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Abstract Tolerance of chilling dictates the geographical distribution, establishment, and productivity of C₄crops. Chilling reduces enzyme rate, limiting the sink for the absorbed light energy leading to the need for quick energy dissipation via non-photochemical quenching (NPQ). Here, we characterize NPQ upon chilling in threeMiscanthusaccessions representing diverse chilling tolerance in C₄grasses. High chilling tolerant accessions accumulate substantial amounts of zeaxanthin during chilling nights in both field and growth chamber settings. Chilling-induced zeaxanthin accumulation in the dark enhances rate of NPQ induction by 66% in the following morning. Based on our data, the emerging ways for the unique regulation of NPQ include post-translational regulation of violaxanthin de-epoxidase (VDE), VDE cofactor accessibility, and absence of transcriptional upregulation of zeaxanthin conversion back to violaxanthin. In the future, more studies will be required to obtain further evidence for these ways contributions to the chilling-dark regulation of NPQ. Engineering dark accumulation of zeaxanthin will help improve crop chilling tolerance and promote sustainable production by allowing early spring planting to maximize the use of early-season soil moisture. Driving the engineered trait by chilling inducible promoter would ensure the minimization of a trade-off between photoprotection and photosynthesis efficiency. 

Abstract Water supply limitations will likely impose increasing restrictions on future crop production, underlining a need for crops that use less water per mass of yield. Water use efficiency (WUE) therefore becomes a key consideration in developing resilient and productive crops. In this study, we hypothesized that it is possible to improve WUE under drought conditions via modulation of chloroplast signals for stomatal opening by up-regulation of non-photochemical quenching (NPQ). Nicotiana tabacum plants with strong overexpression of the PsbS gene encoding PHOTOSYSTEM II SUBUNIT S, a key protein in NPQ, were grown under differing levels of drought. The PsbS-overexpressing lines lost 11% less water per unit CO2 fixed under drought and this did not have a significant effect on plant size. Depending on growth conditions, the PsbS-overexpressing lines consumed from 4–30% less water at the whole-plant level than the corresponding wild type. Leaf water and chlorophyll contents showed a positive relation with the level of NPQ. This study therefore provides proof of concept that up-regulation of NPQ can increase WUE, and as such is an important step towards future engineering of crops with improved performance under drought.