S-protein mutants indicate a functional role for SBP in the self-incompatibility reaction of Papaver rhoeas.

The self-incompatibility response involves S-allele specific recognition between stigmatic S proteins and incompatible pollen, resulting in S-specific pollen inhibition. In Papaver rhoeas, the pollen S gene product is predicted to be a receptor that interacts with the stigmatic S protein in an S specific manner. We recently identified an S protein binding protein (SBP) in pollen that binds stigmatic S proteins, although apparently not in an S-allele-specific manner. In order to investigate the functional significance of the interaction between S proteins and SBP, we constructed mutant derivatives of the S1 protein and tested their SBP-binding activity and their biological activity. Here we present an evaluation of nine mutant derivatives of the S1 protein. Western ligand blotting was used to show that mutations to amino acid residues in predicted loops 2 and 6 of the S1 protein cause significant reductions in their SBP-binding activity. These same mutants show a concomitant reduction in their ability to inhibit incompatible pollen. This establishes a direct link between SBP binding and inhibition of incompatible pollen and implicates SBP as a pollen component playing a key role in the self-incompatibility reaction. We discuss the possible nature of the contribution of SBP in the S-specific rejection of incompatible pollen.

Identification of residues in a hydrophilic loop of the Papaver rhoeas S protein that play a crucial role in recognition of incompatible pollen.

The self-incompatibility response involves S allele-specific recognition between stigmatic S proteins and incompatible pollen. This response results in pollen inhibition. Defining the amino acid residues within the stigmatic S proteins that participate in S allele-specific inhibition of incompatible pollen is essential for the elucidation of the molecular basis of the self-incompatibility response. We have constructed mutant derivatives of the S1 protein from Papaver rhoeas by using site-directed mutagenesis and have tested their biological activity. This has enabled us to identify amino acid residues in the stigmatic S proteins of P. rhoeas that are required for S-specific inhibition of incompatible pollen. We report here the identification of several amino acid residues in the predicted hydrophilic loop 6 of the P. rhoeas stigmatic S1 protein that are involved in the inhibition of S1 pollen. Mutation of the only hypervariable amino acid, which is situated in this loop, resulted in the complete loss of ability of the S protein to inhibit S1 pollen. This clearly demonstrates that this residue plays a crucial role in pollen recognition and may also participate in defining allelic specificity. We have also established the importance of highly conserved amino acids adjacent to this hypervariable site. Our studies demonstrate that both variable and conserved amino acids in the region of the S protein corresponding to surface loop 6 are key elements that play a role in the recognition and inhibition of incompatible pollen in the pollen-pistil self-incompatibility reaction.

A putative receptor protein kinase gene in Ipomoea trifida.

We have characterized a cDNA clone, IRK1, for a putative receptor kinase from a stigma cDNA library of Ipomoea trifida. IRK1 protein contains an extracellular receptor-like domain and the consensus sequences diagnostic of serine/threonine protein kinase. Both the pattern of gene expression and the results of RFLP analysis indicate that the IRK1 gene is not primarily involved in the self-incompatibility system of Ipomoea.

Molecular chaperon produced by an intracellular symbiont.

Symbionin, that is selectively produced by an intracellular symbiont harbored by the aphid bacteriocyte, is structurally homologous to the Escherichia coli groEL protein, a heat shock protein functioning as a molecular chaperon. It was shown that symbionin has ATPase activity and, in the presence of Mg-ATP, is converted into lower molecular mass species. Like the groEL protein, symbionin was able to reconstitute dimeric ribulose 1,5-bisphosphate carboxylase/oxygenase holoenzyme from its unfolded subunits in vitro, suggesting that this protein functions as a molecular chaperon in the endosymbiont. The groES-homologous protein did exist in the endosymbiont, but its amount was small relative to that of symbionin.

Heterochromatic differentiation in barley chromosomes revealed by C- and N-banding techniques.

Heterochromatin distribution in barley chromosomes was investigated by analyzing the C- and N-banding patterns of four cultivars. Enzymatic maceration and air drying were employed for the preparation of the chromosome slides. Although the two banding patterns were generally similar to each other, a clear difference was observed between them at the centromeric sites on all chromosomes. Every centromeric site consisted of N-banding positive and C-banding negative (N(+) C(-)) heterochromatin in every cultivar examined. An intervarietal polymorphism of heterochromatin distribution was confirmed in each of the banding techniques. The appearance frequencies of some bands were different between the two banding techniques and among the cultivars. The heterochromatic differentiation observed is discussed with respect to cause.

Identification and characterization of somatic rice chromosomes by imaging methods.

Somatic rice chromosomes from 30 spreads were analyzed by imaging methods. Morphological characters of each of the 12 rice chromosomes were obtained both by the imaging methods and by visual inspection. The numerical data of relative length, arm ratio, and condensation pattern (CP) were statistically analyzed. The descriptive morphological information obtained was also summarized into numbers of "key characters" or essential short sentences to characterize the traits. The fitness probability or the appearing frequencies of the key character for each of the 30 chromosomes was calculated. Altogether, 118 key characters were extracted to distinguish each rice chromosome. Furthermore, several "discriminants" or critical key characters were determined among the key characters, and a discrimination chart or flowchart to identify all the rice chromosomes was constructed using the discriminants.

High resolution bands in maize chromosomes by G-banding methods.

It was demonstrated that G-bands are unequivocally present in plant chromosomes, in contrast to what had been formerly believed by plant cytologists. Maize chromosomes prepared by an enzymatic maceration method and treated with trypsin or SDS showed clear G-bands spreading along the chromosomes. The most critical point during the G-banding procedures was the post-fixation with glutaraldehyde solution. Banding patterns were processed by using the chromosome image analyzing system and a clearer image was obtained. Gbanding technique and the image manipulation method described here can be applied to many plant species, and would contribute new information in the field of plant cytology and genetics.