Natural variation in LONELY GUY-Like 1 regulates rice grain weight under warmer night conditions.

Global nighttime temperatures are rising at twice the rate of daytime temperatures and pose a challenge for rice (Oryza sativa) production. High nighttime temperature (HNT) stress affects rice yield by reducing grain weight, size, and fertility. Although the genes associated with these yield parameters have been identified and characterized under normal temperatures, the genetic basis of grain weight regulation under HNT stress remains less explored. We examined the natural variation for rice single grain weight (SGW) under HNT stress imposed during grain development. A genome-wide association analysis identified several loci associated with grain weight under HNT stress. A locus, single grain weight 1 (SGW1), specific to HNT conditions resolved to LONELY GUY-Like 1 (LOGL1), which encodes a putative cytokinin activation enzyme. We demonstrated that LOGL1 contributes to allelic variation at SGW1. Accessions with lower LOGL1 transcript abundance had higher grain weight under HNT. This was supported by higher grain weight of logl1 mutants relative to the wild type under HNT. Compared to logl1 mutants, LOGL1 over-expressers showed increased sensitivity to HNT. We showed that LOGL1 regulates the thiamine biosynthesis pathway, which is under circadian regulation, which in turn is likely perturbed by HNT stress. These findings provide a genetic source to enhance rice adaptation to warming night temperatures and improve our mechanistic understanding of HNT stress tolerance pathways.

Rice Chalky Grain 5 regulates natural variation for grain quality under heat stress.

Heat stress occurring during rice (Oryza sativa) grain development reduces grain quality, which often manifests as increased grain chalkiness. Although the impact of heat stress on grain yield is well-studied, the genetic basis of rice grain quality under heat stress is less explored as quantifying grain quality is less tractable than grain yield. To address this, we used an image-based colorimetric assay (Red, R; and Green, G) for genome-wide association analysis to identify genetic loci underlying the phenotypic variation in rice grains exposed to heat stress. We found the R to G pixel ratio (RG) derived from mature grain images to be effective in distinguishing chalky grains from translucent grains derived from control (28/24°C) and heat stressed (36/32°C) plants. Our analysis yielded a novel gene, rice Chalky Grain 5 (OsCG5) that regulates natural variation for grain chalkiness under heat stress. OsCG5 encodes a grain-specific, expressed protein of unknown function. Accessions with lower transcript abundance of OsCG5 exhibit higher chalkiness, which correlates with higher RG values under stress. These findings are supported by increased chalkiness of OsCG5 knock-out (KO) mutants relative to wildtype (WT) under heat stress. Grains from plants overexpressing OsCG5 are less chalky than KOs but comparable to WT under heat stress. Compared to WT and OE, KO mutants exhibit greater heat sensitivity for grain size and weight relative to controls. Collectively, these results show that the natural variation at OsCG5 may contribute towards rice grain quality under heat stress.

Endoplasmic reticulum stress pathway mediates the early heat stress response of developing rice seeds.

A transient heat stress occurring during early seed development in rice (Oryza sativa) reduces seed size by altering endosperm development. However, the relationship between the timing of the stress and specific developmental stage on heat sensitivity is not well-understood. To address this, we imposed a series of non-overlapping heat stress treatments and found that young seeds are most sensitive during the first two days after flowering. Temporal transcriptome analysis of developing, heat stressed (35°C) seeds during this window shows that Inositol-requiring enzyme 1 (IRE1)-mediated endoplasmic reticulum (ER) stress response and jasmonic acid (JA) pathways are the early (1-3 h) drivers of heat stress response. We propose that increased JA levels under heat stress may precede ER stress response as JA application promotes the spliced form of OsbZIP50, an ER response marker gene linked to IRE1-specific pathway. This study presents temporal and mechanistic insights into the role of JA and ER stress signalling during early heat stress response of rice seeds that impact both grain size and quality. Modulating the heat sensitivity of the early sensing pathways and downstream endosperm development genes can enhance rice resilience to transient heat stress events.

