Adaptive evolution of the enigmatic Takakia now facing climate change in Tibet.

The most extreme environments are the most vulnerable to transformation under a rapidly changing climate. These ecosystems harbor some of the most specialized species, which will likely suffer the highest extinction rates. We document the steepest temperature increase (2010-2021) on record at altitudes of above 4,000 m, triggering a decline of the relictual and highly adapted moss Takakia lepidozioides. Its de-novo-sequenced genome with 27,467 protein-coding genes includes distinct adaptations to abiotic stresses and comprises the largest number of fast-evolving genes under positive selection. The uplift of the study site in the last 65 million years has resulted in life-threatening UV-B radiation and drastically reduced temperatures, and we detected several of the molecular adaptations of Takakia to these environmental changes. Surprisingly, specific morphological features likely occurred earlier than 165 mya in much warmer environments. Following nearly 400 million years of evolution and resilience, this species is now facing extinction.

EAR APICAL DEGENERATION1 regulates maize ear development by maintaining malate supply for apical inflorescence.

Ear length (EL) is a key trait that contributes greatly to grain yield in maize (Zea mays). While numerous quantitative trait loci for EL have been identified, few causal genes have been studied in detail. Here we report the characterization of ear apical degeneration1 (ead1) exhibiting strikingly shorter ears and the map-based cloning of the casual gene EAD1. EAD1 is preferentially expressed in the xylem of immature ears and encodes an aluminum-activated malate transporter localizing to the plasma membrane. We show that EAD1 is a malate efflux transporter and loss of EAD1 leads to lower malate contents in the apical part of developing inflorescences. Exogenous injections of malate rescued the shortened ears of ead1. These results demonstrate that EAD1 plays essential roles in regulating maize ear development by delivering malate through xylem vessels to the apical part of the immature ear. Overexpression of EAD1 led to greater EL and kernel number per row and the EAD1 genotype showed a positive association with EL in two different genetic segregating populations. Our work elucidates the critical role of EAD1 in malate-mediated female inflorescence development and provides a promising genetic resource for enhancing maize grain yield.

Rice EIL1 interacts with OsIAAs to regulate auxin biosynthesis mediated by the tryptophan aminotransferase MHZ10/OsTAR2 during root ethylene responses.

Ethylene plays essential roles in adaptive growth of rice (Oryza sativa). Understanding of the crosstalk between ethylene and auxin (Aux) is limited in rice. Here, from an analysis of the root-specific ethylene-insensitive rice mutant mao hu zi 10 (mhz10), we identified the tryptophan aminotransferase (TAR) MHZ10/OsTAR2, which catalyzes the key step in indole-3-pyruvic acid-dependent Aux biosynthesis. Genetically, OsTAR2 acts downstream of ethylene signaling in root ethylene responses. ETHYLENE INSENSITIVE3 like1 (OsEIL1) directly activated OsTAR2 expression. Surprisingly, ethylene induction of OsTAR2 expression still required the Aux pathway. We also show that Os indole-3-acetic acid (IAA)1/9 and OsIAA21/31 physically interact with OsEIL1 and show promotive and repressive effects on OsEIL1-activated OsTAR2 promoter activity, respectively. These effects likely depend on their EAR motif-mediated histone acetylation/deacetylation modification. The special promoting activity of OsIAA1/9 on OsEIL1 may require both the EAR motifs and the flanking sequences for recruitment of histone acetyltransferase. The repressors OsIAA21/31 exhibit earlier degradation upon ethylene treatment than the activators OsIAA1/9 in a TIR1/AFB-dependent manner, allowing OsEIL1 activation by activators OsIAA1/9 for OsTAR2 expression and signal amplification. This study reveals a positive feedback regulation of ethylene signaling by Aux biosynthesis and highlights the crosstalk between ethylene and Aux pathways at a previously underappreciated level for root growth regulation in rice.

The PIF1-miR408-PLANTACYANIN repression cascade regulates light-dependent seed germination.

