Somatic Embryogenesis Receptor-like Kinase Research Articles

Identifying receptor-like kinases that enable Caulobacter RHG1 to promote plant growth in Arabidopsis thaliana

ABSTRACT Plants express an array of receptor-like kinases (RLKs) to control development and communicate with their environment. Many RLKs are uncharacterized and some of them are expected to regulate plant responses to plant growth-promoting rhizobacteria (PGPR). Despite documented effects induced by Caulobacter RHG1, the underlying signaling pathways and the involved RLKs remain uncharted. Through a targeted RLK mutant screening, we aimed to decipher the receptors that steer the Caulobacter RHG1-induced growth promotion in Arabidopsis thaliana. We identified four RLKs that are pivotal in the RHG1-Arabidopsis interaction, including the coreceptors SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE 1 (SERK1) and BRASSINOSTEROID INSENSITIVE 1-ASSOCIATED KINASE 1 (BAK1/SERK3), which act redundantly in the RHG1-Arabidopsis interaction, possibly by interplaying with the unknown RLK AT3G28040 and the immunity-related ELONGATION FACTOR-TU RECEPTOR (EFR). These results shed new light on the molecular dynamics orchestrating plant responses to PGPR, and concurrently contribute a crucial piece to the intricate puzzle of RLK interactions. Key policy highlights Four RLKs; BAK1, SERK1, EFR, and AT3G28040 (RRHG) are involved in the RHG1-Arabidopsis interaction. BAK1 and SERK1, two well-described co-receptors, act redundantly and play a pivotal role in the RHG1-driven growth promotion, possibly by interplaying with the unknown RLK AT3G28040 and the immunity-related RLK EFR. Most known development – and immunity-related RLKs barely influence RHG1-driven plant growth promotion in Arabidopsis.

Tilletia horrida glycoside hydrolase family 128 protein, designated ThGhd_7, modulates plant immunity by blocking reactive oxygen species production.

Tilletia horrida is an important soilborne fungal pathogen that causes rice kernel smut worldwide. We found a glycoside hydrolase family 128 protein, designated ThGhd_7, caused cell death in Nicotiana benthamiana leaves. The predicted signal peptide (SP) of ThGhd_7 targets it for secretion. However, loss of the SP did not affect its ability to induce cell death. The 23-201 amino acid sequence of ThGhd_7 was sufficient to trigger cell death in N. benthamiana. ThGhd_7 expression was induced and upregulated during T. horrida infection. ThGhd_7 localised to both the cytoplasm and nucleus of plant cells, and nuclear localisation was required to induce cell death. The ability of ThGhd_7 to trigger cell death in N. benthamiana depends on RAR1 (required for Mla12 resistance), SGT1 (suppressor of G2 allele of Skp1), and BAK1/SERK3 (somatic embryogenesis receptor-like kinase 3). Heterologous overexpression of ThGhd_7 in rice reduced reactive oxygen species (ROS) production and enhanced susceptibility to T. horrida. Further research revealed that ThGhd_7 interacted with and destabilised OsSGT1, which is required for ROS production and is a positive regulator of rice resistance to T. horrida. Taken together, these findings suggest that T. horrida employs ThGhd_7 to disrupt ROS production and thereby promote infection.

DsRNA-induced immunity targets plasmodesmata and is suppressed by viral movement proteins.

