Phytohormone profiling in an evolutionary framework.

The genomes of charophyte green algae, close relatives of land plants, typically do not show signs of developmental regulation by phytohormones. However, scattered reports of endogenous phytohormone production in these organisms exist. We performed a comprehensive analysis of multiple phytohormones in Viridiplantae, focusing mainly on charophytes. We show that auxin, salicylic acid, ethylene and tRNA-derived cytokinins including cis-zeatin are found ubiquitously in Viridiplantae. By contrast, land plants but not green algae contain the trans-zeatin type cytokinins as well as auxin and cytokinin conjugates. Charophytes occasionally produce jasmonates and abscisic acid, whereas the latter is detected consistently in land plants. Several phytohormones are excreted into the culture medium, including auxin by charophytes and cytokinins and salicylic acid by Viridiplantae in general. We note that the conservation of phytohormone biosynthesis and signaling pathways known from angiosperms does not match the capacity for phytohormone biosynthesis in Viridiplantae. Our phylogenetically guided analysis of established algal cultures provides an important insight into phytohormone biosynthesis and metabolism across Streptophyta.

Light Supplementation in Pitaya Orchards Induces Pitaya Flowering in Winter by Promoting Phytohormone Biosynthesis.

The interaction between light and phytohormones is crucial for plant growth and development. The practice of supplementing light at night during winter to promote pitaya flowering and thereby enhance yield has been shown to be crucial and widely used. However, it remains unclear how supplemental winter light regulates phytohormone levels to promote flowering in pitaya. In this study, through analyzing the transcriptome data of pitaya at four different stages (NL, L0, L1, L2), we observed that differentially expressed genes (DEGs) were mainly enriched in the phytohormone biosynthesis pathway. We further analyzed the data and found that cytokinin (CK) content first increased at the L0 stage and then decreased at the L1 and L2 stages after supplemental light treatment compared to the control (NL). Gibberellin (GA), auxin (IAA), salicylic acid (SA), and jasmonic acid (JA) content increased during the formation of flower buds (L1, L2 stages). In addition, the levels of GA, ethylene (ETH), IAA, and abscisic acid (ABA) increased in flower buds after one week of development (L2f). Our results suggest that winter nighttime supplemental light can interact with endogenous hormone signaling in pitaya, particularly CK, to regulate flower bud formation. These results contribute to a better understanding of the mechanism of phytohormone interactions during the induction of flowering in pitaya under supplemental light in winter.

Cytokinin-deficient Chlamydomonas reinhardtii CRISPR-Cas9 mutants show reduced ability to prime resistance of tobacco against bacterial infection.

Although microalgae have only recently been recognized as part of the plant and soil microbiome, their application as biofertilizers has a tradition in sustainable crop production. Under consideration of their ability to produce the plant growth-stimulating hormone cytokinin (CK), known to also induce pathogen resistance, we have assessed the biocontrol ability of CK-producing microalgae. All pro- and eukaryotic CK-producing microalgae tested were able to enhance the tolerance of tobacco against Pseudomonas syringae pv. tabaci (PsT) infection. Since Chlamydomonas reinhardtii (Cre) proved to be the most efficient, we functionally characterized its biocontrol ability. We employed the CRISPR-Cas9 system to generate the first knockouts of CK biosynthetic genes in microalgae. Specifically, we targeted Cre Lonely Guy (LOG) and isopentenyltransferase (IPT) genes, the key genes of CK biosynthesis. While Cre wild-type exhibits a strong protection, the CK-deficient mutants have a reduced ability to induce plant defence. The degree of protection correlates with the CK levels, with the IPT mutants showing less protection than the LOG mutants. Gene expression analyses showed that Cre strongly stimulates tobacco resistance through defence gene priming. This study functionally verifies that Cre primes defence responses with CK, which contributes to the robustness of the effect. This work contributes to elucidate microalgae-mediated plant defence priming and identifies the role of CKs. In addition, these results underscore the potential of CK-producing microalgae as biologicals in agriculture by combining biofertilizer and biocontrol ability for sustainable and environment-friendly crop management.

[Identification and expression profile analysis of rice CRF gene family].

