Duplication and suppression of chloroplast protein translocation genes in maize

The HCF106 (high chlorophyll fluorescence) gene of maize encodes a chloroplast membrane protein required for translocation of a subset of proteins across the thylakoid membrane. Mutations in HCF106 caused by the insertion of Robertson's Mutator transposable elements have been mapped to chromosome 2S. Here we show that there is a closely related homolog of HCF106 encoded elsewhere in the maize genome (HCF106c) that can partially compensate for these mutations. This homolog maps on chromosome 10L and is part of the most recent set of segmental duplications in the maize genome. Triple mutants that are disrupted in both the HCF106 and Sec-dependent protein translocation pathways provide evidence that they act independently. The HCF106c gene accounts for a previously reported exception to the correlation between epigenetic suppression of hcf106 and methylation of Mutator transposons. We also demonstrate that insertions of Robertson's Mutator elements into either introns or promoters can lead to mutations whose phenotypes are suppressed in the absence of Mu activity, while alleles with insertions in both positions are not suppressed. The implications of these observations are discussed.     

Evolution of seed storage protein genes: Legumin genes of Ginkgo biloba

Legumin-like seed storage proteins have been intensively studied in crop plants. However, little is known about the molecular evolution of these proteins and their genes and it was assumed that they originated from an ancestral gene that already existed at the beginning of angiosperm evolution. We have evidence for the ubiquitous occurrence of homologous proteins in gymnosperms as well. We have characterized the major seed storage globulin from Ginkgo biloba by amino acid sequencing, which reveals clear homology to legumin-like proteins from angiosperms. The Ginkgo legumin is encoded by a gene family; we describe two of its members. The promoter regions contain sequence motifs which are known to function as regulatory elements involved in seed-specific expression of angiosperm legumins, although the tissues concerned are different in gymnosperms and angiosperms. The Ginkgo legumin gene structure is divergent from that of angiosperms and suggests that the evolution of legumin genes implicated loss of introns. From our data and from functional approaches recently described it becomes obvious that the posttranslational processing site of legumin precursors is less conserved than hitherto assumed. Finally, we present a phylogenetic analysis of legumin encoding sequences and discuss their utility as molecular markers for the reconstruction of seed plant evolution.Correspondence to: K.-P. Häger     

Effects of sulfur nutrition on expression of the soybean seed storage protein genes in transgenic petunia

The 7S seed storage protein (β-conglycinin) of soybean (Glycine max [L]. Merr.) has three major subunits; α, α′, and β. Accumulation of the β-subunit, but not the α- and α′-subunits, has been shown to be repressed by exogenously applied methionine to the immature cotyledon culture system (LP Holowach, JF Thompson, JT Madison [1984] Plant Physiol 74: 576-583) and to be enhanced under sulfate deficiency in soybean plants (KR Gayler, GE Sykes [1985] Plant Physiol 78: 582-585). Transgenic petunia (Petunia hybrida) harboring either the α′- or β-subunit gene were constructed to test whether the patterns of differential expression were retained in petunia. Petunia regulates these genes in a similar way as soybean in response to sulfur nutritional stimuli, i.e. (a) expression of the β-subunit gene is repressed by exogenous methionine in in vitro cultured seeds, whereas the α′-subunit gene expression is not affected; and (b) accumulation of the β-subunit is enhanced by sulfur deficiency. The pattern of accumulation of major seed storage protein of petunia was not affected by these treatments. These results indicate that this mechanism of gene regulation in response to sulfur nutrition is conserved in petunia even though it is not used to regulate its own major seed storage proteins.     

Chromosomal location of seed storage protein genes in the genome ofDasypyrum villosum (L.) Candargy

Summary Genes coding for glutenin-like subunits and for several prolamin subunits with electrophoretic mobilities (lactate-PAGE) corresponding to those of omega- and gamma-gliadins of wheat were located inDasypyrum villosum chromosome1V. Genes controlling four gliadinlike subunits with electrophoretic mobilities corresponding to those of alpha- and gamma-gliadins were located on the short arm of chromosome6V and on the long arm of chromosome4V. N-terminal amino acid sequences of these four components were also determined and homology with alpha-type gliadins was demonstrated. The presence of genes coding for glutenin- and gliadin-like subunits on chromosomes1V and6V demonstrates homoeology between theD. villosum chromosomes1V and6V and the chromosomes of homoeologous groups 1 and 6 in wheat. It is likely that the additional locusGli-V3 on chromosome4V originated by translocation from theGli-V2 locus.     

