F. Garcia-Olmedo

Synthesis and processing of thionin precursors in developing endosperm from barley (Hordeum vulgare L.).

Thionin is a lysine‐rich polypeptide (mol. wt. 5000) which is synthesized in developing barley endosperm from ˜8 days to ˜30 days after anthesis. Two thionin precursors (THP1 and THP2) have been identified using monospecific antibodies (A‐TH) prepared against the mature protein. THP1, which is the only polypeptide recognized in vitro by A‐TH, is encoded by a 7.5S mRNA obtained from membrane‐bound polysomes, and its alkylated derivative has an apparent mol. wt. of 17 800. THP2, which is selected together with mature thionin by A‐TH among labelled proteins in vivo, differs from THP1 in apparent mol. wt. (17 400 alkylated) and in electrophoretic mobility at pH 3.2. Both THP1 and THP2 are competed out of the antigen‐antibody complex by purified thionin. The conversion of THP2 into thionin, which has been demonstrated in a pulse‐chase experiment in vivo, is a post‐translational process. As it has not been possible to detect THP1 in vivo it is assumed that it is converted co‐translationally into THP2. Final deposition of thionin as an extrinsic membrane protein, possibly associated with the endoplasmic reticulum, has been tentatively established on the basis of subcellular fractionation experiments.

Genetics of CM-proteins (A-hordeins) in barley

SummaryThe CM-proteins, which are the main components of the A-hordeins, include four previously described proteins (CMa-1, CMb-1, CMc-1, CMd-1), plus a new one, CMe-1, which has been tentatively included in this group on the basis of its solubility properties and electrophoretic mobility. The variability of the five proteins has been investigated among 38 Hordeum vulgare cultivars and 17 H. spontaneum accessions. Proteins CMa-1, CMc-1 and CMd-1 were invariant within the cultivated species; CMd was also invariant in the wild one. The inheritance of variants CMb-1/CMb-2 and CMe-1/CMe-2,2′ was studied in a cross H. spontaneum x H. vulgare. The first two proteins were inherited as codominantly expressed allelic variations of a single mendelian gene. Components CMe-2,2′ were jointly inherited and codominantly expressed with respect to CMe-1. Gene CMb and gene(s) CMe were found to be unlinked. The chromosomal locations of genes encoding CM-proteins were investigated using wheat-barley addition lines. Genes CMa and CMc were associated with chromosome 1, and genes CMb and CMd with chromosome 4. These gene locations further support the proposed homoeology of chromosomes 1 and 4 of barley with chromosomes groups 7 and 4 of wheat, respectively. Gene(s) CMe has been assigned to chromosome 3 of barley. The accumulation of protein CMe-1 is totally blocked in the “high lysine” mutant Riso 1508 and partially so in the high lysine barley Hiproly.

Heterogeneity of wheat endosperm proteolipids (CM proteins)

Proteins extracted with CHCl3-MeOH from wheat endosperm have been fractionated by Sephadex G-100 and the 15 000–20 000 MW range fraction, designated CM protein, has been examined by combined electrofocusing (pH range 5–8) and electrophoresis (pH 3.2) and the heterogeneity of the electrophoretic components has been ascertained. It has been shown by joint mapping and by sequential extraction that CM proteins are extracted by 70% EtOH but not by H2O, although they can be made water-soluble after dialysis against an acid buffer, pH 3.2, 3 M urea, without losing their solubility in CHCl3-MeOH mixtures. It is concluded that CM proteins fit the definition of a Folch—Lees proteolipid. The Triticum aestivum (genomes ABD) map can be reconstructed by mixing T. durum (AB) and Aegilops squarrosa (D). The low intragenomic variability of CM protein is confirmed.

Chromosomal control of non-gliadin proteins from the 70% ethanol extract of wheat endosperm

SummaryThe non-gliadin fraction of the 70% ethanol extracts of compensated nulli-tetrasomics and ditelosomics of Triticum aestivum cv. Chinese Spring has been analyzed by combined electrofocusing and electrophoresis. Seventeen of the 21 protein map components of the euploid have been ascribed to eight chromosomes: 4Aβ, 3BS, 6BS, 7BS, 3Dβ, 4D, 5D and 7DS.The relationship of the different map components with other proteins previously associated with the same chromosomes is discussed.

Homeologous proteins synthesis controlled by homeologous chromosomes in wheat.