Cytokinin response factors (CRFs), as unique transcription factors in plants, play crucial roles in regulating development, phytohormone signaling pathway, and stress responses. In this study, we identified nine CRF genes from the rice genome by conducting a BLAST analysis using the protein sequences of twelve Arabidopsis AtCRFs. These genes are located on seven different rice chromosomes. We conducted a comprehensive analysis of the conserved domains, physicochemical properties, secondary structures, and phylogenetic relationships of rice CRF proteins using various online tools and local software. Additionally, we analyzed the exon-intron structures and cis-acting elements of OsCRFs, and found an abundance of elements relevant to phytohormone response and stress response on the promoters of rice CRF genes. Spatial-temporal expression pattern analysis revealed that four of the OsCRFs were barely expressed in all tested samples, while the other five were highly expressed in the leaf, panicle, or seed of rice. Microarray data showed that OsCRF genes are regulated to varying degrees by abscisic acid, auxin, cytokinin, and jasmonic acid. Furthermore, through analyzing the RNA-seq data, we found that OsCRFs are primarily involved in plant response to temperature stress (chilling and heat), with several OsCRFs also implicated in drought response, while hardly any respond to salt stress. This study provides an important basis for the functional characterization of rice CRF family genes.

CEPs suppress auxin signaling but promote cytokinin signaling to inhibit root growth in Arabidopsis.

C-terminally encoded peptides (CEPs) are peptide hormones that function as mobile signals coordinating crucial developmental programs in plants. Previous studies have revealed that CEPs exert negative regulation on root development through interaction with CEP receptors (CEPRs), CEP DOWNSTREAMs (CEPDs), the cytokinin receptor ARABIDOPSIS HISTIDINE KINASE (AHKs) and the transcriptional repressor Auxin/Indole-3-Acetic Acid (AUX/IAA). However, the precise molecular mechanisms underlying CEPs-mediated regulation of root development via auxin and cytokinin signaling pathways still necessitate further detailed investigation. In this study, we examined prior research and elucidated the underlying molecular mechanisms. The results showed that both synthetic AtCEPs and overexpression of AtCEP5 markedly supressed primary root elongation and lateral root (LR) formation in Arabidopsis. Molecular biology and genetics elucidated how CEPs inhibit root growth by suppressing auxin signaling while promoting cytokinin signaling. In summary, this study elucidated the inhibitory effects of AtCEPs on Arabidopsis root growth and provided insights into their potential molecular mechanisms, thus enhancing our comprehension of CEP-mediated regulation of plant growth and development.

Longistylin A from Cajanus cajan (L.) Millsp. disturbs glycerophospholipid metabolism and cytokinin biosynthesis of Nocardia seriolae.

ETHNOPHARMACOLOGICAL RELEVANCE: Nocardiosis is an uncommon infectious disease that bears certain similarities to tuberculosis, with a continuous increase in its incidence and a poor prognosis. In traditional Chinese medicine, the leaves of Cajanus cajan (L.) Millsp. are employed to treat wounds, malaria, coughs, and abdominal pain. AIM OF THE STUDY: In this study, we investigated the effects and mechanisms of longistylin A (LGA), a natural stilbene isolated from C. cajan, as a potential antibiotic against nocardiosis. MATERIALS AND METHODS: LGA was isolated from the leaves of C. cajan and assessed using a minimum bactericidal concentration (MBC) determination against Nocardia seriolae. Multi-omics analysis encompassing genes, proteins, and metabolites was conducted to investigate the impact of LGA treatment on N. seriolae. Additionally, quantitative analysis of 40 cytokinins in N. seriolae mycelium was performed to assess the specific effects of LGA treatment on cytokinin levels. Cryo-scanning electron microscopy was utilized to examine morphological changes induced by LGA treatment, particularly in the presence of exogenous trans-zeatin-O-glucoside (tZOG). The therapeutic effect of LGA was investigated by feeding N. seriolae-infected largemouth bass. RESULTS: LGA exhibited significant efficacy against N. seriolae, with MBC value of 2.56 µg/mL. Multi-omics analysis revealed that LGA disrupted glycerophospholipid metabolism and hormone biosynthesis by notably reducing the expression of glycerol-3-phosphate dehydrogenase and calmodulin-like protein. Treatment with LGA markedly disrupted 12 distinct cytokinins in N. seriolae mycelium. Additionally, the addition of exogenous tZOG counteracted the inhibitory effects of LGA on filamentous growth, resulting in mycelial elongation and branching. Furthermore, LGA treatment improved the survival rate of largemouth bass infected with N. seriolae. CONCLUSIONS: We found for the first time that LGA from C. cajan exhibited significant efficacy against N. seriolae by interfering with glycerophospholipid metabolism and cytokinin biosynthesis.