Light-dependent seed germination is a vital process for many seed plants. A decisive event in light-induced germination is degradation of the central repressor PHYTOCHROME INTERACTING FACTOR 1 (PIF1). The balance between gibberellic acid (GA) and abscisic acid (ABA) helps to control germination. However, the cellular mechanisms linking PIF1 turnover to hormonal balancing remain elusive. Here, employing far-red light-induced Arabidopsis thaliana seed germination as the experimental system, we identified PLANTACYANIN (PCY) as an inhibitor of germination. It is a blue copper protein associated with the vacuole that is both highly expressed in mature seeds and rapidly silenced during germination. Molecular analyses showed that PIF1 binds to the miR408 promoter and represses miR408 accumulation. This in turn posttranscriptionally modulates PCY abundance, forming the PIF1-miR408-PCY repression cascade for translating PIF1 turnover to PCY turnover during early germination. Genetic analysis, RNA-sequencing, and hormone quantification revealed that PCY is necessary and sufficient to maintain the PIF1-mediated seed transcriptome and the low-GA-high-ABA state. Furthermore, we found that PCY domain organization and regulation by miR408 are conserved features in seed plants. These results revealed a cellular mechanism whereby PIF1-relayed external light signals are converted through PCY turnover to internal hormonal profiles for controlling seed germination.

Brassinosteroid-Activated BRI1-EMS-SUPPRESSOR 1 Inhibits Flavonoid Biosynthesis and Coordinates Growth and UV-B Stress Responses in Plants.

UV-B light is a potential stress factor in plants, but how plants coordinate growth and UV-B stress responses is not well understood. Here, we report that brassinosteroid (BR) signaling inhibits UV-B stress responses in Arabidopsis (Arabidopsis thaliana) and various crops by controlling flavonol biosynthesis. We further demonstrate that BRI1-EMS-SUPPRESSOR 1 (BES1) mediates the tradeoff between plant growth and UV-B defense responses. BES1, a master transcription factor involved in BR signaling, represses the expression of transcription factor genes MYB11, MYB12, and MYB111, which activate flavonol biosynthesis. BES1 directly binds to the promoters of these MYBs in a BR-enhanced manner to repress their expression, thereby reducing flavonol accumulation. However, exposure to broadband UV-B down-regulates BES1 expression, thus promoting flavonol accumulation. These findings demonstrate that BR-activated BES1 not only promotes growth but also inhibits flavonoid biosynthesis. UV-B stress suppresses the expression of BES1 to allocate energy to flavonoid biosynthesis and UV-B stress responses, allowing plants to switch from growth to UV-B stress responses in a timely manner.

The GDSL Lipase MHZ11 Modulates Ethylene Signaling in Rice Roots.

Ethylene plays important roles in plant growth and development, but the regulation of ethylene signaling is largely unclear, especially in crops such as rice (Oryza sativa). Here, by analysis of the ethylene-insensitive mutant mao huzi 11 (mhz11), we identified the GDSL lipase MHZ11, which modulates ethylene signaling in rice roots. MHZ11 localized to the endoplasmic reticulum membrane and has acyl-hydrolyzing activity. This activity affects the homeostasis of sterols in rice roots and is required for root ethylene response. MHZ11 overexpression caused constitutive ethylene response in roots. Genetically, MHZ11 acts with the ethylene receptor ETHYLENE RESPONSE SENSOR2 (OsERS2) upstream of CONSTITUTIVE TRIPLE RESPONSE2 (OsCTR2) and ETHYLENE INSENSITIVE2 (OsEIN2). The mhz11 mutant maintains more OsCTR2 in the phosphorylated form whereas MHZ11 overexpression promotes ethylene-mediated inhibition of OsCTR2 phosphorylation. MHZ11 colocalized with the ethylene receptor OsERS2, and its effect on OsCTR2 phosphorylation requires ethylene perception and initiation of ethylene signaling. The mhz11 mutant overaccumulated sterols and blocking sterol biosynthesis partially rescued the mhz11 ethylene response, likely by reducing receptor-OsCTR2 interaction and OsCTR2 phosphorylation. We propose that MHZ11 reduces sterol levels to impair receptor-OsCTR2 interactions and OsCTR2 phosphorylation for triggering ethylene signaling. Our study reveals a mechanism by which MHZ11 participates in ethylene signaling for regulation of root growth in rice.