Allelic variation in rice Fertilization Independent Endosperm 1 contributes to grain width under high night temperature stress.

A higher minimum (night-time) temperature is considered a greater limiting factor for reduced rice yield than a similar increase in maximum (daytime) temperature. While the physiological impact of high night temperature (HNT) has been studied, the genetic and molecular basis of HNT stress response remains unexplored. We examined the phenotypic variation for mature grain size (length and width) in a diverse set of rice accessions under HNT stress. Genome-wide association analysis identified several HNT-specific loci regulating grain size as well as loci that are common for optimal and HNT stress conditions. A novel locus contributing to grain width under HNT conditions colocalized with Fie1, a component of the FIS-PRC2 complex. Our results suggest that the allelic difference controlling grain width under HNT is a result of differential transcript-level response of Fie1 in grains developing under HNT stress. We present evidence to support the role of Fie1 in grain size regulation by testing overexpression (OE) and knockout mutants under heat stress. The OE mutants were either unaltered or had a positive impact on mature grain size under HNT, while the knockouts exhibited significant grain size reduction under these conditions.

Divergent phenotypic response of rice accessions to transient heat stress during early seed development.

Increasing global surface temperatures is posing a major food security challenge. Part of the solution to address this problem is to improve crop heat resilience, especially during grain development, along with agronomic decisions such as shift in planting time and increasing crop diversification. Rice is a major food crop consumed by more than 3 billion people. For rice, thermal sensitivity of reproductive development and grain filling is well-documented, while knowledge concerning the impact of heat stress (HS) on early seed development is limited. Here, we aim to study the phenotypic variation in a set of diverse rice accessions for elucidating the HS response during early seed development. To explore the variation in HS sensitivity, we investigated aus (1), indica (2), temperate japonica (2), and tropical japonica (4) accessions for their HS (39/35°C) response during early seed development that accounts for transition of endosperm from syncytial to cellularization, which broadly corresponds to 24 and 96 hr after fertilization (HAF), respectively, in rice. The two indica and one of the tropical japonica accessions exhibited severe heat sensitivity with increased seed abortion; three tropical japonicas and an aus accession showed moderate heat tolerance, while temperate japonicas exhibited strong heat tolerance. The accessions exhibiting extreme heat sensitivity maintain seed size at the expense of number of fully developed mature seeds, while the accessions showing relative resilience to the transient HS maintained number of fully developed seeds but compromised on seed size, especially seed length. Further, histochemical analysis revealed that all the tested accessions have delayed endosperm cellularization upon exposure to the transient HS by 96 HAF; however, the rate of cellularization was different among the accessions. These findings were further corroborated by upregulation of cellularization-associated marker genes in the developing seeds from the heat-stressed samples.

SeedExtractor: An Open-Source GUI for Seed Image Analysis.

Accurate measurement of seed size parameters is essential for both breeding efforts aimed at enhancing yields and basic research focused on discovering genetic components that regulate seed size. To address this need, we have developed an open-source graphical user interface (GUI) software, SeedExtractor that determines seed size and shape (including area, perimeter, length, width, circularity, and centroid), and seed color with capability to process a large number of images in a time-efficient manner. In this context, our application takes ∼2 s for analyzing an image, i.e., significantly less compared to the other tools. As this software is open-source, it can be modified by users to serve more specific needs. The adaptability of SeedExtractor was demonstrated by analyzing scanned seeds from multiple crops. We further validated the utility of this application by analyzing mature-rice seeds from 231 accessions in Rice Diversity Panel 1. The derived seed-size traits, such as seed length, width, were used for genome-wide association analysis. We identified known loci for regulating seed length (GS3) and width (qSW5/GW5) in rice, which demonstrates the accuracy of this application to extract seed phenotypes and accelerate trait discovery. In summary, we present a publicly available application that can be used to determine key yield-related traits in crops.