Nitric oxide and cytokinin cross-talk and their role in plant hypoxia response.

Nitric oxide (NO) and cytokinins (CKs) are known for their crucial contributions to plant development, growth, senescence, and stress response. Despite the importance of both signals in stress responses, their interaction remains largely unexplored. The interplay between NO and CKs emerges as particularly significant not only regarding plant growth and development but also in addressing plant stress response, particularly in the context of extreme weather events leading to yield loss. In this review, we summarize NO and CKs metabolism and signaling. Additionally, we emphasize the crosstalk between NO and CKs, underscoring its potential impact on stress response, with a focus on hypoxia tolerance. Finally, we address the most urgent questions that demand answers and offer recommendations for future research endeavors.

DNAL7, a new allele of NAL11, has major pleiotropic effects on rice architecture.

MAIN CONCLUSION: dnal7, a novel allelic variant of the OsHSP40, affects rice plant architecture and grain yield by coordinating auxins, cytokinins, and gibberellic acids. Plant height and leaf morphology are the most important traits of the ideal plant architecture (IPA), and discovering related genes is critical for breeding high-yield rice. Here, a dwarf and narrow leaf 7 (dnal7) mutant was identified from a γ-ray treated mutant population, which exhibits pleiotropic effects, including dwarfing, narrow leaves, small seeds, and low grain yield per plant compared to the wild type (WT). Histological analysis showed that the number of veins and the distance between adjacent small veins (SVs) were significantly reduced compared to the WT, indicating that DNAL7 controls leaf size by regulating the formation of veins. Map-based cloning and transgenic complementation revealed that DNAL7 is allelic to NAL11, which encodes OsHSP40, and the deletion of 2 codons in dnal7 destroyed the His-Pro-Asp (HPD) motif of OsHSP40. In addition, expression of DNAL7 in both WT and dnal7 gradually increased with the increase of temperature in the range of 27-31 °C. Heat stress significantly affected the seedling height and leaf width of the dnal7 mutant. A comparative transcriptome analysis of WT and dnal7 revealed that DNAL7 influenced multiple metabolic pathways, including plant hormone signal transduction, carbon metabolism, and biosynthesis of amino acids. Furthermore, the contents of the cytokinins in leaf blades were much higher in dnal7 than in the WT, whereas the contents of auxins were lower in dnal7. The contents of bioactive gibberellic acids (GAs) including GA1, GA3, and GA4 in shoots were decreased in dnal7. Thus, DNAL7 regulates rice plant architecture by coordinating the balance of auxins, cytokinins, and GAs. These results indicate that OsHSP40 is a pleiotropic gene, which plays an important role in improving rice yield and plant architecture.

OsRR26, a type-B response regulator, modulates salinity tolerance in rice via phytohormone-mediated ROS accumulation in roots and influencing reproductive development.

MAIN CONCLUSION: OsRR26 is a cytokinin-responsive response regulator that promotes phytohormone-mediated ROS accumulation in rice roots, regulates seedling growth, spikelet fertility, awn development, represses NADPH oxidases, and negatively affects salinity tolerance. Plant two-component systems (TCS) play a pivotal role in phytohormone signaling, stress responses, and circadian rhythm. However, a significant knowledge gap exists regarding TCS in rice. In this study, we utilized a functional genomics approach to elucidate the role of OsRR26, a type-B response regulator in rice. Our results demonstrate that OsRR26 is responsive to cytokinin, ABA, and salinity stress, serving as the ortholog of Arabidopsis ARR11. OsRR26 primarily localizes to the nucleus and plays a crucial role in seedling growth, spikelet fertility, and the suppression of awn development. Exogenous application of cytokinin led to distinct patterns of reactive oxygen species (ROS) accumulation in the roots of both WT and transgenic plants (OsRR26OE and OsRR26KD), indicating the potential involvement of OsRR26 in cytokinin-mediated ROS signaling in roots. The application of exogenous ABA resulted in varied cellular compartmentalization of ROS between the WT and transgenic lines. Stress tolerance assays of these plants revealed that OsRR26 functions as a negative regulator of salinity stress tolerance across different developmental stages in rice. Physiological and biochemical analyses unveiled that the knockdown of OsRR26 enhances salinity tolerance, characterized by improved chlorophyll retention and the accumulation of soluble sugars, K+ content, and amino acids, particularly proline.