Brassinosteroid homeostasis is critical for the functionality of the Medicago truncatula pulvinus.

Many plant species open their leaves during the daytime and close them at night as if sleeping. This leaf movement is known as nyctinasty, a unique and intriguing phenomenon that been of great interest to scientists for centuries. Nyctinastic leaf movement occurs widely in leguminous plants, and is generated by a specialized motor organ, the pulvinus. Although a key determinant of pulvinus development, PETIOLULE-LIKE PULVINUS (PLP), has been identified, the molecular genetic basis for pulvinus function is largely unknown. Here, through an analysis of knockout mutants in barrelclover (Medicago truncatula), we showed that neither altering brassinosteroid (BR) content nor blocking BR signal perception affected pulvinus determination. However, BR homeostasis did influence nyctinastic leaf movement. BR activity in the pulvinus is regulated by a BR-inactivating gene PHYB ACTIVATION TAGGED SUPPRESSOR1 (BAS1), which is directly activated by PLP. A comparative analysis between M. truncatula and the non-pulvinus forming species Arabidopsis and tomato (Solanum lycopersicum) revealed that PLP may act as a factor that associates with unknown regulators in pulvinus determination in M. truncatula. Apart from exposing the involvement of BR in the functionality of the pulvinus, these results have provided insights into whether gene functions among species are general or specialized.

Creation of fragrant sorghum by CRISPR/Cas9.

Sorghum, the fifth largest cereal crop, has high value as a staple food and raw material for liquor and vinegar brewing. Due to its high biomass and quality, it is also used as the second most planted silage resource. No fragrant sorghums are currently on the market. Through CRISPR/Cas9-mediated knockout of SbBADH2, we obtained sorghum lines with extraordinary aromatic smell in both seeds and leaves. Animal feeding experiments showed that fragrant sorghum leaves were attractable. We believe this advantage will produce great value in the sorghum market for both grain and whole biomass forage.

AtCPS V326M significantly affect the biosynthesis of gibberellins.

Gibberellins are a class of typical phytohormones, which regulate plant growth and development. The contents of gibberellins dramatically affect the morphology and biomass of plant. The encoding protein of copalyl diphosphate synthase gene (CPS) catalyzes the first-step in the biosynthetic pathway of gibberellins. The mutation in this gene may significantly affect the contents of gibberellins in plants. In this study, we found an EMS-triggered mutant, ga1-168, showing short roots, short hypocotyls, late flowering and dwarf. Map-based cloning revealed that the causal gene of ga1-168 was AtCPS-168, an allele of AtCPS gene. The encoding protein of AtCPS-168 was AtCPS V326M which was resulted from a single-point mutation (guanine to adenine at nucleotide 2768) of AtCPS gene. Protein domain analysis showed that V326 was located in the Terpene_synth domain. The allelism test demonstrated that AtCPS-168 was an allele of AtCPS gene. The transgenic complementation of ga1-168 indicated that AtCPS V326M led to the dwarf and bushy phenotype of ga1-168. The endogenous gibberellins contents analysis suggested that the gibberellins contents of ga1-168 were much lower than that of wild-type. The exogenous GA3 application assay uncovered that application of GA3 can complement the dwarf and bushy phenotype of ga1-168 caused by low endogenous gibberellins contents. Therefore, this study suggested that it is an elegant way to create the ideal plant architecture and height by site-directed mutating the gibberellin biosynthetic genes.

Natural variations in the promoter of Awn Length Inhibitor 1 (ALI-1) are associated with awn elongation and grain length in common wheat.

