Suppression of Hydroxycinnamate Network Formation in Cell Walls of Rice Shoots Grown under Microgravity Conditions in Space

<div><p>Network structures created by hydroxycinnamate cross-links within the cell wall architecture of gramineous plants make the cell wall resistant to the gravitational force of the earth. In this study, the effects of microgravity on the formation of cell wall-bound hydroxycinnamates were examined using etiolated rice shoots simultaneously grown under artificial 1 <i>g</i> and microgravity conditions in the Cell Biology Experiment Facility on the International Space Station. Measurement of the mechanical properties of cell walls showed that shoot cell walls became stiff during the growth period and that microgravity suppressed this stiffening. Amounts of cell wall polysaccharides, cell wall-bound phenolic acids, and lignin in rice shoots increased as the shoot grew. Microgravity did not influence changes in the amounts of cell wall polysaccharides or phenolic acid monomers such as ferulic acid (FA) and <i>p</i>-coumaric acid, but it suppressed increases in diferulic acid (DFA) isomers and lignin. Activities of the enzymes phenylalanine ammonia-lyase (PAL) and cell wall-bound peroxidase (CW-PRX) in shoots also increased as the shoot grew. PAL activity in microgravity-grown shoots was almost comparable to that in artificial 1 <i>g</i>-grown shoots, while CW-PRX activity increased less in microgravity-grown shoots than in artificial 1 <i>g</i>-grown shoots. Furthermore, the increases in expression levels of some class III peroxidase genes were reduced under microgravity conditions. These results suggest that a microgravity environment modifies the expression levels of certain class III peroxidase genes in rice shoots, that the resultant reduction of CW-PRX activity may be involved in suppressing DFA formation and lignin polymerization, and that this suppression may cause a decrease in cross-linkages within the cell wall architecture. The reduction in intra-network structures may contribute to keeping the cell wall loose under microgravity conditions.</p></div

Activity of PAL in rice shoots.

<p>Activity is expressed as the amounts of <i>t</i>-cinnamic acid (CA) produced during the assay. Data are means ± SE from three independent samples. Statistically significant difference between on-orbit (artificial) 1 <i>g</i> and microgravity conditions was not detected.</p><p>Activity of PAL in rice shoots.</p

Rice seedlings grown for 4 days under artificial (on-orbit) 1 <i>g</i> and microgravity conditions in orbit.

<p>Lower panels show magnified images of 4 d seedlings whose roots were excised.</p

Amounts of FA and <i>p</i>-coumaric acid (<i>p</i>-CA) in the cell walls of rice shoots.

<p>FA and <i>p</i>-coumaric acid were analyzed by HPLC on a reversed-phase column. The figure shows means ± SE from three independent samples. As to FA and <i>p</i>-coumaric acid contents, statistically significant difference between artificial (on-orbit) 1 <i>g</i> and microgravity conditions was not detected.</p

Amounts of DFA isomers in the cell walls of rice shoots.

<p>DFA isomers were analyzed by HPLC on a reversed-phase column. The figure shows means ± SE from three independent samples. *Mean values significantly different between artificial (on-orbit) 1 <i>g</i> and microgravity conditions (Student's <i>t</i>-test, P < 0.05).</p

Activity of CW-PRX in rice shoots measured using two substrates, guaiacol and FA.

<p>Activity is expressed as the increase in the absorbance at 470 nm for guaiacol and as the decrease in the absorbance at 310 nm for FA. The figure shows means ± SE from three independent samples. *Mean values significantly different between artificial (on-orbit) 1 <i>g</i> and microgravity conditions (Student's <i>t</i>-test, P < 0.05).</p

Root-tip-mediated inhibition of hydrotropism is accompanied with the suppression of asymmetric expression of auxin-inducible genes in response to moisture gradients in cucumber roots

<div><p>In cucumber seedlings, gravitropism interferes with hydrotropism, which results in the nearly complete inhibition of hydrotropism under stationary conditions. However, hydrotropic responses are induced when the gravitropic response in the root is nullified by clinorotation. Columella cells in the root cap sense gravity, which induces the gravitropic response. In this study, we found that removing the root tip induced hydrotropism in cucumber roots under stationary conditions. The application of auxin transport inhibitors to cucumber seedlings under stationary conditions suppressed the hydrotropic response induced by the removal of the root tip. To investigate the expression of genes related to hydrotropism in de-tipped cucumber roots, we conducted transcriptome analysis of gene expression by RNA-Seq using seedlings exhibiting hydrotropic and gravitropic responses. Of the 21 and 45 genes asymmetrically expressed during hydrotropic and gravitropic responses, respectively, five genes were identical. Gene ontology (GO) analysis indicated that the category auxin-inducible genes was significantly enriched among genes that were more highly expressed in the concave side of the root than the convex side during hydrotropic or gravitropic responses. Reverse transcription followed by quantitative polymerase chain reaction (RT-qPCR) analysis revealed that root hydrotropism induced under stationary conditions (by removing the root tip) was accompanied by the asymmetric expression of several auxin-inducible genes. However, intact roots did not exhibit the asymmetric expression patterns of auxin-inducible genes under stationary conditions, even in the presence of a moisture gradient. These results suggest that the root tip inhibits hydrotropism by suppressing the induction of asymmetric auxin distribution. Auxin transport and distribution not mediated by the root tip might play a role in hydrotropism in cucumber roots.</p></div