The Long-Distance Transport of Some Plant Hormones and Possible Involvement of Lipid-Binding and Transfer Proteins in Hormonal Transport.

Adaptation to changes in the environment depends, in part, on signaling between plant organs to integrate adaptive response at the level of the whole organism. Changes in the delivery of hormones from one organ to another through the vascular system strongly suggest that hormone transport is involved in the transmission of signals over long distances. However, there is evidence that, alternatively, systemic responses may be brought about by other kinds of signals (e.g., hydraulic or electrical) capable of inducing changes in hormone metabolism in distant organs. Long-distance transport of hormones is therefore a matter of debate. This review summarizes arguments for and against the involvement of the long-distance transport of cytokinins in signaling mineral nutrient availability from roots to the shoot. It also assesses the evidence for the role of abscisic acid (ABA) and jasmonates in long-distance signaling of water deficiency and the possibility that Lipid-Binding and Transfer Proteins (LBTPs) facilitate the long-distance transport of hormones. It is assumed that proteins of this type raise the solubility of hydrophobic substances such as ABA and jasmonates in hydrophilic spaces, thereby enabling their movement in solution throughout the plant. This review collates evidence that LBTPs bind to cytokinins, ABA, and jasmonates and that cytokinins, ABA, and LBTPs are present in xylem and phloem sap and co-localize at sites of loading into vascular tissues and at sites of unloading from the phloem. The available evidence indicates a functional interaction between LBTPs and these hormones.

PGI1-mediated vascular oxidative pentose phosphate pathway modulates photosynthesis via long-distance cytokinin signaling.

In Arabidopsis, the plastidial isoform of phosphoglucose isomerase, PGI1, mediates growth and photosynthesis, likely due to its involvement in the vascular production of cytokinins (CK). To examine this hypothesis, we characterized pgi1-2 knockout plants impaired in PGI1 and pgi1-2 plants specifically expressing PGI1 in root tips and vascular tissues. Moreover, to investigate whether the phenotype of pgi1-2 plants is due to impairments in the plastidial oxidative pentose phosphate pathway (OPPP) or the glycolytic pathway, we characterized pgl3-1 plants with reduced OPPP and pfk4pfk5 knockout plants impaired in plastidial glycolysis. Compared with wild-type (WT) leaves, pgi1-2 leaves exhibited weaker expression of photosynthesis- and 2-C-methyl-D-erythritol 4-P (MEP) pathway-related proteins, and stronger expression of oxidative stress protection-related enzymes. Consistently, pgi1-2 leaves accumulated lower levels of chlorophyll, and higher levels of tocopherols, flavonols and anthocyanins than the WT. Vascular- and root tip-specific PGI1 expression countered the reduced photosynthesis, low MEP pathway-derived CK content, dwarf phenotype and the metabolic characteristics of pgi1-2 plants, reverting them to WT-like levels. Moreover, pgl3-1, but not pfk4pfk5 plants phenocopied pgi1-2. Histochemical analyses of plants expressing GUS under the control of promoter regions of genes encoding plastidial OPPP enzymes exhibited strong GUS activity in root tips and vascular tissues. Overall, our findings show that root tip and vascular PGI1-mediated plastidial OPPP activity affects photosynthesis and growth through mechanisms involving long-distance modulation of the leaf proteome by MEP pathway-derived CKs.

Cytokinin and indole-3-acetic acid crosstalk is indispensable for silicon mediated chromium stress tolerance in roots of wheat seedlings.