Wheat awn plays a vital role in photosynthesis, grain production, and drought tolerance. However, the systematic identification or cloning of genes controlling wheat awn development is seldom reported. Here, we conducted a genome-wide association study (GWAS) with 364 wheat accessions and identified 26 loci involved in awn length development, including previously characterized B1, B2, Hd, and several rice homologs. The dominant awn suppressor B1 was fine mapped to a 125-kb physical interval, and a C2 H2 zinc finger protein Awn Length Inhibitor 1 (ALI-1) was confirmed to be the underlying gene of the B1 locus through the functional complimentary test with native awnless allele. ALI-1 expresses predominantly in the developing spike of awnless individuals, transcriptionally suppressing downstream genes. ALI-1 reduces cytokinin content and simultaneously restrains cytokinin signal transduction, leading to a stagnation of cell proliferation and reduction of cell numbers during awn development. Polymorphisms of four single nucleotide polymorphisms (SNPs) located in ALI-1 promoter region are diagnostic for the B1/b1 genotypes, and these SNPs are associated with awn length (AL), grain length (GL) and thousand-grain weight (TGW). More importantly, ali-1 was observed to increase grain length in wheat, which is a valuable attribute of awn on grain weight, aside from photosynthesis. Therefore, ALI-1 pleiotropically regulates awn and grain development, providing an alternative for grain yield improvement and addressing future climate changes.

A wheat dominant dwarfing line with Rht12, which reduces stem cell length and affects gibberellic acid synthesis, is a 5AL terminal deletion line.

Dwarfing and semi-dwarfing are important agronomic traits that have great potential for the improvement of wheat yields. Rht12, a dominant gibberellic acid (GA)-responsive dwarfing gene from the gamma-ray-induced wheat mutant Karcagi 522M7K, is located in the long arm of chromosome 5A, which is closely linked with the locus Xwmc410. Rht12 is likely an ideal gene for GA biosynthesis and deactivation research in common wheat. However, information on the Rht12 locus and sequence is lacking. In this study, Rht12 significantly shortened stem cell length and decreased GA biosynthetic components. Using bulked segregant RNA-Seq, wheat 660k single nucleotide polymorphism chip detection, and newly developed simple sequence repeat markers, Rht12 was mapped to a 11.21-Mb region at the terminal end of chromosome 5AL, and was found to be closely linked with the Xw5ac207SSR marker with a 10.73-Mb fragment deletion in all of the homologous dwarfing plants. Transcriptome analyses of the remaining 483-kb region showed significantly higher expression of the TraesCS5A01G543100 gene encoding the GA metabolic enzyme GA 2-ß-dioxygenase in dwarfing plants than in high stalk plants, suggesting that Rht12 reduces plant height by activating TaGA2ox-A14. Taken together, our findings will promote cloning and functional studies of Rht12 in common wheat.

The Arabidopsis TRM61/TRM6 complex is a bona fide tRNA N1-methyladenosine methyltransferase.

tRNA molecules, which contain the most abundant post-transcriptional modifications, are crucial for proper gene expression and protein biosynthesis. Methylation at N1 of adenosine 58 (A58) is critical for maintaining the stability of initiator methionyl-tRNA (tRNAiMet) in bacterial, archaeal, and eukaryotic tRNAs. However, although research has been conducted in yeast and mammals, it remains unclear how A58 in plant tRNAs is modified and involved in development. In this study, we identify the nucleus-localized complex AtTRM61/AtTRM6 in Arabidopsis as tRNA m1A58 methyltransferase. Deficiency or a lack of either AtTRM61 or AtTRM6 leads to embryo arrest and seed abortion. The tRNA m1A level decreases in conditionally complemented Attrm61/LEC1pro::AtTRM61 plants and this is accompanied by reduced levels of tRNAiMet, indicating the importance of the tRNA m1A modification for tRNAiMet stability. Taken together, our results demonstrate that tRNA m1A58 modification is necessary for tRNAiMet stability and is required for embryo development in Arabidopsis.

Arabidopsis WRKY46, WRKY54, and WRKY70 Transcription Factors Are Involved in Brassinosteroid-Regulated Plant Growth and Drought Responses.

