Rice prolamins are categorized into three groups by molecular size (10, 13, or 16 kDa), while the 13 kDa prolamins are assigned to four subgroups (Pro13a-I, Pro13a-II, Pro13b-I, and Pro13b-II) based on cysteine residue content. Since lowering prolamin content in rice is essential to minimize indigestion and allergy risks, we generated four knockout lines using CRISPR-Cas9, which selectively reduced the expression of a specific subgroup of the 13 kDa prolamins. These four mutant rice lines also showed the compensatory expression of glutelins and non-targeted prolamins and were accompanied by low grain weight, altered starch content, and atypically-shaped starch granules and protein bodies. Transcriptome analysis identified 746 differentially expressed genes associated with 13 kDa prolamins during development. Correlation analysis revealed negative associations between genes in Pro13a-I and those in Pro13a-II and Pro13b-I/II subgroups. Furthermore, alterations in the transcription levels of 9 ER stress and 17 transcription factor genes were also observed in mutant rice lines with suppressed expression of 13 kDa prolamin. Our results provide profound insight into the functional role of 13 kDa rice prolamins in the regulatory mechanisms underlying rice seed development, suggesting their promising potential application to improve nutritional and immunological value.

Caleosin/peroxygenases (CLO/PXGs) play critical functional roles during plant development, oxylipin metabolism, and the response to abiotic/biotic stressors and environmental toxins. In Oryza sativa, peroxygenase-9 (OsPXG9) catabolizes intermediates in oxylipin biosynthesis produced by lipoxygenase-9 (9-LOX) and scavenges HOOH and CuOOH by transferring oxygen to hydroxy fatty acids (HFAs) but not to the free fatty acids. The resulting epoxide derivatives of HFAs are then enzymatically or non-enzymatically hydrolyzed into the corresponding trihydroxy derivatives. Results presented here demonstrate OsPXG9′s specificity for catabolizing products of the 9-LOX (and not for the 13-LOX) pathway of oxylipin biosynthesis. Overexpression of OsPXG9 reduces ROS (reactive oxygen species) abundance and reduces drought- and salt-stress-induced apoptotic cell death. The high expression level of OsPXG9 also stimulates drought- and salt-induced but not basal expression of antioxidant enzymes/pathways in plants, thereby increasing cellular resistance to drought. These results suggest that OsPXG9 decreases ROS abundance and is essential to increase resilience in rice plants exposed to exogenous or endogenous abiotic stress.

Caleosin/peroxygenases (CLO/PXGs) play critical functional roles during plant development, oxylipin metabolism and the response to abiotic/biotic stressors and environmental toxins. In Oryza sativa, peroxygenase 9 (OsPXG9) catabolizes intermediates in oxylipin biosynthesis produced by lipoxygenase 9 (9-LOX) and scavenges HOOH and CuOOH by transferring oxygen to hydroxy fatty acids (HFAs) but not to the free fatty acids. The resulting epoxide derivatives of HFAs are then enzymatically or non-enzymatically hydrolyzed into the corresponding trihydroxy derivatives. Results presented here demonstrate OsPXG9’s specificity for catabolizing products of the 9-LOX (and not for the 13-LOX) pathway of oxylipin biosynthesis. Overexpression of OsPXG9 reduces ROS abundance, reduces drought- and salt stress-induced apoptotic cell death and stimulates drought- and salt-induced but not basal expression of antioxidant enzymes/pathways in plants, thereby increasing cellular resistance to drought. These results suggest that OsPXG9 decreases ROS abundance and is essential to increase resilience in rice plants exposed to exogenous or endogenous abiotic stress.

Plant oxylipins are a diverse group of oxygenated fatty acid derivatives that function as important signaling molecules in plant responses to abiotic stress. Although jasmonates, particularly jasmonic acid (JA) and its derivatives, have been extensively studied, increasing evidence demonstrates that other oxylipin classes, including 12-oxophytodienoic acid (OPDA), green leaf volatiles (GLVs), reactive electrophilic oxylipins (RES), and peroxygenase (PXG)-derived oxylipins, also contribute to stress adaptation. This review summarizes current understanding of the molecular mechanisms underlying oxylipin signal perception, transduction, and regulation, with particular emphasis on interactions with other plant hormone pathways, reactive oxygen species (ROS), calcium (Ca²⁺), and mitogen-activated protein kinase (MAPK) signaling. We further examine transcriptional, post-transcriptional, and post-translational mechanisms that regulate oxylipin responses and discuss their integration across individual and combined abiotic stresses. Particular attention is given to experimentally established mechanisms while distinguishing emerging or unresolved signaling processes. Understanding these interconnected signaling mechanisms will be important for developing strategies to improve crop resilience under global climate change.

Plant oxylipins are a diverse group of oxygenated fatty acid derivatives that function as important signaling molecules in plant responses to abiotic stress. Although jasmonates, particularly jasmonic acid (JA) and its derivatives, have been extensively studied, increasing evidence demonstrates that other oxylipin classes, including 12-oxophytodienoic acid (OPDA), green leaf volatiles (GLVs), reactive electrophilic oxylipins (RES), and peroxygenase (PXG)-derived oxylipins, also contribute to stress adaptation. This review summarizes current understanding of the molecular mechanisms underlying oxylipin signal perception, transduction, and regulation, with particular emphasis on interactions with other plant hormone pathways, reactive oxygen species (ROS), Calcium (Ca²⁺), and mitogen-activated protein kinase (MAPK) signaling. We further examine transcriptional, post-transcriptional, and post-translational mechanisms that regulate oxylipin responses and discuss their integration across individual and combined abiotic stresses. Particular attention is given to experimentally established mechanisms while distinguishing emerging or unresolved signaling processes. Understanding these interconnected signaling mechanisms will be important for developing strategies to improve crop resilience under global climate change.

Rice prolamins are categorized into three groups by molecular size (10, 13, or 16 kDa), while the 13 kDa prolamins are assigned to four subgroups (Pro13a-I, Pro13a-II, Pro13b-I, and Pro13b-II) based on cysteine residue content. Here, we generated four knockout strains of rice using CRISPR-Cas9, each of which demonstrated selectively reduced expression of specific subgroups of the 13 kDa prolamins. These four mutant rice lines also demonstrated compensatory expression of glutelins and non-targeted prolamins, which resulted in low grain weight, altered starch content and atypically-shaped starch granules and protein bodies. Transcriptome analysis identified 746 transcripts that were differentially expressed in the mutant rice lines during development. These transcripts played roles primarily in RNA processing, protein synthesis, stress response, and transport. After selective suppression/knockout of genes in the Pro13a-I subgroup, compensatory upregulation of genes in the Pro13a-II and Pro13b-I/II subgroups was observed. Compensatory expression of 9 ER stress and 17 transcription factor genes was also observed in mutant rice in which expression of 13 kDa prolamin genes was suppressed. Our results provide valuable insight into the regulatory mechanisms underlying rice seed development and the specific roles played by 13 kDa rice prolamins and their target genes.