Variability and Phylogeny of the Pepper Phytoene Synthase Paralogs PSY1 and PSY2 in Species of Various Capsicum Complexes.

Genes homologous to PSY1 and PSY2 that encode phytoene synthase isoforms in Capsicum species C. baccatum, C. chinense, C. frutescens, C. tovarii, C. eximium, and C. chacoense were identified. High conservatism of functionally significant sites of phytoene synthases of the analyzed accessions was revealed. It was found that only PSY1-based clustering of pepper species corresponds to the traditional Capsicum phylogeny; C. eximium was a part of the Purple corolla complex, and C. chacoense was equidistant from Annuum and Baccatum clades. The absence of significant differences between PSY1 and PSY2 of yellow-fruited C. chinense and red-fruited pepper accessions was shown. The yellow color of C. chinense fruit may be the result of both decreased PSY1 expression and increased PSY2 transcription. Thus, it was demonstrated that the acquired fruit pigmentation retains strict phylogenetic limitations, which, however, can be overcome using artificial selection for the activity of phytoene synthase PSY1.

Structural and functional features of phytoene synthase isoforms PSY1 and PSY2 in pepper Capsicum annuum L. cultivars.

The fruits of various pepper cultivars are characterized by a different color, which is determined by the pigment ratio; carotenoids dominate in ripe fruits, while chlorophylls, in immature fruits. A key regulator of carotenoid biosynthesis is the phytoene synthase encoded by the PSY gene. The Capsicum annuum genome contains two isoforms of this enzyme, localized in leaf (PSY2) and fruit (PSY1) plastids. In this work, the complete PSY1 and PSY2 genes were identified in nine C. annuum cultivars, which differ in ripe fruit color. PSY1 and PSY2 sequence variability was 2.43 % (69 SNPs) and 1.21 % (36 SNPs). The most variable were PSY1 proteins of the cultivars 'Maria' (red-fruited) and 'Sladkij shokolad' (red-brown-fruited). All identified PSY1 and PSY2 homologs contained the phytoene synthase domain HH-IPPS and the transit peptide. In the PSY1 and PSY2 HH-IPPS domains, functionally significant sites were determined. For all accessions studied, the active sites (YAKTF and RAYV), aspartate-rich substrate-Mg2+-binding sites (DELVD and DVGED), and other functional residues were shown to be conserved. Transit peptides were more variable, and their similarity in the PSY1 and PSY2 proteins did not exceed 78.68 %. According to the biochemical data obtained, the largest amounts of chlorophylls and carotenoids across the cultivars studied were detected in immature and ripe fruits of the cv. 'Sladkij shokolad' and 'Shokoladnyj'. Also, ripe fruits of the cv. 'Nesozrevayuschij' (green-fruited) were marked by significant chlorophyll content, but a minimum of carotenoids. The PSY1 and PSY2 expression patterns were determined in the fruit pericarp at three ripening stages in 'Zheltyj buket', 'Sladkij shokolad', 'Karmin' and 'Nesozrevayuschij', which have different ripe fruit colors: yellow, red-brown, dark red and green, respectively. In the leaves of the cultivars studied, PSY1 expression levels varied significantly. All cultivars were characterized by increased PSY1 transcription as the fruit ripened; the maximum transcription level was found in the ripe fruit of 'Sladkij shokolad', and the lowest, in 'Nesozrevayuschij'. PSY2 transcripts were detected not only in the leaves and immature fruits, but also in ripe fruits. Assessment of a possible correlation of PSY1 and PSY2 transcription with carotenoid and chlorophyll content revealed a direct relationship between PSY1 expression level and carotenoid pigmentation during fruit ripening. It has been suggested that the absence of a typical pericarp pigmentation pattern in 'Nesozrevayuschij' may be associated with impaired chromoplast formation.

Identification and Expression Analysis of Chitinase Genes in Pitchers of Nepenthes sp. during Development.

