Suppression of seed germination at supraoptimal high temperature (thermoinhibiton) during summer is crucial for Arabidopsis (Arabidopsis thaliana) to establish vegetative and reproductive growth in appropriate seasons. Abscisic acid (ABA) and gibberellins (GAs) are well known to be involved in germination control, but it remains unknown how these hormone actions (metabolism and responsiveness) are altered at high temperature. Here, we show that ABA levels in imbibed seeds are elevated at high temperature and that this increase is correlated with up-regulation of the zeaxanthin epoxidase gene ABA1/ZEP and three 9-cis-epoxycarotenoid dioxygenase genes, NCED2, NCED5, and NCED9. Reverse-genetic studies show that NCED9 plays a major and NCED5 and NCED2 play relatively minor roles in high temperature-induced ABA synthesis and germination inhibition. We also show that bioactive GAs stay at low levels at high temperature, presumably through suppression of GA 20-oxidase genes, GA20ox1, GA20ox2, and GA20ox3, and GA 3-oxidase genes, GA3ox1 and GA3ox2. Thermoinhibition-tolerant germination of loss-of-function mutants of GA negative regulators, SPINDLY (SPY) and RGL2, suggests that repression of GA signaling is required for thermoinibition. Interestingly, ABA-deficient aba2-2 mutant seeds show significant expression of GA synthesis genes and repression of SPY expression even at high temperature. In addition, the thermoinhibition-resistant germination phenotype of aba2-1 seeds is suppressed by a GA biosynthesis inhibitor, paclobutrazol. We conclude that high temperature stimulates ABA synthesis and represses GA synthesis and signaling through the action of ABA in Arabidopsis seeds.

Seeds monitor the environment to germinate at the proper time, but different species respond differently to environmental conditions, particularly light and temperature. In Arabidopsis thaliana, light promotes germination but high temperature suppresses germination. We previously reported that light promotes germination by repressing SOMNUS (SOM). Here, we examined whether high temperature also regulates germination through SOM and found that high temperature activates SOM expression. Consistent with this, som mutants germinated more frequently than the wild type at high temperature. The induction of SOM mRNA at high temperature required abscisic acid (ABA) and gibberellic acid biosynthesis, and ABA-INSENSITIVE3 (ABI3), ABI5, and DELLAs positively regulated SOM expression. Chromatin immunoprecipitation assays indicated that ABI3, ABI5, and DELLAs all target the SOM promoter. At the protein level, ABI3, ABI5, and DELLAs all interact with each other, suggesting that they form a complex on the SOM promoter to activate SOM expression at high temperature. We found that high-temperature-inducible genes frequently have RY motifs and ABA-responsive elements in their promoters, some of which are targeted by ABI3, ABI5, and DELLAs in vivo. Taken together, our data indicate that ABI3, ABI5, and DELLAs mediate high-temperature signaling to activate the expression of SOM and other high-temperature-inducible genes, thereby inhibiting seed germination.

The endophytic bacteria in the seeds of rice plants (Oryza sativa, cultivar Kinuhikari) cultivated on an experimental plot adjacent to a paddy field were studied as the seeds matured by comparing them with the bacteria on the surface of the seeds. Endophytic and surface bacteria were isolated using a nutrient broth and a diluted nutrient broth agar medium. The isolates were identified based on 16S rRNA gene sequences. Three genera (Paenibacillus, Acidovorax and Pantoea) and 2 genera (Stenotrophomonas and Rhizobium) were specific to the inside and to the surface of the seeds, respectively. Six genera (Bacillus, Curtobacterium, Methylobacterium, Sphingomonas, Xanthomonas and Micrococcus) were common to both the inside and the surface. As the seed matured, the flora of culturable endophytic bacteria changed in a different manner from that of culturable surface bacteria. More isolates tolerant of high osmotic pressure were found among the endophytes than among the surface bacteria, especially at the later stages of the maturation process. An increasing number of endophytic isolates exhibited amylase activity at the later stages.