Plant steroid hormones, brassinosteroids (BRs), play important roles in growth and development. BR signaling controls the activities of BRASSINOSTERIOD INSENSITIVE1-EMS-SUPPRESSOR1/BRASSINAZOLE-RESISTANT1 (BES1/BZR1) family transcription factors. Besides the role in promoting growth, BRs are also implicated in plant responses to drought stress. However, the molecular mechanisms by which BRs regulate drought response have just begun to be revealed. The functions of WRKY transcription factors in BR-regulated plant growth have not been established, although their roles in stress responses are well documented. Here, we found that three Arabidopsis thaliana group III WRKY transcription factors, WRKY46, WRKY54, and WRKY70, are involved in both BR-regulated plant growth and drought response as the wrky46 wrky54 wrky70 triple mutant has defects in BR-regulated growth and is more tolerant to drought stress. RNA-sequencing analysis revealed global roles of WRKY46, WRKY54, and WRKY70 in promoting BR-mediated gene expression and inhibiting drought responsive genes. WRKY54 directly interacts with BES1 to cooperatively regulate the expression of target genes. In addition, WRKY54 is phosphorylated and destabilized by GSK3-like kinase BR-INSENSITIVE2, a negative regulator in the BR pathway. Our results therefore establish WRKY46/54/70 as important signaling components that are positively involved in BR-regulated growth and negatively involved in drought responses.

Cellular and molecular characterization of a thick-walled variant reveal a pivotal role of shoot apical meristem in transverse development of bamboo culm.

Little is known about the mechanisms underlying the development of bamboo culm. Using anatomical, mathematical modeling, and genomics methods, we investigated the role of shoot apical meristem (SAM) in the development of the transverse morphology of bamboo culm and explored the underlying cellular and molecular processes. We discovered that maintenance of SAM morphology that can produce circular culm and increase in SAM cell numbers, especially corpus cells, is the means by which bamboo makes a larger culm with a regular pith cavity and culm wall during development. A less cellular form of SAM with a lower proportion of corpus cells causes an abnormal higher ratio of wall component cells to pith cells, which breaks the balance of their interaction and triggers the random invasion of wall component cells into pith tissues during development, and finally results in the various thick culm walls of Phyllostachys nidularia f. farcta. The smaller SAM also results in a lower level of hormones such as cytokinin and auxin, and down-regulates hormone signaling and the downstream functional genes such as those related to metabolism, which finally results in a dwarf and smaller diameter culm with lower biomass. These results provide an important perspective on the culm development of bamboo, and support a plausible mechanism causing the size-reduced culm and various thick culm walls of P. nidularia f. farcta.

Pursuing extreme sensitivity for determination of endogenous brassinosteroids through direct fishing from plant matrices and eliminating most interferences with boronate affinity magnetic nanoparticles.

Brassinosteroids (BRs) are important plant hormones regulating plant growth and development. High-performance analytical methods for quantifying endogenous BRs are important for studying the molecular mechanisms of BR action. Herein we developed a high-performance sample pretreatment method based on boronate affinity magnetic nanoparticles (BAMNPs). The high specificity of boronate affinity enables direct fishing of BRs from plant matrices. The strong binding energy makes it possible to remove most contaminants in plant matrices with a small loss of target BRs. Besides these advantages, the novel two-step oxidation-hydrolysis elution system raised BR recoveries to 70.5%-98.2%, which was much higher than other boronate affinity applications. The high cleanliness of the final eluents lowered the matrix effects to 85.2%-92.4%. As a result, this method enables simultaneously good recoveries of endogenous BRs and thorough removal of matrix interferences, which greatly improves the sensitivity of BR analysis and reduces the use of plant materials for routine analysis to <10 mg. In addition, the sample handling time can be shortened to <3 h due to the operating convenience of BAMNPs and their easy separation from plant powders. Based on these advantages of BAMNP solid phase extraction, the organ-specific BR distribution analysis in Arabidopsis and rice tissues demonstrates excellent sensitivity, good reproducibility and high throughput of the method. Graphical abstract A high-sensitivity and time-saving UPLC-MS/MS-based quantification method for brassinosteroids (BRs) was developed through directly fishing BRs from plant matrices and eliminating most matrix interferences with as-prepared boronate affinity magnetic nanoparticles (BAMNPs).

An improved simplified high-sensitivity quantification method for determining brassinosteroids in different tissues of rice and Arabidopsis.

