). Unfortunately, it is generally very the enzyme myo-inositol-3-phosphate synthase (Ins3P poorly available for monogastric animals because of synthase, MIPS) (Loewus et al., 1990; Yoshida et al., their lack of phytase activity. A possible solution to 1999; Loewus and Murthy 2000). The product of this this problem is to produce, with either transgenic or reaction, Ins3P, is then dephosphorylated by inositol mutation methods, cereals that accumulate less phytic monophosphatase to yield myo-inositol, which in addi-P and contain more free P in the seed. Several mutants tion to being phosphorylated to hexakis phosphate (phy-have been isolated in recent years in maize, barley tic acid) during seed maturation, plays a central role in (Hordeum vulgare L.), rice (Oryza sativa L.), and soy-several metabolic processes and in signal transduction bean [Glycine max (L.) Merr.] (Raboy and Gerbasi in the plant Ray-tian et al., 2000). The results of several experiments, in chaudhuri et al., 1997). which monogastric animals were fed with lpa mutant Thus, the free myo-inositol level may influence both flours, demonstrated an improvement in animal phos-plant growth and development and responses to vari-phate nutrition and a consequent decrease of the amount ations in environmental conditions in different ways of excreted inorganic P in the manure, resulting in an (Munnik et al., 1998; Stevenson et al., 2000). For in-alleviation of the associated environmental problems stance, perturbation in growth patterns may occur be-cause of alterations in cell wall extensibility caused by R. Pilu and E. Cassani, Dipartimento di Produzione Vegetale, Uni-variations in myo-inositol-derived wall components. Like- about by variations in the supply of myo-inositol-derived 2097 ACR were used as donors in initial crosses with W64A, W22, an increased salt tolerance in transgenic plants. How-B73, and K6 maize inbred lines. The BC 2 F 2 seeds were used ever, the higher amount of myo-inositol did not result for quantitative analysis of P fractions. The lpa1 mutant stock in salt tolerance or alteration of a number of other plant used for the allelism test was provided by Dr. Victor Raboy, characteristics linked with putative functions of myo-USDA ARS, Aberdeen, ID. inositol-derived metabolites. Nevertheless, other approaches based on the genera-Assay for High Phosphate Levels in Maize Kernels tion of transgenic plants, in which an antisense RNA strategy allows the suppression of Ins3P synthase activ-Seeds were individually ground in a mortar with a steel pestle. A total of 100 mg of the resulting flour was then ex-ity, or based on the study of MIPS defective mutants, tracted with 1 mL 0.4 M HCl at 4C overnight. Samples were might produce more information. These approaches might mixed briefly and 100 L were removed and supplemented disclose the consequences of myo-inositol deficiency on with 900 L Chen's reagent (12 M H 2 SO 4 : 2.5% ammonium both phytic acid and raffinosaccharide accumulation as molybdate: 10% ascorbic acid: H 2 O [1:1:1:2,v/v/v/v]) in micro-well as on any process influenced by inositol-derived titer plates (Chen et al., 1956). In the case of high phosphate compounds. content, a dark-blue colored phosphomolybdate complex formed
Crop Science 01/1996; 45(45):2096-2105. · 1.64 Impact Factor
