Identification, Expression Analysis, and Target Prediction of Flax Genotroph MicroRNAs Under Normal and Nutrient Stress Conditions.
Cultivated flax (Linum usitatissimum L.) is an important plant valuable for industry. Some flax lines can undergo heritable phenotypic and genotypic changes (LIS-1 insertion being the most common) in response to nutrient stress and are called plastic lines. Offspring of plastic lines, which stably inherit the changes, are called genotrophs. MicroRNAs (miRNAs) are involved in a crucial regulatory mechanism of gene expression. They have previously been assumed to take part in nutrient stress response and can, therefore, participate in genotroph formation. In the present study, we performed high-throughput sequencing of small RNAs (sRNAs) extracted from flax plants grown under normal, phosphate deficient and nutrient excess conditions to identify miRNAs and evaluate their expression. Our analysis revealed expression of 96 conserved miRNAs from 21 families in flax. Moreover, 475 novel potential miRNAs were identified for the first time, and their targets were predicted. However, none of the identified miRNAs were transcribed from LIS-1. Expression of seven miRNAs (miR168, miR169, miR395, miR398, miR399, miR408, and lus-miR-N1) with up- or down-regulation under nutrient stress (on the basis of high-throughput sequencing data) was evaluated on extended sampling using qPCR. Reference gene search identified ETIF3H and ETIF3E genes as most suitable for this purpose. Down-regulation of novel potential lus-miR-N1 and up-regulation of conserved miR399 were revealed under the phosphate deficient conditions. In addition, the negative correlation of expression of lus-miR-N1 and its predicted target, ubiquitin-activating enzyme E1 gene, as well as, miR399 and its predicted target, ubiquitin-conjugating enzyme E2 gene, was observed. Thus, in our study, miRNAs expressed in flax plastic lines and genotrophs were identified and their expression and expression of their targets was evaluated using high-throughput sequencing and qPCR for the first time. These data provide new insights into nutrient stress response regulation in plastic flax cultivars.
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Nataliya V
NV
Melnikova
Laboratory of Structural and Functional Genomics, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Alexey A
AA
Dmitriev
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of SciencesMoscow, Russia; Department of Higher Plants, Lomonosov Moscow State UniversityMoscow, Russia.
Maxim S
MS
Belenikin
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Nadezhda V
NV
Koroban
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of SciencesMoscow, Russia; Department of Higher Plants, Lomonosov Moscow State UniversityMoscow, Russia.
Anna S
AS
Speranskaya
Department of Higher Plants, Lomonosov Moscow State University Moscow, Russia.
Anastasia A
AA
Krinitsina
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
George S
GS
Krasnov
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Valentina A
VA
Lakunina
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Anastasiya V
AV
Snezhkina
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Asiya F
AF
Sadritdinova
Laboratory of Genetics, All-Russian Research Institute for Flax Torzhok, Russia.
Natalya V
NV
Kishlyan
Laboratory of Genetics, All-Russian Research Institute for Flax Torzhok, Russia.
Tatiana A
TA
Rozhmina
Laboratory of Genetics of Microorganisms, Vavilov Institute of General Genetics, Russian Academy of Sciences Moscow, Russia.
Kseniya M
KM
Klimina
Laboratory of Molecular Karyology, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Alexandra V
AV
Amosova
Laboratory of Molecular Karyology, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Alexander V
AV
Zelenin
Laboratory of Molecular Karyology, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Olga V
OV
Muravenko
Laboratory of Molecular Karyology, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Nadezhda L
NL
Bolsheva
Laboratory of Post-Genomic Research, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences Moscow, Russia.
Anna V
AV
Kudryavtseva
Linum usitatissimum; gene expression; genotroph; miR399; microRNA; nutrient stress; qPCR; reference gene