BOTRYOID POLLEN 1 regulates ROS-triggered PCD and pollen wall development by controlling UDP-sugar homeostasis in rice.

Uridine diphosphate (UDP)-sugars are important metabolites involved in the biosynthesis of polysaccharides and may be important signaling molecules. UDP-glucose 4-epimerase (UGE) catalyzes the interconversion between UDP-Glc and UDP-Gal, whose biological function in rice (Oryza sativa) fertility is poorly understood. Here, we identify and characterize the botryoid pollen 1 (bp1) mutant and show that BP1 encodes a UGE that regulates UDP-sugar homeostasis, thereby controlling the development of rice anthers. The loss of BP1 function led to massive accumulation of UDP-Glc and imbalance of other UDP-sugars. We determined that the higher levels of UDP-Glc and its derivatives in bp1 may induce the expression of NADPH oxidase genes, resulting in a premature accumulation of reactive oxygen species (ROS), thereby advancing programmed cell death (PCD) of anther walls but delaying the end of tapetal degradation. The accumulation of UDP-Glc as metabolites resulted in an abnormal degradation of callose, producing an adhesive microspore. Furthermore, the UDP-sugar metabolism pathway is not only involved in the formation of intine but also in the formation of the initial framework for extine. Our results reveal how UDP-sugars regulate anther development and provide new clues for cellular ROS accumulation and PCD triggered by UDP-Glc as a signaling molecule.

The OsMYC2-JA feedback loop regulates diurnal flower-opening time via cell wall loosening in rice.

Diurnal flower-opening time (DFOT), the time of spikelet opening during the day, is an important trait for hybrid rice (Oryza sativa L.) seed production. Hybrids between indica and japonica rice varieties have strong heterosis, but the parental lines usually have different, nonoverlapping DFOTs. This reduces the success of hybrid seed production in crosses between indica and japonica subspecies, thus hindering the utilization of indica and japonica inter-subspecies heterosis. However, little is known about the molecular mechanisms regulating DFOT in rice. Here, we obtained japonica rice lines with a DFOT 1.5 h earlier than the wild type by overexpressing OsMYC2, a gene encoding a key transcription factor in the jasmonate (JA) signaling pathway. OsMYC2 is activated by JA signaling and directly regulates the transcription of genes related to JA biosynthesis and cell wall metabolism. Overexpressing OsMYC2 led to significantly increased JA contents and decreased cellulose and hemicellulose contents in lodicule cells, as well as the softening of lodicule cell walls. This may facilitate the swelling of lodicules, resulting in early diurnal flower-opening. These results suggest that the OsMYC2-JA feedback loop regulates DFOT in rice via cell wall remodeling. These findings shed light on the understanding of regulatory mechanism of the DFOT of plants, which should promote the development of indica and japonica varieties suitable for hybrid rice breeding.

SET DOMAIN GROUP 711-mediated H3K27me3 methylation of cytokinin metabolism genes regulates organ size in rice.

Organ size shapes plant architecture during rice (Oryza sativa) growth and development, affecting key factors influencing yield, such as plant height, leaf size, and seed size. Here, we report that the rice Enhancer of Zeste [E(z)] homolog SET DOMAIN GROUP 711 (OsSDG711) regulates organ size in rice. Knockout of OsSDG711 produced shorter plants with smaller leaves, thinner stems, and smaller grains. We demonstrate that OsSDG711 affects organ size by reducing cell length and width and increasing cell number in leaves, stems, and grains. The result of chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) using an antitrimethylation of histone H3 lysine 27 (H3K27me3) antibody showed that the levels of H3K27me3 associated with cytokinin oxidase/dehydrogenase genes (OsCKXs) were lower in the OsSDG711 knockout line Ossdg711. ChIP-qPCR assays indicated that OsSDG711 regulates the expression of OsCKX genes through H3K27me3 histone modification. Importantly, we show that OsSDG711 directly binds to the promoters of these OsCKX genes. Furthermore, we measured significantly lower cytokinin contents in Ossdg711 plants than in wild-type plants. Overall, our results reveal an epigenetic mechanism based on OsSDG711-mediated modulation of H3K27me3 levels to regulate the expression of genes involved in the cytokinin metabolism pathway and control organ development in rice. OsSDG711 may be an untapped epigenetic resource for ideal plant type improvement.

Controlling diurnal flower-opening time by manipulating the jasmonate pathway accelerates development of indica-japonica hybrid rice breeding.