Quantification of brassinosteroids is essential and extremely important to study the molecular mechanisms of their physiological roles in plant growth and development. Herein, we present a simple, material and cost-saving high-performance method for determining endogenous brassinosteroids (BRs) in model plants. This new method enables simultaneous enrichment of a wide range of bioactive BRs such as brassinolide, castasterone, teasterone, and typhasterol with ion exchange solid-phase extraction and high-sensitivity quantitation of these BRs based on isotope dilution combined with internal standard approach. For routine analysis, the consumption of plant materials was reduced to one-twentieth of previously reported and the overall process could be completed within 1 day compared with previous 3 to 4 days. The strategy was validated by profiling BRs in different ecotypes and mutants of rice (Oryza sativa) and Arabidopsis (Arabidopsis thaliana), and the BR distributions in different model plants tissues were determined with the new method. The method allows plant physiologists to monitor the dynamics and distributions of BRs with 1 gram fresh weight of model plant tissues, which will speed up the process for the molecular mechanism research of BRs with these model plants in future work.

Tomato CYP94C1 inactivates bioactive JA-Ile to attenuate jasmonate-mediated defense during fruit ripening.

As the master regulators of the ET signaling pathway, EIL transcription factors directly activate the expression of CYP94C1 to inactivate bioactive JA-Ile, thereby attenuating JA-mediated defense during fruit ripening. Knockout of CYP94C1 improves tomato fruit resistance to necrotrophs without compromising fruit quality.

A dual role of boronate affinity in high-sensitivity detection of vicinal diol brassinosteroids from sub-gram plant tissues via UPLC-MS/MS.

Based on the dual role of specific boronate affinity, making use of both novel self-synthesized boronate affinity-functionalized magnetic nanoparticles and a high-efficiency organic boronic acid-type derivatization reagent, we report a simple, convenient and highly-sensitive method for detection of endogenous brassinosteroids from real plant materials.

SnoRNP is essential for thermospermine-mediated development in Arabidopsis thaliana.

Polyamines have been discovered for hundreds of years and once considered as a class of phytohormones. Polyamines play critical roles in a range of developmental processes. However, the molecular mechanisms of polyamine signaling pathways remain poorly understood. Here, we measured the contents of main types of polyamines, and found that endogenous level of thermospermine (T-Spm) in Arabidopsis thaliana is comparable to those of classic phytohormones and is significantly lower than those of putrescine (Put), spermidine (Spd), and spermine (Spm). We further found a nodule-like structure around the junction area connecting the shoot and root of the T-Spm biosynthetic mutant acl5 and obtained more than 50 suppressors of acl5nodule structure (san) through suppressor screening. An in-depth study of two san suppressors revealed that NAP57 and NOP56, core components of box H/ACA and C/D snoRNPs, were essential for T-Spm-mediated nodule-like structure formation and plant height. Furthermore, analyses of rRNA modifications showed that the overall levels of pseudouridylation and 2'-O-methylation were compromised in san1 and san2 respectively. Taken together, these results establish a strong genetic relationship between rRNA modification and T-Spm-mediated growth and development, which was previously undiscovered in all organisms.

A well-defined block copolymer serving as surfactants in separation of 1,4-dihydropyridines by open-tubular capillary eletrochromatography.

A novel series of diblock copolymers, poly(butyl methacrylate)(n) -block-poly(glycidyl methacrylate)(m) [P(BMA)(n) -b-P(GMA)(m) ], were synthesized by atom transfer radical polymerization and developed as covalent coating of capillaries. The excellent performance of this coating in separation of three 1,4-dihydropyridines (DHPs) derivatives (amlodipine, nicardipine, nitrendipine) was achieved when the diblock copolymers self-assembled into micelles, which was confirmed by transmission electron microscopy, dynamic light scattering, and atom force microscopy. Meanwhile, the effects of block ratio n/m, pH value, buffer concentration, and organic solvents on the separation of 1,4-DHPs were investigated in detail. Then, the relationship between the morphologies of copolymers and the separation resolutions of 1,4-DHPs was discussed. Furthermore, the proposed method exhibited good run-to-run and column-to-column precision with relative standard deviations of electroosmotic flow less than 3.0%. It was also validated with linearity of three 1,4-DHPs in the range of 0.01-1.80 mM (r(2) ≥ 99.7%), efficient recovery (94-103%), and good repeatability (≤ 3.8%). In addition, three 1,4-DHPs were successfully separated in the spiked human serum sample, which indicated the potential utility of this method in biological sample analysis.