The rising heavy metal contamination of soils imposes toxic impacts on plants as well as other life forms. One such highly toxic and carcinogenic heavy metal is hexavalent chromium [Cr(VI)] that has been reported to prominently retard the plant growth. The present study investigated the potential of silicon (Si, 10 µM) to alleviate the toxicity of Cr(VI) (25 µM) on roots of wheat (Triticum aestivum L.) seedlings. Application of Si to Cr(VI)-stressed wheat seedlings improved their overall growth parameters. This study also reveals the involvement of two phytohormones, namely auxin and cytokinin and their crosstalk in Si-mediated mitigation of the toxic impacts of Cr(VI) in wheat seedlings. The application of cytokinin alone to wheat seedlings under Cr(VI) stress reduced the intensity of toxic effects of Cr(VI). In combination with Si, cytokinin application to Cr(VI)-stressed wheat seedlings significantly minimized the decrease induced by Cr(VI) in different parameters such as root-shoot length (10.8% and 13%, respectively), root-shoot fresh mass (11.3% and 10.1%, respectively), and total chlorophyll and carotenoids content (13.4% and 6.8%, respectively) with respect to the control. This treatment also maintained the regulation of proline metabolism (proline content, and P5CS and PDH activities), ascorbate-glutathione (AsA-GSH) cycle and nutrient homeostasis. The protective effect of Si and cytokinin against Cr(VI) stress was minimized upon supplementation of an inhibitor of polar auxin transport- 2,3,5-triiodobenzoic acid (TIBA) which suggested a potential involvement of auxin in Si and cytokinin-mediated mitigation of Cr(VI) toxicity. The exogenous addition of a natural auxin - indole-3-acetic acid (IAA) confirmed auxin is an active member of a signaling cascade along with cytokinin that aids in Si-mediated Cr(VI) toxicity alleviation as IAA application reversed the negative impacts of TIBA on wheat roots treated with Cr(VI), cytokinin and Si. The results of this research are also confirmed by the gene expression analysis conducted for nutrient transporters (Lsi1, CCaMK, MHX, SULT1 and ZIP1) and enzymes involved in the AsA-GSH cycle (APX, GR, DHAR and MDHAR). The overall results of this research indicate towards possible induction of a crosstalk between cytokinin and IAA upon Si supplementation which in turn stimulates physiological, biochemical and molecular changes to exhibit protective effects against Cr(VI) stress. Further, the information obtained suggests probable employment of Si, cytokinin and IAA alone or combined in agriculture to maintain plant productivity under Cr(VI) stress and data regarding expression of key genes can be used to develop new crop varieties with enhanced resistance against Cr(VI) stress together with its reduced load in seedlings.

Two transcription factors, RhERF005 and RhCCCH12, regulate rose resistance to Botrytis cinerea by modulating cytokinin levels.

Gray mold caused by the necrotrophic fungal pathogen Botrytis cinerea is one of the most destructive diseases in rose (Rosa spp.). Rose infection by B. cinerea leads to severe economic losses due to necrosis, tissue collapse, and rot. In rose, cytokinins (CKs) positively regulate a defense response to B. cinerea, but little is known about the underlying molecular mechanisms. Here, we characterized two ethylene/jasmonic acid-regulated transcription factors, RhEFR005 and RhCCCH12, that bind to the promoter region of PATHOGENESIS-RELATED 10.1 (RhPR10.1) and promote its transcription, leading to decreased susceptibility to B. cinerea. The RhEFR005/RhCCCH12-RhPR10.1 module regulated cytokinin content in rose, and the susceptibility of RhEFR005-, RhCCCH12-, and RhPR10.1-silenced rose petals can be rescued by exogenous CK. In summary, our results reveal that the RhERF005/RhCCCH12-RhPR10.1 module regulates the CK-induced defense response of rose to B. cinerea.

MsMYB62-like as a negative regulator of anthocyanin biosynthesis in Malus spectabilis.