Inter-subspecific indica-japonica hybrid rice (Oryza sativa) has the potential for increased yields over traditional indica intra-subspecies hybrid rice, but limited yield of F1 hybrid seed production (FHSP) hinders the development of indica-japonica hybrid rice breeding. Diurnal flower-opening time (DFOT) divergence between indica and japonica rice has been a major contributing factor to this issue, but few DFOT genes have been cloned. Here, we found that manipulating the expression of jasmonate (JA) pathway genes can effectively modulate DFOT to improve the yield of FHSP in rice. Treating japonica cultivar Zhonghua 11 (ZH11) with methyl jasmonate (MeJA) substantially advanced DFOT. Furthermore, overexpressing the JA biosynthesis gene OPDA REDUCTASE 7 (OsOPR7) and knocking out the JA inactivation gene CHILLING TOLERANCE 1 (OsHAN1) in ZH11 advanced DFOT by 1- and 2-h respectively; and knockout of the JA signal suppressor genes JASMONATE ZIM-DOMAIN PROTEIN 7 (OsJAZ7) and OsJAZ9 resulted in 50-min and 1.5-h earlier DFOT respectively. The yields of FHSP using japonica male-sterile lines GAZS with manipulated JA pathway genes were significantly higher than that of GAZS wildtype. Transcriptome analysis, cytological observations, measurements of elastic modulus and determination of cell wall components indicated that the JA pathway could affect the loosening of the lodicule cell walls by regulating their composition through controlling sugar metabolism, which in turn influences DFOT. This research has vital implications for breeding japonica rice cultivars with early DFOT to facilitate indica-japonica hybrid rice breeding.

An E3 ubiquitin ligase CSIT2 controls critical sterility-inducing temperature of thermo-sensitive genic male sterile rice.

Thermo-sensitive genic male sterile (TGMS) lines are the core of two-line hybrid rice (Oryza sativa). However, elevated or unstable critical sterility-inducing temperatures (CSITs) of TGMS lines are bottlenecks that restrict the development of two-line hybrid rice. However, the genes and molecular mechanisms controlling CSIT remain unknown. Here, we report the CRITICAL STERILITY-INDUCING TEMPERATURE 2 (CSIT2) that encodes a really interesting new gene (RING) type E3 ligase, controlling the CSIT of thermo-sensitive male sterility 5 (tms5)-based TGMS lines through ribosome-associated protein quality control (RQC). CSIT2 binds to the large and small ribosomal subunits and ubiquitinates 80S ribosomes for dissociation, and may also ubiquitinate misfolded proteins for degradation. Mutation of CSIT2 inhibits the possible damage to ubiquitin system and protein translation, which allows more proteins such as catalases to accumulate for anther development and inhibits abnormal accumulation of reactive oxygen species (ROS) and premature programmed cell death (PCD) in anthers, partly rescuing male sterility and raised the CSIT of tms5-based TGMS lines. These findings reveal a mechanism controlling CSIT and provide a strategy for solving the elevated or unstable CSITs of tms5-based TGMS lines in two-line hybrid rice.

Environment-sensitive genic male sterility in rice and other plants.

Environment-sensitive genic male sterility is a type of male sterility that is affected by both genetic and environmental factors. Environment-sensitive genic male sterile lines are not only used in two-line hybrid breeding but are also good materials for studying plant-environment interactions. In this study we review the research progress on environment-sensitive genic male sterility in rice from the perspectives of epigenetic, transcriptional, posttranscriptional, posttranslational and metabolic mechanisms as well as signal transduction processes. While significant progress has been made in the genetics, gene cloning and understanding of the molecular mechanisms of environment-sensitive genic male sterility in recent years, the relevant regulatory network is still poorly understood in rice. We therefore also review studies of environment-sensitive genic male sterility in Arabidopsis and other crops, hoping to promote research in this field in rice. Finally, we analyse the challenges posed by environment-sensitive genic male sterility and provide corresponding suggestions. This review will contribute towards an understanding the molecular genetics of environment-sensitive genic male sterility and its application in two-line hybrid breeding in rice and other species.

Advances on the Study of Diurnal Flower-Opening Times of Rice.

