Метаболизм каллозы в волокнах льна при гравиответе: анализ экспрессии генов

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Abstract

Двигательные реакции растений, относящиеся к тропизмам, обычно осуществляются с использованием механизмов роста растяжением. Однако в данном исследовании изучался гравитропизм на уровне клеток (первичные флоэмные волокна), закончивших свой рост и формирующих утолщенную третичную клеточную стенку. Проведены инвентаризация и анализ экспрессии генов ферментов, ответственных за метаболизм каллозы, на разных стадиях развития флоэмных волокон льна (Linum usitatissimum L.) и при гравиответе. Выявлены гены предполагаемых β-1,3-глюкансинтаз (GSLs) и β-1,3-глюканаз (BGs), имеющие дифференциальный характер экспрессии в исследуемых клетках, среди которых отмечены гены с максимальным уровнем экспрессии на определенной стадии развития. В основном при гравитропизме экспрессия генов β-1,3-глюкансинтаз была понижена, тогда как для генов β-1,3-глюканаз были характерны различные профили экспрессии, среди которых выявлены гены с повышенным уровнем экспрессии только при гравиответе (LusBG1 и LusBG3). Полученные данные позволили предположить наличие активного метаболизма каллозы в клеточной стенке исследуемых волокон на разных стадиях развития и доминирование деградации каллозы в ходе гравиответа. Результаты работы закладывают основу для дальнейших исследований функции каллозы в развитии волокон и реализации двигательной реакции растений.

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Н. Н. Ибрагимова

Казанский институт биохимии и биофизики – обособленное структурное подразделение Федерального государственного бюджетного учреждения науки Федерального исследовательского центра “Казанский научный центр Российской академии наук”

Author for correspondence.
Email: nibra@yandex.ru
Russian Federation, Казань

Н. Е. Мокшина

Казанский институт биохимии и биофизики – обособленное структурное подразделение Федерального государственного бюджетного учреждения науки Федерального исследовательского центра “Казанский научный центр Российской академии наук”

Email: nibra@yandex.ru
Russian Federation, Казань

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Supplementary files

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2. Fig. 1. Scheme of sampling (isolated flax fibers) for gene expression assessment. Expression data are uploaded to FIBexDB [18]; iFIBa – intrusively growing fibers, tFIBa – early stage of TCR (tertiary cell wall) formation, tFIBb – late stage of TCR formation; 8, 24, 96 h – time of sample fixation after stem tilting; PUL – pulling side, OPP – opposite to PUL-side. The right side of the table shows the project numbers in NCBI, which published raw data of transcriptome analyses. Normalized data for all transcriptome experiments can be found in FIBexDB (https://ssl.cres-t.org/fibex/flax/).

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3. Fig. 2. List of β-1,3-glucan synthase genes (PF02364, PF04652) and their expression in flax fibers at different stages of development and during gravistimulation (in TGR) divided into clusters 1–4. Genes, the expression of which was significantly reduced during gravistimulation in flax stem fibers, are shown in blue; genes, the expression of which significantly increased in fibers during gravistimulation compared to the expression in fibers of control plants (tFIBb), are shown in red. * (c) – Differences from control (tFIBb) and (b) – differences with tFIBa are significant at q < 0.05. Genes, the expression of which was not considered, are highlighted in gray. iFIBa – intrusively growing fibers, tFIBa – early stage of TCR (tertiary cell wall) formation. tFIBb – late stage of TCS formation, which served as a control (CTR) in the experiment with gravistimulation. 8, 24, 96 h – time of sample fixation after stem tilting; PUL – pulling side, OPP – opposite to PUL-side. Lus – sequences of flax Linum usitatissimum, At – sequences of Arabidopsis thaliana.

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4. Fig. 3. Expression of β-1,3-glucan synthase genes (PF02364) in flax stem fibers from clusters 1–4 with a maximum at different stages of development and a decrease in expression during gravioresponse (a), as well as sharply decreasing expression in fibers with a mature cell wall (tFIBb) compared to iFIBa and tFIBa, but activated during gravioresponse (b). * – Differences from the control are significant at q < 0.05. iFIBa – intrusively growing fibers, tFIBa – early stage of TCR (tertiary cell wall) formation. tFIBb – late stage of TCR formation. 8, 24, 96 h – time of sample fixation after stem tilting; PUL – pulling side, OPP – side opposite to PUL. The black dotted line indicates the expression level in the control plants without gravistimulation.

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5. Fig. 4. Genes of β-1,3-glucan synthase (PF02364), homologous to ATGSL7 (a) and ATGSL2 (b), having a high level of expression in flax fibers at different stages of development, but decreasing expression under gravistimulation. * – Differences with the control are significant at q < 0.05. iFIBa – intrusively growing fibers, tFIBa – early stage of TCR (tertiary cell wall) formation. tFIBb – late stage of TCR formation. 8, 24, 96 h – time of sample fixation after stem bending; PUL – pulling side, OPP – side opposite to PUL. TGR – total gene reads. The black dotted line indicates the expression level in the control plants without gravistimulation.

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6. Fig. 5. List of genes encoding glycoside hydrolases of family 17 (PF00332) and their expression in flax stem fibers (in TGR). To the left of the heat map are flax genes (and their closest homologs in Arabidopsis thaliana) exhibiting more than a 3-fold change in expression during the gravioresponse (in at least one sample); genes with the highest expression levels only during the gravioresponse are highlighted. * (c) – Differences from the control (tFIBb) and b – differences with tFIBa are significant at q < 0.05. iFIBa – intrusively growing fibers, tFIBa – early stage of TCR (tertiary cell wall) formation. tFIBb – late stage of TCR formation, which served as a control (CTR) in the experiment with gravistimulation. 8, 24, 96 h – time of sample fixation after stem tilting; PUL – pulling side, OPP – opposite PUL side. Lus – sequences of flax Linum usitatissimum, Аt – sequences of Arabidopsis thaliana. BG – β-1,3-glucanase. Samples with maximum gene expression are marked on the right. g – predicted GPI anchor (coefficient 0.173–0.503).

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7. Fig. 6. Genes of β-1,3-glucanases, the expression of which increased in PUL fibers under gravistimulation compared to OPP. The domain organization of the corresponding protein sequences is shown at the top: SP – signal peptide, GH17 – Glycosyde hydrolase family 17. * – The difference with the control is significant at q < 0.05; t-test – the differences between the samples are significant at P < 0.05. iFIBa – intrusively growing fibers, tFIBa – early stage of TCR (tertiary cell wall) formation, tFIBb – late stage of TCR formation. 8, 24, 96 h – time of sample fixation after stem tilting; PUL – pulling side, OPP – side opposite to PUL. TGR – total gene reads. The black dotted line indicates the expression level in the control plants without gravistimulation.

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