Fifteen chitinases of classes I-V were identified in the transcriptomes of pitchers and adult leaves of the carnivorous plant Nepenthes sp. Ten of these chitinases were identified for the first time, including the chitinases of classes II and V. The expression levels of all found chitinase genes in leaves and at three stages of pitcher development were determined. The maximum level of transcriptional activity in an open pitcher was observed for the genes encoding chitinase NChi4 (class II) and its isoforms. The expression levels of these genes significantly increased as the pitcher developed. In addition, for the first time, transcription of the genes encoding chitinases of all five classes was detected in the leaves of this plant.

Coexpression of YABBY1 and YABBY3 Genes in Lateral Organs of Tomato Species (Solanum, Section Lycopersicon).

The interspecific polymorphism and the expression patterns of the genes encoding the YABBY1 and YABBY3 transcription factors of cultivated tomato Solanum lycopersicum and wild species S. chmielewskii, S. peruvianum, and S. habrochaites are characterized. The possibility of an inverse relationship between the level of YABBY1 and YABBY3 genes coexpression and the size of leaves and flowers of the tomato species studied is shown. The phylogeny of the genes suggests an earlier emergence of YABBY1 compared to YABBY3 as well as the origin of these paralogs from a common ancestor before the divergence of dicotyledonous plants into Rosids and Asterids.

[Identification and expression analysis of receptor-like kinase gene ERECTA in mycoheterotrophic plant Monotropa hypopitys].

The precise spatial-temporal coordination of cell division and differentiation is necessary for the correct formation of tissues, organs, and the organism as a whole. This coordination has been implemented by the intercellular communication mediated by signaling molecules and receptors that selectively recognize them. Membrane receptor kinases of ERECTA family regulate inflorescence and flower structure, the formation of root epidermis and adaptation responses. The characterization of the ERECTA genes of flowering plant pinesap Monotropa hypopitys with unique development features can enrich the knowledge about the kinase ERECTA functions and conserved development processes with their participation. Transcriptomic and genomic search with the subsequent structural-phylogenetic analysis identified the mRNA of a gene of serine-threonine kinase receptor with leucine-rich repeats of MhyERL1, which is the only ortholog of the ERECTA family kinases of pinesap. A quantitative analysis of the MhyERL1 gene transcripts has revealed its expression in all analyzed pinesap tissues with maximum levels in the flowers. MhyERL1 is probably involved in defining the inflorescence and flower architecture, and the formation of the pinesap root epidermis. The cascades involving ERL1 are apparently conserved. The exception are pathways associated with the development of above-ground vegetative structures, and the immune response to fungal pathogens probably lost in the process of the pinesap adaptation to unfavorable environmental conditions.

Identification and characterization of the flower meristem identity gene MhyLFY in mycoheterotrophic plant Monotropa hypopitys.

The gene encoding the transcription factor LEAFY was identified in the genome of the mycoheterotrophic plant, pinesap Monotropa hypopitys. In the transcriptomes of roots, bracts, and flowers of flowering pinesaps, the MhyLFY gene expression was absent. These data suggest the conservativeness of the LFY-dependent mechanism of flower meristem identity and flower formation in heterotrophic species with some differences associated to the specificity of development and the structure of such plants. The pinesap flowering under the control of the transcription factor MhyLFY may be initiated either in an embryonic inflorescence during spring dormancy release of adventitious root buds or in an inflorescence of a growing reproductive stem after photoperiodic induction.

Identification and expression analysis of chitinase genes in parasitic plant Monotropa hypopitys.

Genes encoding six chitinases, five of which belong to classes I (MhCHI3 and MhCHI4), IV (MhCHI1), V (MhCHI5), and VII (MhCHI2), were identified in the transcriptome of the parasitic mixoheterotrophic plant Monotropa hypopitys. The transcription level of MhCHI5 and MhCHI1 was low; however, in the leaves (bracts) and roots it was higher than in flowers. MhCHI4 transcripts were detected primarily in the flowers and were almost absent in the roots, whereas the expression level of MhCHI3 was relatively high in all organs but maximum in the leaves (bracts).