Crabapple is a valuable tree species in gardens due to its captivating array of flower and leaf colors, rendering it a favored choice in landscaping. The economic and ornamental values of Malus crabapple are closely associated with the biosynthesis of anthocyanin, a pigment responsible for its vibrant hues. The intricate regulation of anthocyanin biosynthesis involves the concerted activity of various genes. However, the specific mechanism governing this process in crabapple warrants in-depth exploration. In this study, we explored the inhibitory role of MsMYB62-like in anthocyanin biosynthesis. We identified MsDFR and MsANS as two downstream target genes of MsMYB62-like. These genes encode enzymes integral to the anthocyanin biosynthetic pathway. The findings demonstrate that MsMYB62-like directly binds to the promoters of MsDFR and MsANS, resulting in the downregulation of their expression levels. Additionally, our observations indicate that the plant hormone cytokinins exert a suppressive effect on the expression levels of MsMYB62-like, while concurrently upregulating MsDFR and MsANS. This study reveals that the MsMYB62-like-MsDFR/MsANS module plays an important role in governing anthocyanin levels in Malus crabapple. Notably, the regulatory interplay is modulated by the plant hormone cytokinins.

Unveiling terahertz wave stress effects and mechanisms in Pinellia ternata: Challenges, insights, and future directions.

This review aims to elucidate the intricate effects and mechanisms of terahertz (THz) wave stress on Pinellia ternata, providing valuable insights into plant responses. The primary objective is to highlight the imperative for future research dedicated to comprehending THz wave impacts across plant structures, with a specific focus on the molecular intricacies governing root system structure and function, from shoots to roots. Notably, this review highlights the accelerated plant growth induced by THz waves, especially in conjunction with other environmental stressors, and the subsequent alterations in cellular homeostasis, resulting in the generation of reactive oxygen species (ROS) and an increase in brassinosteroids. Brassinosteroids are explored for their dual role as toxic by-products of stress metabolism and vital signal transduction molecules in plant responses to abiotic stresses. The paper further investigates the spatio-temporal regulation and long-distance transport of phytohormones, including growth hormone, cytokinin, and abscisic acid (ABA), which significantly influence the growth and development of P. ternata under THz wave stress. With a comprehensive review of Reactive oxygen species (ROS) and Brassinosteroid Insensitive (BRI) homeostasis and signalling under THz wave stress, the article elucidates the current understanding of BRI involvement in stress perception, stress signalling, and domestication response regulation. Additionally, it underscores the importance of spatio-temporal regulation and long-distance transport of key plant hormones, such as growth hormone, cytokinin, and ABA, in determining root growth and development under THz wave stress. The study of how plants perceive and respond to environmental stresses holds fundamental biological significance, and enhancing plant stress tolerance is crucial for promoting sustainable agricultural practices and mitigating the environmental burdens associated with low-tolerance crop cultivation.

Structures and mechanisms of the Arabidopsis cytokinin transporter AZG1.

Cytokinins are essential for plant growth and development, and their tissue distributions are regulated by transmembrane transport. Recent studies have revealed that members of the 'Aza-Guanine Resistant' (AZG) protein family from Arabidopsis thaliana can mediate cytokinin uptake in roots. Here we present 2.7 to 3.3 Å cryo-electron microscopy structures of Arabidopsis AZG1 in the apo state and in complex with its substrates trans-zeatin (tZ), 6-benzyleaminopurine (6-BAP) or kinetin. AZG1 forms a homodimer and each subunit shares a similar topology and domain arrangement with the proteins of the nucleobase/ascorbate transporter (NAT) family. These structures, along with functional analyses, reveal the molecular basis for cytokinin recognition. Comparison of the AZG1 structures determined in inward-facing conformations and predicted by AlphaFold2 in the occluded conformation allowed us to propose that AZG1 may carry cytokinins across the membrane through an elevator mechanism.

Conserved CKI1-mediated signaling is required for female germline specification in Marchantia polymorpha.

In land plants, gametes derive from a small number of dedicated haploid cells.1 In angiosperms, one central cell and one egg cell are differentiated in the embryo sac as female gametes for double fertilization, while in non-flowering plants, only one egg cell is generated in the female sexual organ, called the archegonium.2,3 The central cell specification of Arabidopsis thaliana is controlled by the histidine kinase CYTOKININ-INDEPENDENT 1 (CKI1), which is a two-component signaling (TCS) activator sharing downstream regulatory components with the cytokinin signaling pathway.4,5,6,7 Our phylogenetic analysis suggested that CKI1 orthologs broadly exist in land plants. However, the role of CKI1 in non-flowering plants remains unclear. Here, we found that the sole CKI1 ortholog in the liverwort Marchantia polymorpha, MpCKI1, which functions through conserved downstream TCS components, regulates the female germline specification for egg cell development in the archegonium. In M. polymorpha, the archegonium develops three-dimensionally from a single cell accumulating MpBONOBO (MpBNB), a master regulator for germline initiation and differentiation.8 We visualized female germline specification by capturing the distribution pattern of MpBNB in discrete stages of early archegonium development, and found that MpBNB accumulation is restricted to female germline cells. MpCKI1 is required for the proper MpBNB accumulation in the female germline, and is critical for the asymmetric cell divisions that specify the female germline cells. These results suggest that CKI1-mediated TCS originated during early land plant evolution and participates in female germ cell specification in deeply diverged plant lineages.

