Interaction between tobacco Ribulose-l,5-biphosphate Carboxylase/Oxygenase large subunit (RubisCO-LSU) and the PVY Coat Protein (PVY-CP)

A 54 kDa band (P54) was continually detected with the 30 kDa viral capsid protein (CP) on the SDS-PAGE migration profile of purified potato virus Y (PVY). P54 was observed following the use of two different procedures for the purification of the PVY from infected tobacco. It was a constitutively expressed tobacco protein. The analysis of the PVY preparation showed that P54 has aggregation properties and precipitates, thus pulling down the virus. We used an enzyme-linked immunosorbent assay (ELISA) to study the relationship between P54 and the PVY particles. We performed an inhibition test with monoclonal antibodies (mAb) directed against the PVY-CP, to show that these two components interact. This result was confirmed by western blot. The internal sequence of five major peptides, obtained by C-lysine endoprotease digestion of the P54 followed by HPLC separation, showed 100% homology with the large subunit of the ribulose-1,5-biphosphate carboxylase/oxygenase (RubisCO-LSU) of tobacco. MAb directed against RubisCO-LSU were produced and used to reveal the RubisCO-LSU/PVY complex in infected tobacco extracts. A phage library displaying random heptapeptides was used to isolate several peptides that specifically bound to the native form of the PVY. The sequences of thirty-three phage-displayed peptides, which bound specifically to this virus, present further discontinuous sequence homologies with the RubisCO-LSU. Five peptides (p1 to p5) corresponding to the RubisCO-LSU homologous regions were used for a bacterial two-hybrid system to confirm in vivo direct interactions between the selected RubisCO-LSU regions and the PVY-CP. We propose that the PVY-CP may be involved in the production of mosaics and yellowing symptoms in tobacco through its interaction with RubisCO-LSU.     

Microbial Diversity and Phage–Host Interactions in the Georgian Coastal Area of the Black Sea Revealed by Whole Genome Metagenomic Sequencing

Viruses have the greatest abundance and highest genetic diversity in marine ecosystems. The interactions between viruses and their hosts is one of the hot spots of marine ecology. Besides their important role in various ecosystems, viruses, especially bacteriophages and their gene pool, are of enormous interest for the development of new gene products with high innovation value. Various studies have been conducted in diverse ecosystems to understand microbial diversity and phage–host interactions; however, the Black Sea, especially the Eastern coastal area, remains among the least studied ecosystems in this regard. This study was aimed at to fill this gap by analyzing microbial diversity and bacteriophage–host interactions in the waters of Eastern Black Sea using a metagenomic approach. To this end, prokaryotic and viral metagenomic DNA from two sampling sites, Poti and Gonio, were sequenced on the Illumina Miseq platform and taxonomic and functional profiles of the metagenomes were obtained using various bioinformatics tools. Our metagenomics analyses allowed us to identify the microbial communities, with Proteobacteria, Cyanobacteria, Actinibacteria, and Firmicutes found to be the most dominant bacterial phyla and Synechococcus and Candidatus Pelagibacter phages found to be the most dominant viral groups in the Black Sea. As minor groups, putative phages specific to human pathogens were identified in the metagenomes. We also characterized interactions between the phages and prokaryotic communities by determining clustered regularly interspaced short palindromic repeats (CRISPR), prophage-like sequences, and integrase/excisionase sequences in the metagenomes, along with identification of putative horizontally transferred genes in the viral contigs. In addition, in the viral contig sequences related to peptidoglycan lytic activity were identified as well. This is the first study on phage and prokaryote diversity and their interactions in the Eastern coastal area of the Black Sea using a metagenomic approach.     

Complete chloroplast DNA sequences of Georgian indigenous polyploid wheats (<Emphasis Type="Italic">Triticum</Emphasis> spp.) and B plasmon evolution

Three types of plasmon (A, B and G) are present for genus Triticum. Plasmon B is detected in polyploid species - Triticum turgidum L. and Triticum aestivum L. By now, 21 complete sequences of chloroplast DNA of the genus Triticum is published by different authors. Many inaccuracies can be detected in the sequenced chloroplast DNAs. Therefore, we found it necessary to study of plasmon B evolution to use only those sequences obtained by our method in our laboratory. Complete nucleotide sequences of chloroplast DNA of 11 representatives of Georgian wheat polyploid species were determined. Chloroplast DNA sequencing was performed on an Illumina MiSeq platform. Chloroplast DNA molecules were assembled using the SOAPdenovo computer program. Using T. aestivum L. subsp. macha var. palaeocolchicum as a reference, 5 SNPs were identified in chloroplast DNA of Georgian indigenous polyploid wheats. 38 and 56 bp inversions were observed in paleocolchicum subspecies. The phylogeny tree shows that subspecies macha, durum, carthlicum and palaeocolchicum occupy different positions. According the simplified scheme based on SNP and indel data the ancestral, female parent of all studied polyploid wheats is an unknown X predecesor, from which four lines were formed.