YABBY3-Orthologous Genes in Wild Tomato Species: Structure, Variability, and Expression.

Evolution of the genes encoding YABBY transcription factors is believed to be one of the key reasons for flat leaf emergence from the radially symmetrical stem and gynoecium diversity. YABBY genes determine the identity of the abaxial surface of all aboveground lateral organs in seed plants. In the present study, complete sequences of YABBY3-orthologous genes were identified and characterized in 13 accessions of cultivated and wild tomato species with diverse morphophysiology of leaves, flowers, and fruits. The obtained gene sequences showed high homology (95-99%) and an identical exon-intron structure with the known S. lycopersicum YABBY3 gene, and they contained sequences that encode the conserved HMG-like YABBY and Cys2Cys2-zinc-finger domains. In total, in the analyzed YABBY3 genes, 317 variable sites were found, wherein 8 of 24 exon-specific SNPs were nonsynonymous. In the vegetative and reproductive organs of red-fruited and green-fruited tomato species, YABBY3 gene expression was similar to that in S. pimpinellifolium described earlier, but it demonstrated interspecies differences at the leaf-, bud- and flower-specific expression levels.

[Analysis of the ITS1/ITS2 nuclear spacers and the secondary structure of 5.8S rRNA gene in endemic species Bellevalia sarmatica (Pall. ex Georgi) Woronow and related species of the subfamily scilloideae].

Sequence variability of the ITS spacers and 5.8S rRNA gene was examined in 11 accessions of the subfamily Scilloideae, including seven accessions of rare and endangered species Bellevalia sarmatica from Volgograd region. The intraspecific polymorphism level of the examined ITS1­5.8S­ITS2 sequence of B. sarmatica accessions constituted 1.3%. The phylogenetic position of B. sarmatica within the genus Bellevalia was determined. It was demonstrated that B. sarmatica belonged to the section Nutantes, and the most closely related species were B. webbiana and B. dubia. Nucleotide substitutions in the 5.8S rRNA gene sequence of the analyzed Scilloideae accessions were identified and studied. The predicted secondary structure of 5.8S rRNA gene was constructed. It was demonstrated that in the examined accessions, mutations in the 5.8S rRNA gene were mainly localized in the third hairpin region and had no effect on the secondary structure of the 5.8S rRNA molecule.

5.8S rRNA sequence and secondary structure in Monotropa hypopitys and related Ericaceae species.

This is the first study to investigate the secondary structure of 5.8S rRNA in M. hypopitys and related Ericaceae species. The identified nucleotide substitutions are localized mostly at the 3' and 5' ends of the gene, in the region of the third hairpin, and do not significantly affect the topology of the secondary structure of the 5.8S rRNA molecule.

[Intraspecific Variability of ITS Sequences in the Parasitic Plant Monotropa hypopitys L. from the European Russian Populations].

Intraspecific sequence polymorphism in the ITS region of the nuclear ribosomal operon was examined in ten populations of the parasitic plant Monotropa hypopitys from Belgorod oblast, Vologda oblast, and Kaluga oblast of Russia. Based on an analysis of the identified nucleotide substitutions and indels in the ITS regions, the M. hypopitys populations were divided into two groups (A and B). Four new ITS haplotypes of M. hypopitys were identified.

[5.8S rDNA variability in Allium species belonging to the third evolutionary group].

Sequence analysis of 5.8S rDNA in 67 accessions of the subgenus Allium and six other subgenera belonging to the third evolutionary group of Allium genus (Friesen et al., 2006) was performed. Nucleotide substitutions in 5.8S rDNA sequences ofAllium accessions were identified and, studied for the first time. The probable secondary structure of 5.8S rRNA was constructed. It was shown that mutations in 5.8S rDNA do not involve conserved motifs, and they did not significantly affect the Secondary structure of the RNA molecule in Allium accessions.