The principal goal of rice (Oryza sativa L.) breeding is to increase the yield. In the past, hybrid rice was mainly indica intra-subspecies hybrids, but its yield has been difficult to improve. The hybridization between the indica and japonica subspecies has stronger heterosis; the utilization of inter-subspecies heterosis is important for long-term improvement of rice yields. However, the different diurnal flower-opening times (DFOTs) between the indica and japonica subspecies seriously reduce the efficiency of cross-pollination and yield and increase the cost of indica-japonica hybrid rice seeds, which has become one of the main constraints for the development of indica-japonica hybrid rice breeding. The DFOT of plants is adapted to their growing environment and is also closely related to species stability and evolution. Herein, we review the structure and physiological basis of rice flower opening, the factors that affect DFOT, and the progress of cloning and characterization of DFOT genes in rice. We also analyze the problems in the study of DFOT and provide corresponding suggestions.

A cytosolic pentatricopeptide repeat protein is essential for tapetal plastid development by regulating OsGLK1 transcript levels in rice.

Most plant pentatricopeptide repeat (PPR) proteins localize to and function inside plastids and mitochondria. However, the function of PPRs that only localize to the cytoplasm remains unknown. Here, we demonstrated that the rice (Oryza sativa) PPR protein CYTOPLASM-LOCALIZED PPR1 (OsCPPR1) contributes to pollen development and localizes to the cytoplasm. Knocking down OsCPPR1 led to abnormal plastid development in tapetal cells, prolonged tapetal programmed cell death (PCD) and tapetum degradation, and significantly reduced pollen fertility. Transcriptome analysis revealed that the transcript level of OsGOLDEN-LIKE1 (OsGLK1), which encodes a transcription factor that regulates plastid development and maintenance, was significantly higher in the OsCPPR1 knockdown plants compared to wild-type plants. We further determined that OsCPPR1 downregulates OsGLK1 transcription by directly binding to the single-stranded regions of OsGLK1 mRNAs. Overexpression of OsGLK1 resulted in abnormal tapetum and plastid development, similar to that seen in OsCPPR1 knockdown plants, and suppression of OsGLK1 partially restored pollen fertility in the OsCPPR1 knockdown plants. We therefore conclude that OsCPPR1 suppresses OsGLK1 in the regulation of plastid development and PCD in the tapetum. Our work revealed novel functions for a cytosolic PPR, demonstrating the diverse roles of PPRs in plants and identifying a new regulatory mechanism for regulating pollen development in rice.

OsAGO2 controls ROS production and the initiation of tapetal PCD by epigenetically regulating OsHXK1 expression in rice anthers.

Proteins of the ARGONAUTE (AGO) family function in the epigenetic regulation of gene expression. Although the rice (Oryza sativa) genome encodes 19 predicted AGO proteins, few of their functions have thus far been characterized. Here, we show that the AGO protein OsAGO2 regulates anther development in rice. OsAGO2 was highly expressed in anthers. Knockdown of OsAGO2 led to the overaccumulation of reactive oxygen species (ROS) and abnormal anther development, causing premature initiation of tapetal programmed cell death (PCD) and pollen abortion. The expression level of Hexokinase 1 (OsHXK1) increased significantly, and the methylation levels of its promoter decreased, in plants with knocked-down OsAGO2 expression. Overexpression of OsHXK1 also resulted in the overaccumulation of ROS, premature initiation of PCD, and pollen abortion. Moreover, knockdown of OsHXK1 restored pollen fertility in OsAGO2 knockdown plants. Chromatin immunoprecipitation assays demonstrated that OsAGO2 binds directly to the OsHXK1 promoter region, suggesting that OsHXK1 is a target gene of OsAGO2. These results indicate that OsHXK1 controls the appropriate production of ROS and the proper timing of tapetal PCD and is directly regulated by OsAGO2 through epigenetic regulation.

Heterologous Expression of Jatropha curcas Fatty Acyl-ACP Thioesterase A (JcFATA) and B (JcFATB) Affects Fatty Acid Accumulation and Promotes Plant Growth and Development in Arabidopsis.