Interactions between tassel seed genes and other sex determining genes in maize

The tassel seed mutations of maize cause sex reversal of the florets of the tassel, such that the normally staminate florets develop pistils. Although these mutations have been recognized for many years, little is known about how they act. We have tested the hypothesis that the tassel seed genes interact directly with each other and with other genes controlling sex determination in a single genetic pathway by the construction and analysis of double mutants. On the basis of the phenotypes of the double mutants, the tassel seed mutations were placed into two groups: ts1, ts2, Ts5 and ts4, Ts6. Both groups of tassel seed mutations were additive with the masculinizing mutation dwarf, indicating independent modes of action. Interactions of tassel seed mutations with silkless varied, allowing the ordering of the action of the various tassel seed mutations relative to silkless. Both groups of tassel seed mutations were epistatic with regard to sex expression to mutations that alter both architecture of the plant and distribution of male and female florets, Teopod 1, terminal ear, and teosinte branched. Thus, there are at least two separate genetic pathways that control the sex of florets in maize tassels. In addition, analysis of double mutants revealec that all tassel seed genes tested play a role in the regulation of flower morphogenesis as well as pistil suppression. © 1994 Wiley-Liss, Inc.     

Asymmetric localization of seed storage protein RNAs to distinct subdomains of the endoplasmic reticulum in developing maize endosperm cells

Plant storage proteins are synthesized and stored in different compartments of the plant endomembrane system. Developing maize seeds synthesize and accumulate prolamin (zein) and 11S globulin (legumin-1) type proteins, which are sequestered in the endoplasmic reticulum (ER) lumen and storage vacuoles, respectively. Immunofluorescence studies showed that the lumenal chaperone BiP was not randomly distributed within the ER in developing maize endosperm but concentrated within the zein-containing protein bodies. Analysis of the spatial distribution of RNAs in maize endosperm sections by in situ RT-PCR showed that, contrary to the conclusions made in an earlier study [Kim et al. (2002) Plant Cell 14: 655-672], the zein and legumin-1 RNAs are not symmetrically distributed on the ER but, instead, targeted to specific ER subdomains. RNAs coding for 22 kDa alpha-zein, 15 kDa beta-zein, 27 kDa gamma-zein and 10 kDa delta-zein were localized to ER-bounded zein protein bodies, whereas 51 kDa legumin-1 RNAs were distributed on adjacent cisternal ER proximal to the zein protein bodies. These results indicate that the maize storage protein RNAs are targeted to specific ER subdomains in developing maize endosperm and that RNA localization may be a prevalent mechanism to sort proteins within plant cells.     

Chromosomal location of isozyme and seed storage protein genes in Dasypyrum villosum (L.) Candargy

Summary The zymogram phenotypes of glucose-phosphate isomerase (GPI), alcohol dehydrogenase-1 (ADH-1), glutamate oxaloacetate transaminase (GOT), superoxide dismutase (SOD), lipoxygenase (LPX), esterase (EST) and the banding patterns of gliadin and glutenin seed storage proteins were determined for Triticum aestivum cv. Chinese Spring (CS), Dasypyrum villosum, the octoploid amphiploid T. aestivum cv. Chinese Spring D. villosum (CS × v) (2n=8x=56; AABBDDVV), and for five CS-D. villosum disomic addition lines. The genes Gpi-V1, Adh-V1, Got-V2, and Sod-V2 coding for GPI-1, ADH-1, GOT-2, and SOD-2 isozymes were located in D. villosum on chromosome 1V, 4V, 6V, and 7V, respectively. Genes coding for gliadin- and glutenin-like subunits are located in D. villosum chromosomes 1V. There are no direct evidence for chromosomal location of genes coding for GOT-3, EST-1 and LPX-2 isozymes. The linkage between genes coding for glutenin-like proteins and GPI-1 isozymes in chromosome 1V is evidence of homoeology between chromosome 1V and the chromosomes of homoeologous group 1 in wheat.Research supported by the National Research Council (Italy) and National Science Foundation (USA). International cooperative project, Grant No. 85.01504.06 (CNR)     

Antigenic relationship of legume seed proteins to the 7S seed storage protein of soybean

Extracts enriched for globulin proteins were prepared from the seeds of a large number of legume species and were tested for homology to antisera prepared against the glycosylated 7S seed storage protein of the soybean (Glycine max). Electrophoretic identification and subsequent analysis of proteins precipitated with 7S antisera was useful at relatively short taxonomic distances, particularly within the tribe Phaseoleae, to which G. max belongs. Glycine and most other members of the subtribe Glycininae are unusual within the Phaseoleae in having high molecular weight ($\text{> 70 000}$ dalton) subunit polypeptides. Seeds from other plants representing other subtribes of the Phaseoleae also contained proteins that cross-reacted with the G. max antisera; the molecular weights of these proteins varied from 30 000 to nearly 90 000 daltons. Homology was detected across a wider range of legume tribes within the subfamily Papilionoideae by enzyme-linked immunosorbent assay (ELISA). The results of these experiments suggest both that the 7S proteins of these tribes are evolutionarily related and that at least some features of these apparently rapidly-evolving proteins are under relatively strong selectional constraint.     