Abstract Two homeologous proteins have been isolated from the endosperm of common wheat (genomes ABD). Synthesis of these two proteins is controlled by the homeologous chromosomes 7B and 7D respectively. However, Aegilops speltoides , the more generally accepted B genome donor, does not synthesize the 7B protein.

Inhibition of eukaryotic cell-free protein synthesis by thionins from wheat endosperm

Thionins are polypeptide toxins of about 5000 molecular weight, present in the endosperms of many Gramineae, which modify membrane permeability and inhibit macromolecular synthesis in cultured mammalian cells. Evidence is presented that they inhibit in vitro protein synthesis at micromolar concentrations in cell-free systems derived from wheat germ or from rabbit reticulocytes. Inhibition seems to occur by direct binding of mRNA by the toxin, as judged by the ability of thionins to mediate retention of RNA in nitrocellulose filters and by the dependence of inhibitory concentrations on the amount of exogenous RNA added to the wheat-germ translation system. Commercial preparations of wheat-germ have been found to include some endosperm contamination (up to 15%), which may result in at least partially inhibitory concentrations of the toxin in the cell-free extracts.

Chromosomal assignment of genes controlling salt-soluble proteins (albumins and globulins) in wheat and related species.

SummarySalt-soluble proteins from the endosperms of wheat, barley, and rye have been separated by nonequilibrium electrofocusing x electrophoresis. Genes encoding 14 of the 25 components observed in wheat have been unambiguously assigned to 10 different chromosomes (1B, 3B, 3D, 4A, 4D, 5B, 6B, 6D, 7B, 7D) by analysis of the compensated nulli-tetrasomic series. Five more wheat proteins seem to be controlled by group 2 chromosomes. Analysis of wheat-barley and wheat-rye addition lines has led to the location of genes for 6 out of 20 barley proteins in 4 different chromosomes (1H, 3H, 4H, 6H; 1H is homoeologous to group 7 chromosomes of wheat) and of genes for 5 out of 20 rye proteins in two different chromosomes (2R, 4R). The relationship between the proteins reported here and previously characterized ones is discussed.

Characterization of cDNA clones of the family of trypsin/α-amylase inhibitors (CM proteins) in barley (Hordeum vulgare L.)

SummaryRecombinants encoding members of the trypsin/α-amylase inhibitors family (also designated CM-proteins) were selected from a cDNA library prepared from developing barley endosperm. Inserts in two of the clones, pUP-13 and pUP-38, were sequenced and found to encode proteins which clearly belong to this family, as judged from the extensive homology of the deduced sequences with that of the barley trypsin inhibitor CMe, the only member of the group for which a complete amino acid sequence has been obtained by direct protein sequencing. These results, together with previously obtained N-terminal sequences of purified CM-proteins, imply that there are at least six different members of this dispersed gene family in barley. The relationship of this protein family to the B-3 hordein and to reserve prolamins from related species is discussed in terms of their genome structure and evolution.

Endosperm sterol phenotype and germination in wheat

Abstract Free and conjugated sterols of endosperm, coats, scutellum, coleoptile and roots have been analysed at different germination stages in two wheat cultivars with different endosperm sterol phenotypes. It seems that sterol metabolism of the developing tissues, namely coleoptile and roots, is not affected by the sterol conjugation profile of the endosperm. Enough sterol is present in the mature embryo to supply the germinating axis during the observation period (144 hr at 16°). The data suggest that sterol is transferred from scutellum to coleoptile and roots during germination.

Biochemical evidence of chromosome homoeology among related plant genera

Abstract Biochemical markers associated with homoelogous chromosome groups 3 and 7 of Triticum aestivum L. have been investigated in genetic stocks carrying chromosomes or chromosomal segments of the same homoeology groups from Agropyron elongatum and Secale cereale. Chromosomes 3Ag of Agropyron and 3R of Secale control proteins a3 and b3 with the same properties as proteins 5, 6 and 7 associated with 3B and 3D of Triticum. It is concluded that genes for proteins 5, a3 and b3 are located in segments proximal to the centromere in the β arms of chromosomes 3D and 3Ag, respectively. Proteins 3, 4 and 11, controlled by 7D-short arm of Triticum, are replaced by proteins a7, b7 and c7, when that chromosome is replaced by 7 Ag. Genes for these proteins are located proximal to the centromere in the short arms of chromosomes 7D and 7 Ag. Finally, a gene that controls sterol esterification is similarly located in the short arms of chromosomes 7D and 7 Ag.