Are high-lysine cereal crops still a challenge?

The essential amino acids lysine and threonine are synthesized in higher plants via a pathway starting with aspartate that also leads to the formation of methionine and isoleucine. Lysine is one of most limiting amino acids in plants consumed by humans and livestock. Recent genetic, molecular, and biochemical evidence suggests that lysine synthesis and catabolism are regulated by complex mechanisms. Early kinetic studies utilizing mutants and transgenic plants that over-accumulate lysine have indicated that the major step for the regulation of lysine biosynthesis is at the enzyme dihydrodipicolinate synthase. Despite this tight regulation, recent strong evidence indicates that lysine catabolism is also subject to control, particularly in cereal seeds. The challenge of producing crops with a high-lysine concentration in the seeds appeared to be in sight a few years ago. However, apart from the quality protein maize lines currently commercially available, the release of high-lysine crops has not yet occurred. We are left with the question, is the production of high-lysine crops still a challenge?

Are high-lysine cereal crops still a challenge?

The essential amino acids lysine and threonine are synthesized in higher plants via a pathway starting with aspartate that also leads to the formation of methionine and isoleucine. Lysine is one of most limiting amino acids in plants consumed by humans and livestock. Recent genetic, molecular, and biochemical evidence suggests that lysine synthesis and catabolism are regulated by complex mechanisms. Early kinetic studies utilizing mutants and transgenic plants that over-accumulate lysine have indicated that the major step for the regulation of lysine biosynthesis is at the enzyme dihydrodipicolinate synthase. Despite this tight regulation, recent strong evidence indicates that lysine catabolism is also subject to control, particularly in cereal seeds. The challenge of producing crops with a high-lysine concentration in the seeds appeared to be in sight a few years ago. However, apart from the quality protein maize lines currently commercially available, the release of high-lysine crops has not yet occurred. We are left with the question, is the production of high-lysine crops still a challenge?.

Biochemical changes during the development of witches' broom: the most important disease of cocoa in Brazil caused by Crinipellis perniciosa.

Witches' broom disease (WBD) is caused by the hemibiotrophic basidiomycete fungus Crinipellis perniciosa, which is one of the most important diseases of cocoa in the western hemisphere. In this study, the contents of soluble sugars, amino acids, alkaloids, ethylene, phenolics, tannins, flavonoids, pigments, malondialdehyde (MDA), glycerol, and fatty acids were analysed in cocoa (Theobroma cacao) shoots during the infection and development of WBD. Alterations were observed in the content of soluble sugars (sucrose, glucose, and fructose), asparagine and alkaloids (caffeine and theobromine), ethylene, and tannins. Ethylene and tannins increased prior to symptom development and declined with the death of the infected tissues. Furthermore, MDA and glycerol concentrations were higher in infected tissue than in the controls, while fatty acid composition changed in the infected tissues. Chlorophylls a and b were lower throughout the development of the disease while carotenoids and xanthophylls dropped in the infected tissue by the time of symptom development. These results show co-ordinated biochemical alterations in the infected tissues, indicating major stress responses with the production of ethylene. Ethylene levels are hypothesized to play a key role in broom development. Some of the other biochemical alterations are directly associated with ethylene synthesis and may be important for the modification of its effect on the infected tissues.

Nrs1, a repetitive element linked to pisatin demethylase genes on a dispensable chromosome of Nectria haematococca.

We have identified a repetitive DNA element in Nectria haematococca mating population VI, isolate T-2. This repetitive sequence has been called Nrs1. DNA hybridization analysis indicates the sequence is found in several isolates of the fungus pathogenic to Pisum sativum. A 2,027-bp clone containing the Nrs1-2 allele contains a long polyA sequence, imperfect RNA polymerase III promoter sequences, multiple inverted repeats, and the potential for extensive secondary structure similar to known RNA polymerase III transcripts and related retroelements. Ten of the 11 HindIII restriction fragments from isolate T-2 DNA that hybridize to Nrs1-2 segregate in a manner consistent with a 1:1 ratio for random ascospore progeny. The 10 restriction fragment length polymorphism (RFLP) loci define three linkage groups and correspond to three chromosome-sized DNAs from T-2 separated by pulsed field gel electrophoresis. Three RFLP loci defined by hybridization to the gene for pisatin demethylase and localized on the 1.6 million base pair (Mb) chromosome were genetically linked to each other and to several Nrs1 loci. These sequences recombined despite the fact that no obvious homolog exists for the 1.6-Mb chromosome in one parent strain. Allelic RFLPs corresponding to the gene sequence of cutinase were unlinked to Nrs1 loci.

Molecular cloning and characterization of a sym plasmid locus that regulates cultivar-specific nodulation of soybean by Rhizobium fredii USDA257.

Rhizobium fredii strain USDA257 produces nitrogen-fixing nodules on primitive soybean cultivars such as Peking but fails to nodulate agronomically improved cultivars such as McCall. Transposon-mutant 257DH4 has two new phenotypes: it nodulates McCall, and its ability to do so is sensitive to the presence of parental strain USDA257, i.e. it is subject to competitive nodulation blocking. We have isolated a cosmid containing DNA that corresponds to the site of transposon insertion in 257DH4 and have localized Tn5 on an 8.0 kb EcoRI fragment. The 5596 bp DNA sequence that surrounds the insertion site contains seven open reading frames. Five of these, designated nolBTU, ORF4, and nolV, are closely spaced and of the same polarity. nolW and nolX are of the opposite polarity. The initiation codon for nolW lies 155 bp upstream from that of nolB, and its is separated from nolX by 281 bp. The predicted NolT and NolW proteins have putative membrane-spanning regions. The N-terminus of the hypothetical NolW protein also has limited homology to NodH of Rhizobium meliloti, but none of the deduced protein sequences has significant homology to known nodulation gene products. Site-directed mutagenesis with mudII1734 confirms that inactivation of nolB, nolT, nolU, nolV, nolW, or nolX extends host range for nodulation to McCall soybean. This phenotype could not be genetically dissected from sensitivity to competitive nodulation blocking. Expression of nolBTU and nolX is induced as much as 30-fold by flavonoid signal molecules, even though these genes lack nod-box promoters. Histochemical staining of McCall roots inoculated with nolB-, nolU-, or nolX-lacZ fusions verifies that these genes are expressed continuously from preinfection to the stage of the functional nodule. Although a nolU-ORF4-nolV clone hybridizes to a single 8.0 kb EcoRI fragment from 10 strains of R. fredii and broad-host-range Rhizobium sp. NGR234, hybridizing sequences are not detectable in other rhizobia.