A fern spore storage protein is genetically similar to the 1.7 S seed storage protein ofBrassica napus

The ostrich fern, Matteuccia struthiopteris L., contains two globulin spore storage proteins of 2.2 S and 11.3 S, with physical characteristics similar to those of seed storage proteins of Brassica napus (rapeseed) and Raphanus sativus (radish). By the use of a cloned cDNA that encodes the 1.7 S B. napus storage protein (napin), gene sequences that hybridized with napin were detected in fern nuclear DNA, and a 900-nucleotide homologous mRNA was detected in developing spores. In vitro translation of this fern mRNA produced a 22-kD polypeptide comparable in size to the 21-kD precursor polypeptide identified in Brassica. No hybridizations were observed between the Brassica 12 S clone and either fern DNA or developing spore mRNA.     

Characterization of seed storage protein patterns of Heliotropium digynum

Heliotropium digynum, is a shrub that has ecological importance. The height of the plant differs from one population to another and the difference in length of the inflorescence can be attributed to environmental factors, such as rainfall or type of soil and temperature. To date, no study has shed light on estimation in seed samples of H. digynum in Saudi Arabia. So, the aim is to evaluate and characterize the protein patterns of seed storage proteins of H. digynum to be used as fingerprint of this plant in Saudi Arabia. It is collected from different locations in the central region of Saudi Arabia and total protein extraction from plant was compared in SDS-PAGE. The genetic relationships among all cultivars were analyzed using UPGMA and NJ using Total Lab TL and in the same way using Jaccard Similarity Coefficient dendrogram using STATISTICA (ver.8) software. Results, our data show that amounts of protein are different, although they are of the same type or from the same geographical region. Amounts ranged between 22 and 1.5 mg/g of dry weight. Less amount of protein was obtained from the group of samples collected from Dir’iyah area, and the highest amount of protein was from the group of samples collected from Dyrab area in general.     

Genetic regulation of storage protein content in maize endosperm

Sodium dodecylsulfate-polyacrylamide gel electrophoresis reveals that zein prepared from normal maize inbred (Zea mays L.) contains six separable components. Z1 and Z2 are the predominant species, with molecular weights of 21,800 and 19,000 daltons. Amino acid analysis of these two components shows that both are rich in glutamic acid, leucine, and proline, but low in lysine. Of the four minor bands, Z3, Z4, Z5, and Z6, the latter two exist only in trace amounts. A mutation at the opaque-2 locus severely suppresses the synthesis of Z1. The nonallelic mutant, opaque-7, strongly suppresses the synthesis of Z3 and Z4, while slightly reducing Z2. On the other hand, the floury-2 mutant appears to reduce the synthesis of these six proteins in the same relative proportion. In the double mutant combinations, opaque-2 apparently is epistatic to opaque-7 and floury-2 in the synthesis of zein components. The glutelin fraction shows a more complex banding pattern; however, qualitative differences are not apparent among the mutant lines examined.This research was supported in part by a grant from the Lilly Endowment.Journal Paper No. 6100 of the Purdue University Agricultural Experimental Station.     

Polymorphism and inheritance of seed storage protein in sunflower

The data on polymorphism and inheritance of the seed storage protein helianthinin are presented. The results of hybrid analysis indicate that in the annual sunflower Helianthus annuus, helianthinin synthesis is controlled by at least three loci: HelA, HelB, HelB, and HelC. Codominant alleles controlling different electrophoretic variants of polypeptides were identified at each of the loci. The HelA locus was inherited independently of HelB and HelC in a series of dihybrid crosses. The frequencies of recombination between loci HelB and HelC estimated in F2 and BC of two crossing combinations were respectively 21.8 and 19.0%. Segregation of the Hel-C-controlled variants in the progenies from the crosses of cultured sunflower with annual wild species and forms corresponded to that theoretically expected for Mendelian inheritance. The maternal type of helianthinin inheritance was observed in the progenies from the crosses of inbred H. annuus lines with perennial diploid and polyploid Helianthus species. Altered expression of the HelC locus was detected in some hybrid combinations. These alterations appeared in early (F1, F2) hybrid generations and were similar in different hybrid combinations. They did not depend on the perennial paternal species being more influenced by the maternal genotype and by the mode of obtaining hybrids (in an embryo culture or in the field). These results are explained by "genomic shock" generated by hybridization of genetically incompatible species.