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. 2021 Dec 6;12:758785. doi: 10.3389/fpls.2021.758785

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Copyright © 2021 Jia, Liu, Geng and Jiang.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Correlation between ACS2/6 activation and stomatal density and rate of stomatal clustering. (A) Images of stomata distributed on strips of leaf abaxial epidermis under differential interference contrast (DIC) microscopy. Stomata and stomatal clusters are colored in blue for easier identification, and stomatal clusters are indicated by black arrows. The number of stomata is given in parentheses in each image. Sixth rosette were collected under drought or normal watering conditions from 28-day-old seedlings of the WT control; mutants acs2-1, acs6-1, and acs2-1acs6-1, overexpression lines ACS2-OE(#1) and ACS6-OE(#1), and complementation lines ACS2/acs2-1(#1) and ACS6/acs6-1(#1). Experiments were performed three times with similar results. The white scale bar represents 100 μm. (B,C) Statistical analysis of stomatal density (B) and percentage of clustered stomata (C) under normal and drought condition. Number of stomata and stomatal clusters on sixth leaves of 25 seedlings were counted. Values are means ± SD (n = 25 repeats). Letters indicate significant differences (P < 0.05, two-way ANOVA). (D,E) After treated immature leaves by ACC (0–20 μM) for 6 days, stomatal density (D) and percentage of clustered stomata (E) were analyzed, respectively. A total of 25 leaves from 25 seedlings were used to analyze the numbers of stomata and percentage of clustered stomata. Values are means ± SD. (Student’s t-test, ^**P < 0.01).