Ribosomal gene variation in soybean (Glycine) and its relatives

Summary The genes encoding the 18S25S ribosomal RNA gene repeat in soybean (Glycine max) and its relatives in the genus Glycine are surveyed for variation in repeat length and restriction enzyme site locations. Within the wild species of subgenus Glycine, considerable differences in repeat size occur, with a maximum observed in G. falcata. Repeat length and site polymorphisms occur in several species, but within individual plants only single repeat types are observed. The rDNA of the cultivated soybean and its wild progenitor, G. soja are identical at the level of this study, and no variation is found in over 40 accessions of the two species. Data from rDNA mapping studies are congruent with those of previous biosystematic studies, and in some instances give evidence of divergences not seen with other approaches.     

Protein trafficking in plant cells

The cells of higher plants contain distinct subcellular compartments (organelles) that perform specialized functions such as photosynthesis, carbohydrate and lipid metabolism, and so forth. The majority of the protein constituents of plant organelles are formed as cytosolic precursors with N-terminal extensions that direct transport across one or more membrane bilayers in a post- or co-translational fashion. Since the majority of proteins in plant cells are products of nuclear gene expression, there must be precise sorting mechanisms in the cytoplasm that direct proteins to their correct cellular locations. Based on recent studies of protein targeting to chloroplasts and vacuoles, the details of these intracellular sorting mechanisms are becoming clear. The ability to direct proteins to specific compartments within cells provides new opportunities for improvement of plants by genetic manipulation.     

Studies of coat protein-mediated resistance to tobacco mosaic tobamovirus: correlation between assembly of mutant coat proteins and resistance.

Coat protein-mediated resistance (CP-MR) has been widely used to protect transgenic plants against virus diseases. To characterize the mechanisms of CP-MR to tobacco mosaic tobamovirus (TMV) we developed mutants of the coat protein that affected subunit-subunit interactions. Mutant CPs were expressed during TMV replication as well as in transgenic Nicotiana tabacum plants. The mutation T42-->W increased protein aggregation and T28-->W abolished aggregation and assembly, while the mutations T28-->W plus T42-->W and T89-->W altered normal CP subunit-subunit interactions. The mutant T28W was unable to assemble virus-like particles (VLPs) during infection and in transgenic plants failed to aggregate; this protein conferred no protection against challenge of transgenic plants by TMV. The mutant T42W had strong CP subunit-subunit interactions and formed VLPs but not infectious virions. Transgenic lines with this protein exhibited stronger protection against TMV infection than transgenic plants that contained the wild-type (wt) CP. It is proposed that increased resistance conferred by the T42W mutant results from strong interaction between transgenic CP subunits and challenge virus CP subunits. CP carrying the mutation T89-->W formed flexuous and unstable VLPs whereas the double mutant T28W:T42W formed open helical structures that accumulated as paracrystalline arrays. In transgenic plants, T89W and the double mutant CPs showed reduced ability to aggregate and provided lower protection against TMV infection than wt CP. A strong correlation between normal CP subunit-subunit interactions and CP-MR is observed, and a model for CP-MR involving interactions between the transgenic CP and the CP of the challenge virus as well as interference with virus movement is discussed.     

In vitro synthesis of the Bowman-Birk and related soybean protease inhibitors

Poly (A)⁺ RNAs from immature soybean seeds were size fractionated in denaturing sucrose gradients to identify mRNA that directs the cell-free synthesis of the Bowman-Birk protease inhibitor and the related inhibitors PI I–IV. Polypeptides synthesized in vitro were labeled with (³⁵S)-cysteine and (³H)-serine and detected by immunoprecipitation with anti Bowman-Birk and anti PI I–IV sera. Immunoprecipitates of the translation products comigrated on SDS-polyacrylamide gels with the dimeric or trimeric aggregates of the authentic inhibitor proteins, which self-associate under certain conditions. Further evidence that these immunoprecipitates contained authentic polypeptides corresponding to the Bowman-Birk or PI IV inhibitor was shown by sequential amino acid analyses of peptides generated by cleavage with cyanogen bromide.     

In Vitro Synthesis of the alpha and alpha' Subunits of the 7S Storage Proteins (Conglycinin) of Soybean Seeds

Messenger RNAs (mRNAs), isolated from immature soybean (Glycine max L., Merr.) seeds, that bound to oligo(dT)-cellulose were fractionated by centrifugation in sucrose density gradients containing dimethyl sulfoxide. mRNAs with sedimentation values between 21S and 25S coded for the in vitro translation of polypeptides with electrophoretic mobilities similar to those of the α′ and α subunits of the 7S seed storage protein. High pressure liquid chromatographic analyses of the trypsin-induced fragments (“column fingerprinting”) verified that the polypeptides produced in vitro were closely related to authentic α′ and α subunits.     

Engineered resistance against plant virus diseases

The development of genetic engineering techniques has enabled the production of transgenic plants that are resistant to viral diseases. Expressing the coat protein (CP) gene of a virus in Iransgenic plants confers resistance against the virus from which the gene was isolated, and to other closely related strains and viruses. This approach has been demonstrated to be effective in conferring protection against viruses from different virus groups including alfalfa mosaic virus, cucumovirus. ilarvirus, potex-virus, potyvirus, tobamovirus and tobravirus. The data available indicate that several factors may affect the efficiency of the protection obtained including the level of the CP in the transgenic plants, the plant in which the CP gene is expressed and enviromental conditions. These and other aspects of coat protein mediated resistance are discussed.