Cytokinin: From autoclaved DNA to two-component signaling.

Since its first identification in the 1950s as a regulator of cell division, cytokinin has been linked to many physiological processes in plants, spanning growth and development and various responses to the environment. Studies from the last two and one-half decades have revealed the pathways underlying the biosynthesis and metabolism of cytokinin and have elucidated the mechanisms of its perception and signaling, which reflects an ancient signaling system evolved from two-component elements in bacteria. Mutants in the genes encoding elements involved in these processes have helped refine our understanding of cytokinin functions in plants. Further, recent advances have provided insight into the mechanisms of intracellular and long-distance cytokinin transport and the identification of several proteins that operate downstream of cytokinin signaling. Here, we review these processes through a historical lens, providing an overview of cytokinin metabolism, transport, signaling, and functions in higher plants.

Transcription factor RhRAP2.4L orchestrates cell proliferation and expansion to control petal size in rose.

Maintaining proper flower size is vital for plant reproduction and adaption to the environment. Petal size is determined by spatiotemporally regulated cell proliferation and expansion. However, the mechanisms underlying the orchestration of cell proliferation and expansion during petal growth remains elusive. Here, we determined that the transition from cell proliferation to expansion involves a series of distinct and overlapping processes during rose (Rosa hybrida) petal growth. Changes in cytokinin content were associated with the transition from cell proliferation to expansion during petal growth. RNA sequencing identified the AP2/ERF transcription factor gene RELATED TO AP2 4-LIKE (RhRAP2.4L), whose expression pattern positively associated with cytokinin levels during rose petal development. Silencing RhRAP2.4L promoted the transition from cell proliferation to expansion and decreased petal size. RhRAP2.4L regulates cell proliferation by directly repressing the expression of KIP RELATED PROTEIN 2 (RhKRP2), encoding a cell cycle inhibitor. In addition, we also identified BIG PETALub (RhBPEub) as another direct target gene of RhRAP2.4L. Silencing RhBPEub decreased cell size, leading to reduced petal size. Furthermore, the cytokinin signaling protein ARABIDOPSIS RESPONSE REGULATOR 14 (RhARR14) activated RhRAP2.4L expression to inhibit the transition from cell proliferation to expansion, thereby regulating petal size. Our results demonstrate that RhRAP2.4L performs dual functions in orchestrating cell proliferation and expansion during petal growth.

Type I MADS-box transcription factor TaMADS-GS regulates grain size by stabilizing cytokinin signalling during endosperm cellularization in wheat.

Grain size is one of the important traits in wheat breeding programs aimed at improving yield, and cytokinins, mainly involved in cell division, have a positive impact on grain size. Here, we identified a novel wheat gene TaMADS-GS encoding type I MADS-box transcription factor, which regulates the cytokinins signalling pathway during early stages of grain development to modulate grain size and weight in wheat. TaMADS-GS is exclusively expressed in grains at early stage of seed development and its knockout leads to delayed endosperm cellularization, smaller grain size and lower grain weight. TaMADS-GS protein interacts with the Polycomb Repressive Complex 2 (PRC2) and leads to repression of genes encoding cytokinin oxidase/dehydrogenases (CKXs) stimulating cytokinins inactivation by mediating accumulation of the histone H3 trimethylation at lysine 27 (H3K27me3). Through the screening of a large wheat germplasm collection, an elite allele of the TaMADS-GS exhibits higher ability to repress expression of genes inactivating cytokinins and a positive correlation with grain size and weight, thus representing a novel marker for breeding programs in wheat. Overall, these findings support the relevance of TaMADS-GS as a key regulator of wheat grain size and weight.