Emerging evidence indicates that in addition to its well-recognized functions in antiviral RNA silencing, dsRNA elicits pattern-triggered immunity (PTI), likely contributing to plant resistance against virus infections. However, compared to bacterial and fungal elicitor-mediated PTI, the mode-of-action and signaling pathway of dsRNA-induced defense remain poorly characterized. Here, using multicolor in vivo imaging, analysis of GFP mobility, callose staining, and plasmodesmal marker lines in Arabidopsis thaliana and Nicotiana benthamiana, we show that dsRNA-induced PTI restricts the progression of virus infection by triggering callose deposition at plasmodesmata, thereby likely limiting the macromolecular transport through these cell-to-cell communication channels. The plasma membrane-resident SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE 1, the BOTRYTIS INDUCED KINASE1/AVRPPHB SUSCEPTIBLE1-LIKE KINASE1 kinase module, PLASMODESMATA-LOCATED PROTEINs 1/2/3, as well as CALMODULIN-LIKE 41 and Ca2+ signals are involved in the dsRNA-induced signaling leading to callose deposition at plasmodesmata and antiviral defense. Unlike the classical bacterial elicitor flagellin, dsRNA does not trigger a detectable reactive oxygen species (ROS) burst, substantiating the idea that different microbial patterns trigger partially shared immune signaling frameworks with distinct features. Likely as a counter strategy, viral movement proteins from different viruses suppress the dsRNA-induced host response leading to callose deposition to achieve infection. Thus, our data support a model in which plant immune signaling constrains virus movement by inducing callose deposition at plasmodesmata and reveals how viruses counteract this layer of immunity.

PXL1 and SERKs act as receptor–coreceptor complexes for the CLE19 peptide to regulate pollen development

Gametophyte development in angiosperms occurs within diploid sporophytic structures and requires coordinated development; e.g., development of the male gametophyte pollen depends on the surrounding sporophytic tissue, the tapetum. The mechanisms underlying this interaction remain poorly characterized. The peptide CLAVATA3/EMBRYO SURROUNDING REGION-RELATED 19 (CLE19) plays a “braking” role in preventing the harmful overexpression of tapetum transcriptional regulators to ensure normal pollen development in Arabidopsis. However, the CLE19 receptor is unknown. Here, we show that CLE19 interacts directly with the PXY-LIKE1 (PXL1) ectodomain and induces PXL1 phosphorylation. PXL1 is also required for the function of CLE19 in maintaining the tapetal transcriptional regulation of pollen exine genes. Additionally, CLE19 induces the interactions of PXL1 with SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE (SERK) coreceptors required for pollen development. We propose that PXL1 and SERKs act as receptor and coreceptor, respectively, of the extracellular CLE19 signal, thereby regulating tapetum gene expression and pollen development.

Isolation and Expression of Transcription Factors Involved in Somatic Embryo Development by Transcriptome Analysis of Embryogenic Callus of Thuja koraiensis

Thuja koraiensis Nakai (Cupressaceae) is an endangered and ecologically important conifer endemic to Korea. Previously, we established a protocol for micropropagation in T. koraiensis, which involved somatic embryogenesis from embryogenic callus of T. koraiensis. However, the molecular mechanisms underlying somatic embryogenesis remain unclear. Herein, we performed transcriptomic analysis to identify somatic embryogenesis-related genes of T. koraiensis via Illumina RNA sequencing. We conducted de novo transcriptome assembly using a Trinity assembler, which produced 274,077 transcript contigs clustered into 205,843 transcripts (unigenes), with an average length of 825 base pairs. Of all the unigenes, 14.69%, 18.62%, and 7.4% had homologs in the Gene Ontology, NCBI Non-redundant Protein, and NCBI Nucleotide databases, respectively. Among these mRNA sequences, expression of putative embryogenesis-associated transcription factors, namely BABYBOOM (BBM), WUSCHEL-RELATED HOMEOBOX (WOX), and SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE (SERK), was analyzed during somatic embryogenesis. RT-PCR analysis revealed that TkBBM, TkWOX, and TkSERK were highly expressed in embryogenic callus and seedling roots, whereas their expression was reduced in seedling leaves. Our findings provide new insights into the roles of BBM, WOX, and SERK in somatic embryogenesis. Our results may serve as a reference for comparative transcriptome analysis in related species and further aid functional genomics studies in conifers.

The EPFL-ERf-SERK signaling controls integument development in Arabidopsis.

