Genome-wide identification and evolution of the tubulin gene family in Camelina sativa.

BACKGROUND: Tubulins play crucial roles in numerous fundamental processes of plant development. In flowering plants, tubulins are grouped into α-, ß- and γ-subfamilies, while α- and ß-tubulins possess a large isotype diversity and gene number variations among different species. This circumstance leads to insufficient recognition of orthologous isotypes and significantly complicates extrapolation of obtained experimental results, and brings difficulties for the identification of particular tubulin isotype function. The aim of this research is to identify and characterize tubulins of an emerging biofuel crop Camelina sativa. RESULTS: We report comprehensive identification and characterization of tubulin gene family in C. sativa, including analyses of exon-intron organization, duplicated genes comparison, proper isotype designation, phylogenetic analysis, and expression patterns in different tissues. 17 α-, 34 ß- and 6 γ-tubulin genes were identified and assigned to a particular isotype. Recognition of orthologous tubulin isotypes was cross-referred, involving data of phylogeny, synteny analyses and genes allocation on reconstructed genomic blocks of Ancestral Crucifer Karyotype. An investigation of expression patterns of tubulin homeologs revealed the predominant role of N6 (A) and N7 (B) subgenomes in tubulin expression at various developmental stages, contrarily to general the dominance of transcripts of H7 (C) subgenome. CONCLUSIONS: For the first time a complete set of tubulin gene family members was identified and characterized for allohexaploid C. sativa species. The study demonstrates the comprehensive approach of precise inferring gene orthology. The applied technique allowed not only identifying C. sativa tubulin orthologs in model Arabidopsis species and tracking tubulin gene evolution, but also uncovered that A. thaliana is missing orthologs for several particular isotypes of α- and ß-tubulins.

Autophagy formation, microtubule disorientation, and alteration of ATG8 and tubulin gene expression under simulated microgravity in Arabidopsis thaliana.

Autophagy plays an important role in plant growth and development, pathogen invasion and modulates plant response and adaptation to various abiotic stress stimuli. The biogenesis and trafficking of autophagosomes involve microtubules (MTs) as important actors in the autophagic process. However, initiation of autophagy in plants under microgravity has not been previously studied. Here we demonstrate how simulated microgravity induces autophagy development involving microtubular reorganization during period of autophagosome formation. It was shown that induction of autophagy with maximal autophagosome formation in root cells of Arabidopsis thaliana is observed after 6 days of clinostating, along with MT disorganization, which leads to visible changes in root morphology. Gradual decrease of autophagosome number was indicated on 9th and 12th days of the experiment as well as no significant re-orientation of MTs were identified. Respectively, analysis of α- and ß-tubulins and ATG8 gene expression was carried out. In particular, the most pronounced increase of expression on both 6th and 9th days in response to simulated microgravity was detected for non-paralogous AtATG8b, AtATG8f, AtATG8i, and AtTUA2, AtTUA3 genes, as well as for the pair of ß-tubulin duplicates, namely AtTUB2 and AtTUB3. Overall, the main autophagic response was observed after 6 and 9 days of exposure to simulated microgravity, followed by adaptive response after 12 days. These findings provide a key basis for further studies of cellular mechanisms of autophagy and involvement of cytoskeletal structures in autophagy biogenesis under microgravity, which would enable development of new approaches, aimed on enhancing plant adaptation to microgravity.

Overcoming genetic paucity of Camelina sativa: possibilities for interspecific hybridization conditioned by the genus evolution pathway.

Camelina or false flax (Camelina sativa) is an emerging oilseed crop and a feedstock for biofuel production. This species is believed to originate from Western Asian and Eastern European regions, where the center of diversity of the Camelina genus is located. Cultivated Camelina species arose via a series of polyploidization events, serving as bottlenecks narrowing genetic diversity of the species. The genetic paucity of C. sativa is foreseen as the most crucial limitation for successful breeding and improvement of this crop. A potential solution to this challenge could be gene introgression from Camelina wild species or from resynthesized allohexaploid C. sativa. However, both approaches would require a complete comprehension of the evolutionary trajectories that led to the C. sativa origin. Although there are some studies discussing the origin and evolution of Camelina hexaploid species, final conclusions have not been made yet. Here, we propose the most complete integrated evolutionary model for the Camelina genus based on the most recently described findings, which enables efficient improvement of C. sativa via the interspecific hybridization with its wild relatives. We also discuss issues of interspecific and intergeneric hybridization, aimed on improving C. sativa and overcoming the genetic paucity of this crop. The proposed comprehensive evolutionary model of Camelina species indicates that a newly described species Camelina neglecta has a key role in origin of tetra- and hexaploids, all of which have two C. neglecta-based subgenomes. Understanding of species evolution within the Camelina genus provides insights into further research on C. sativa improvements via gene introgression from wild species, and a potential resynthesis of this emerging oilseed crop.

Genome-wide identification and analysis of the cytokinin oxidase/dehydrogenase (ckx) gene family in finger millet (Eleusine coracana).

Cytokinin dehydrogenase/oxidase (CKX) enzymes play a key role in regulating cytokinin (CK) levels in plants by degrading the excess of this phytohormone. CKX genes have proven an attractive target for genetic engineering, as their silencing boosts cytokinin accumulation in various tissues, thereby contributing to a rapid increase in biomass and overall plant productivity. We previously reported a similar effect in finger millet (Eleusine coracana) somaclonal lines, caused by downregulation of EcCKX1 and EcCKX2. However, the CKX gene family has numerous representatives, especially in allopolyploid crop species, such as E. coracana. To date, the entire CKX gene family of E. coracana and its related species has not been characterized. We offer here, for the first time, a comprehensive genome-wide identification and analysis of a panel of CKX genes in finger millet. The functional genes identified in the E. coracana genome are compared with the previously-identified genes, EcCKX1 and EcCKX2. Exon-intron structural analysis and motif analysis of FAD- and CK-binding domains are performed. The phylogeny of the EcCKX genes suggests that CKX genes are divided into several distinct groups, corresponding to certain isotypes. Finally, the phenotypic effect of EcCKX1 and EcCKX2 in partially silencing the SE7 somaclonal line is investigated, showing that lines deficient in CKX-expression demonstrate increased grain yield and greater bushiness, enhanced biomass accumulation, and a shorter vegetation cycle.