Endophytic bacteria are considered to originate from the external environment. To examine the hypothesis that rice (Oryza sativa, cultivar Kinuhikari) seeds are a source of endophytic bacteria, we isolated endophytic bacteria from the shoots, remains of the seeds, and roots of rice seedlings that were aseptically cultivated in vitro from surfacedisinfected seeds. Of the various bacterial strains isolated, the closest relatives, identified by 16S rRNA gene sequencing, were: Bacillus firmus, B. fusiformis, B. pumilus, Caulobacter crescentus, Kocuria palustris, Micrococcus luteus, Methylobacterium fujisawaense, Me. radiotolerans, and Pantoea ananatis. The latter three species have been detected frequently inside both rice seedlings and mature rice plants. These results indicate that rice seeds are an important source of endophytic bacteria. The bacteria that colonize the seed interior appear to infect the subsequent generation via rice seeds and become the dominant endophytic species in the mature plant.Key words: culturable endophytic bacteria, rice seed, infection from seed to shoot, 16S rRNA gene sequence Various types of microorganisms, including fungi, actinomycetes, and other bacteria, are found inside plants and designated endophytes. In general, microorganisms having visibly harmful effects on plants are not considered endophytes. Some endophytic bacteria have beneficial effects on the host, e.g., promoting plant growth, strengthening resistance to pathogens, and increasing the supply of fixed nitrogen to the plant (9). Studies are underway to exploit these beneficial features for agriculture. However, our knowledge of endophytic bacterial ecology, including the origins of these bacteria, remains incomplete.Endophytic bacteria are considered to originate from the external environment and to enter the plant through the stomata, lenticles, wounds (including broken trichomes), areas of emergence of lateral roots, and germinating radicles (12). The notion of seeds as a source of endophytic bacteria remains controversial (9). Sato (39) did not detect endophytic bacteria in rice seeds. Mundt and Hinkle (27) obtained endophytes only from the damaged seeds of 28 plant types other than rice. Endophytic bacteria were detected in cotton seeds at viable population densities that ranged from 1×10 3 to 1×105 CFU g −1 fresh weight, as well as in the seeds of sweet corn at viable population densities of <1×10 CFU g −1 fresh weight (26). Endophytic bacterial populations increased from nondetectable levels in cotton seeds to detectable levels in radicles 4 days after placing surface-disinfected cotton seeds on water agar (1).Rice (Oryza sativa) is the most important cereal crop in the world, feeding more than 50% of the global population (8). In our search for endophytic bacteria in rice plants, we detected culturable endophytic bacteria in rice seeds at viable population densities that ranged from 10 2 to 10 6 CFU g −1 fresh weight (22,31). Endophytes in seeds may be carried over to the subsequent generation. Thus, in the p...

HighlightXyloglucan oligosaccharide metabolism by α-xylosidase impacts xyloglucan remodelling, the mechanical integrity of the primary cell wall of growing tissues, cell expansion, and seed germination.

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Lutein is an industrially important carotenoid pigment, which is essential for photoprotection and photosynthesis in plants. It is crucial for maintaining human health due to its protective ability from ocular diseases. However, its pathway engineering research has scarcely been performed for microbial production using heterologous hosts, such as Escherichia coli, since the engineering of multiple genes are required there. These genes, which include tricky key carotenoid biosynthesis genes typically derived from plants, encode two sorts of cyclases (lycopene ε- and β-cyclase) and cytochrome P450 CYP97C. In this study, upstream genes effective for the increase in carotenoid amounts, such as isopentenyl diphosphate (IPP) isomerase (IDI) gene, were integrated into the E. coli JM101 (DE3) genome. The most efficient set of the key genes (MpLCYe, MpLCYb, and MpCYP97C) was selected from among corresponding genes derived from various plant (or bacterial) species using E. coli that had accumulated carotenoid substrates. Furthermore, to optimize the production of lutein in E. coli, we introduced several sorts of plasmids that contained some of the multiple genes into the genome-inserted strain and compared lutein productivity. Finally, we achieved 11 mg/L as lutein yield at a mini jar level. Here, high-yield production of lutein was successfully performed using E. coli through approaches of pathway engineering. The findings obtained here should be a base reference for substantial lutein production with microorganisms in the future.