Plant fatty acyl-acyl carrier protein (ACP) thioesterases terminate the process of de novo fatty acid biosynthesis in plastids by hydrolyzing the acyl-ACP intermediates, and determine the chain length and levels of free fatty acids. They are of interest due to their roles in fatty acid synthesis and their potential to modify plant seed oils through biotechnology. Fatty acyl-ACP thioesterases (FAT) are divided into two families, i.e., FATA and FATB, according to their amino acid sequence and substrate specificity. The high oil content in Jatropha curcas L. seed has attracted global attention due to its potential for the production of biodiesel. However, the detailed effects of JcFATA and JcFATB on fatty acid biosynthesis and plant growth and development are still unclear. In this study, we found that JcFATB transcripts were detected in all tissues and organs examined, with especially high accumulation in the roots, leaves, flowers, and some stages of developing seeds, and JcFATA showed a very similar expression pattern. Subcellular localization of the JcFATA-GFP and JcFATB-GFP fusion protein in Arabidopsis leaf protoplasts showed that both JcFATA and JcFATB localized in chloroplasts. Heterologous expression of JcFATA and JcFATB in Arabidopsis thaliana individually generated transgenic plants with longer roots, stems and siliques, larger rosette leaves, and bigger seeds compared with those of the wild type, indicating the overall promotion effects of JcFATA and JcFATB on plant growth and development while JcFATB had a larger impact. Compositional analysis of seed oil revealed that all fatty acids except 22:0 were significantly increased in the mature seeds of JcFATA-transgenic Arabidopsis lines, especially unsaturated fatty acids, such as the predominant fatty acids of seed oil, 18:1, 18:2, and 18:3. In the mature seeds of the JcFATB-transgenic Arabidopsis lines, most fatty acids were increased compared with those in wild type too, especially saturated fatty acids, such as 16:0, 18:0, 20:0, and 22:0. Our results demonstrated the promotion effect of JcFATA and JcFATB on plant growth and development, and their possible utilization to modify the seed oil composition and content in higher plants.

Hexokinase gene OsHXK1 positively regulates leaf senescence in rice.

BACKGROUND: Leaf senescence is a highly complex and meticulous regulatory process, and the disruption of any factor involved in leaf senescence might lead to premature or delayed leaf senescence and thus result in reduced or increased crop yields. Despite sincere efforts by scientists, there remain many unsolved problems related to the regulatory factors and molecular mechanisms of leaf senescence. RESULTS: This study successfully revealed that OsHXK1 was highly expressed in senescent leaves of rice. The upregulation of OsHXK1 led to premature senescence of rice leaves, a decreased level of chlorophyll, and damage to the chloroplast structure. The overexpression of OsHXK1 resulted in increases in glucose and ROS levels and produced programmed cell death (PCD) signals earlier at the booting stage. Further analysis showed that expression level of the respiratory burst oxidase homolog (RBOH) genes and OsGLO1 were increased in OsHXK1-overexpressing plants at the booting stage. CONCLUSIONS: Overall, the outcomes of this study suggested that OsHXK1 could act as a positive regulator of rice leaf senescence by mediating glucose accumulation and inducing an increase in ROS.

Initiation and Execution of Programmed Cell Death and Regulation of Reactive Oxygen Species in Plants.

Programmed cell death (PCD) plays crucial roles in plant development and defence response. Reactive oxygen species (ROS) are produced during normal plant growth, and high ROS concentrations can change the antioxidant status of cells, leading to spontaneous cell death. In addition, ROS function as signalling molecules to improve plant stress tolerance, and they induce PCD under different conditions. This review describes the mechanisms underlying plant PCD, the key functions of mitochondria and chloroplasts in PCD, and the relationship between mitochondria and chloroplasts during PCD. Additionally, the review discusses the factors that regulate PCD. Most importantly, in this review, we summarise the sites of production of ROS and discuss the roles of ROS that not only trigger multiple signalling pathways leading to PCD but also participate in the execution of PCD, highlighting the importance of ROS in PCD.

HMS1 interacts with HMS1I to regulate very-long-chain fatty acid biosynthesis and the humidity-sensitive genic male sterility in rice (Oryza sativa).

Environment-sensitive genic male sterility (EGMS) lines are used widely in two-line hybrid breeding in rice (Oryza sativa). At present, photoperiod-sensitive genic male sterility (PGMS) lines and thermo-sensitive genic male sterility (TGMS) lines are predominantly used in two-line hybrid rice, with humidity-sensitive genic male sterility (HGMS) lines rarely being reported. Here, it is shown that HUMIDITY-SENSITIVE GENIC MALE STERILITY 1 (HMS1), encoding a ß-ketoacyl-CoA synthase, plays key roles in the biosynthesis of very-long-chain fatty acids (VLCFAs) and HGMS in rice. The hms1 mutant displayed decreased seed setting under low humidity, but normal seed setting under high humidity. HMS1 catalyzed the biosynthesis of the C26 and C28 VLCFAs, contributing to the formation of bacula and tryphine in the pollen wall, which protect the pollen from dehydration. Under low-humidity conditions, hms1 pollen showed poor adhesion and reduced germination on the stigmas, which could be rescued by increasing humidity. HMS1-INTERACTING PROTEIN (HMS1I) interacted with HMS1 to coregulate HGMS. Furthermore, both japonica and indica rice varieties with defective HMS1 exhibited HGMS, suggesting that hms1 potentially could be used in hybrid breeding. The results herein reveal the novel mechanism of VLCFA-mediated pollen wall formation, which protects pollen from low-humidity stress in rice, and has a potential use in hybrid crop breeding.

Molecular Control and Application of Male Fertility for Two-Line Hybrid Rice Breeding.

The significance of the climate change may involve enhancement of plant growth as well as utilization of the environmental alterations in male fertility (MF) regulation via male sterility (MS) systems. We described that MS systems provide a fundamental platform for improvement in agriculture production and have been explicated for creating bulk germplasm of the two-line hybrids (EGMS) in rice as compared to the three-line, to gain production sustainability and exploit its immense potential. Environmental alterations such as photoperiod and/or temperature and humidity regulate MS in EGMS lines via genetic and epigenetic changes, regulation of the noncoding RNAs, and RNA-metabolism including the transcriptional factors (TFs) implication. Herein, this article enlightens a deep understanding of the molecular control of MF in EGMS lines and exploring the regulatory driving forces that function efficiently during plant adaption under a changing environment. We highlighted a possible solution in obtaining more stable hybrids through apomixis (single-line system) for seed production.

An efficient rice mutagenesis system based on suspension-cultured cells.

Plant mutants are important bio-resources for crop breeding and gene functional studies. Conventional methods for generating mutant libraries by mutagenesis of seeds with physical or chemical agents are of low efficiency. Here, we developed a highly-efficient ethyl methanesulfonate (EMS) mutagenesis system based on suspension-cultured cells, with rice (Oryza sativa L.) as an example. We show that treatment of suspension-cultured tiny cell clusters with 0.4% EMS for 18-22 h followed by differentiation and regeneration produced as high as 29.4% independent mutant lines with visible phenotypic variations, including a number of important agronomic traits such as grain size, panicle size, grain or panicle shape, tiller number and angle, heading date, male sterility, and disease sensitivity. No mosaic mutant was observed in the mutant lines tested. In this mutant library, we obtained a mutant with an abnormally elongated uppermost internode. Sequencing and functional analysis revealed that this is a new allelic mutant of eui (elongated uppermost internode) caused by two point mutations in the first exon of the EUI gene, representing a successful example of this mutagenesis system.

The E3 ubiquitin ligase CSIT1 regulates critical sterility-inducing temperature by ribosome-associated quality control to safeguard two-line hybrid breeding in rice.

Two-line hybrid breeding can fully utilize heterosis in crops. In thermo-sensitive genic male sterile (TGMS) lines, low critical sterility-inducing temperature (CSIT) is vital to safeguard the production of two-line hybrid seeds in rice (Oryza sativa), but the molecular mechanism determining CSIT is unclear. Here, we report the cloning of CSIT1, which encodes an E3 ubiquitin ligase, and show that CSIT1 modulates the CSIT of thermo-sensitive genic male sterility 5 (tms5)-based TGMS lines through ribosome-associated quality control (RQC). Biochemical assays demonstrated that CSIT1 binds to the 80S ribosomes and ubiquitinates abnormal nascent polypeptides for degradation in the RQC process. Loss of CSIT1 function inhibits the possible damage of tms5 to the ubiquitination system and protein translation, resulting in enhanced accumulation of anther-related proteins such as catalase to suppress abnormal accumulation of reactive oxygen species and premature programmed cell death in the tapetum, thereby leading to a much higher CSIT in the tms5-based TGMS lines. Taken together, our findings reveal a regulatory mechanism of CSIT, providing new insights into RQC and potential targets for future two-line hybrid breeding.

Methylesterification of cell-wall pectin controls the diurnal flower-opening times in rice.

Flowers are the core reproductive organ of plants, and flowering is essential for cross-pollination. Diurnal flower-opening time is thus a key trait influencing reproductive isolation, hybrid breeding, and thermostability in plants. However, the molecular mechanisms controlling this trait remain unknown. Here, we report that rice Diurnal Flower Opening Time 1 (DFOT1) modulates pectin methylesterase (PME) activity to regulate pectin methylesterification levels of the lodicule cell walls, which affect lodicule swelling to control diurnal flower-opening time. DFOT1 is specifically expressed in the lodicules, and its expression gradually increases with the approach to flowering but decreases with flowering. Importantly, a knockout of DFOT1 showed earlier diurnal flower opening. We demonstrate that DFOT1 interacts directly with multiple PMEs to promote their activity. Knockout of PME40 also resulted in early diurnal flower opening, whereas overexpression of PME42 delayed diurnal flower opening. Lower PME activity was observed to be associated with higher levels of pectin methylesterification and the softening of cell walls in lodicules, which contribute to the absorption of water by lodicules and cause them to swell, thus promoting early diurnal flower opening. Higher PME activity had the opposite effect. Collectively, our work uncovers a molecular mechanism underlying the regulation of diurnal flower-opening time in rice, which would help reduce the costs of hybrid breeding and improve the heat tolerance of flowering plants by avoiding higher temperatures at anthesis.

The structural characteristics and the substrate recognition properties of RNase ZS1.

TMS5 encodes an RNase ZS1 protein that can process ubiquitin-60S ribosomal protein L40 family (UbL40) mRNAs to regulate thermo-sensitive genic male sterility in rice. Despite the importance of this protein, the structural characteristics and substrate recognition properties of RNase ZS1 remain unclear. Here, we found that the variations in several conservative amino acids alter the activation of RNase ZS1, and its recognition of RNA substrates depends on the structure of RNA. RNase ZS1 acts as a homodimer. The conserved amino acids in or adjacent to enzyme center play a critical role in the enzyme activity of RNase ZS1 and the conserved amino acids that far from active center have little impact on its enzyme activity. The cleavage efficiency of RNase ZS1 for pre-tRNA-MetCAU35 and UbL401 mRNA with cloverleaf-like structure was higher than that of pre-tRNA-AspAUC9 and UbL404 mRNA with imperfect cloverleaf-like structure. This difference implies that the enzyme activity of RNase ZS1 depends on the cloverleaf-like structure of the RNA. Furthermore, the RNase ZS1 activity was not inhibited by the 5' leader sequence and 3' CCA motif of pre-tRNA. These findings provide new insights for studying the cleavage characteristics and substrate recognition properties of RNase ZS.

OsCER1 regulates humidity-sensitive genic male sterility through very-long-chain (VLC) alkane metabolism of tryphine in rice.

Humidity-sensitive genic male sterility (HGMS) is a novel type of environment-sensitive male sterility (EGMS) which plants are male sterile at low humidity and male fertile at high humidity. Previous studies have revealed that OsCER1 contributes to very-long-chain (VLC) alkanes biosynthesis in rice (Oryza sativa L.). Here, applying the CRISPR/Cas9 technique, we obtained two independent OsCER1 knockout lines (OsCER1Cas). Both OsCER1Cas lines exhibited HGMS. Mutant pollen showed defects in adhesion and germination on stigmas at low humidity, whereas high humidity enhanced the pollen germination rate. Transmission electron microscopy (TEM) observations of mutant pollen revealed abnormal tryphine structure, potentially representing the basis of HGMS. Furthermore, co-pollination with mixed OsCER1Cas mutant and maize (Zea mays L.) pollen could rescue the fertility of the mutant, thereby establishing the key role of tryphine in germination on stigmas. OsCER1 knockout might affect VLC alkane metabolism and therefore alter the lipid composition of tryphine. It could lead to the defects in pollen grain adhesion, hydration and germination, resulting in HGMS. This work identified the mechanism of HGMS induced by VLC alkanes in rice and the generality of tryphine in different species of Gramineae.

Expression partitioning between genes duplicated by polyploidy under abiotic stress and during organ development.

Allopolyploidy has been a prominent mode of speciation and a recurrent process during plant evolution and has contributed greatly to the large number of duplicated genes in plant genomes [1-4]. Polyploidy often leads to changes in genome organization and gene expression [5-9]. The expression of genes that are duplicated by polyploidy (termed homeologs) can be partitioned between the duplicates so that one copy is expressed and functions only in some organs and the other copy is expressed only in other organs, indicative of subfunctionalization [10]. To determine how homeologous-gene expression patterns change during organ development and in response to abiotic stress conditions, we have examined expression of the alcohol dehydrogenase gene AdhA in allopolyploid cotton (Gossypium hirsutum). Expression ratios of the two homeologs vary considerably during the development of organs from seedlings and fruits. Abiotic stress treatments, including cold, dark, and water submersion, altered homeologous-gene expression. Most notably, only one copy is expressed in hypocotyls during a water-submersion treatment, and only the other copy is expressed during cold stress. These results imply that subfunctionalization of genes duplicated by polyploidy has occurred in response to abiotic stress conditions. Partitioning of duplicate gene expression in response to environmental stress may lead to duplicate gene retention during subsequent evolution.