As the seed precursor, the ovule produces the female gametophyte (or embryo sac), and the subsequent double fertilization occurs in it. The integuments emerge sequentially from the integument primordia at the early stages of ovule development and finally enwrap the embryo sac gradually during gametogenesis, protecting and nursing the embryo sac. However, the mechanisms regulating integument development are still obscure. In this study, we show that SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASES (SERKs) play essential roles during integument development in Arabidopsis thaliana. The serk1/2/3 triple mutant shows arrested integuments and abnormal embryo sacs, similar defects also found in the triple loss-of-function mutants of ERECTA family (ERf) genes. Ovules of serk1/2/3 er erl1/2 show defects similar to er erl1/2 and serk1/2/3. Results of yeast two-hybrid analyses, bimolecular fluorescence complementation (BiFC) analyses, and co-immunoprecipitation assays demonstrated that SERKs interact with ERf, which depends on EPIDERMAL PATTERNING FACTOR-LIKE (EPFL) family small peptides. The sextuple mutant epfl1/2/3/4/5/6 shows integument defects similar to both of er erl1/2 and serk1/2/3. Our results demonstrate that ERf-SERK-mediated EPFL signaling orchestrates the development of the female gametophyte and the surrounding sporophytic integuments.

Morphophysiological and transcriptome analysis reveal that reprogramming of metabolism, phytohormones and root development pathways governs the potassium (K+) deficiency response in two contrasting chickpea cultivars.

Potassium (K+) is an essential macronutrient for plant growth and development. K+ deficiency hampers important plant processes, such as enzyme activation, protein synthesis, photosynthesis and stomata movement. Molecular mechanism of K+ deficiency tolerance has been partly understood in model plants Arabidopsis, but its knowledge in legume crop chickpea is missing. Here, morphophysiological analysis revealed that among five high yielding desi chickpea cultivars, PUSA362 shows stunted plant growth, reduced primary root growth and low K+ content under K+ deficiency. In contrast, PUSA372 had negligible effect on these parameters suggesting that PUSA362 is K+ deficiency sensitive and PUSA372 is a K+ deficiency tolerant chickpea cultivar. RNA-seq based transcriptome analysis under K+ deficiency revealed a total of 820 differential expressed genes (DEG's) in PUSA362 and 682 DEGs in PUSA372. These DEGs belongs to different functional categories, such as plant metabolism, signal transduction components, transcription factors, ion/nutrient transporters, phytohormone biosynthesis and signalling, and root growth and development. RNA-seq expression of randomly selected 16 DEGs was validated by RT-qPCR. Out of 16 genes, 13 showed expression pattern similar to RNA-seq expression, that verified the RNA-seq expression data. Total 258 and 159 genes were exclusively up-regulated, and 386 and 347 genes were down-regulated, respectively in PUSA362 and PUSA372. 14 DEGs showed contrasting expression pattern as they were up-regulated in PUSA362 and down-regulated in PUSA372. These include somatic embryogenesis receptor-like kinase 1, thaumatin-like protein, ferric reduction oxidase 2 and transcription factor bHLH93. Nine genes which were down-regulated in PUSA362 found to be up-regulated in PUSA372, including glutathione S-transferase like, putative calmodulin-like 19, high affinity nitrate transporter 2.4 and ERF17-like protein. Some important carbohydrate metabolism related genes, like fructose-1,6-bisphosphatase and sucrose synthase, and root growth related Expansin gene were exclusively down-regulated, while an ethylene biosynthesis gene 1-aminocyclopropane-1-carboxylate oxidase 1 (ACO1) was up-regulated in PUSA362. Interplay of these and several other genes related to hormones (auxin, cytokinin, GA etc.), signal transduction components (like CBLs and CIPKs), ion transporters and transcription factors might underlie the contrasting response of two chickpea cultivars to K+ deficiency. In future, some of these key genes will be utilized in genetic engineering and breeding programs for developing chickpea cultivars with improved K+ use efficiency (KUE) and K+ deficiency tolerance traits.

Functional Analyses of a Small Secreted Cysteine-Rich Protein ThSCSP_14 in Tilletia horrida

Tilletia horrida is a biotrophic basidiomycete fungus that causes rice kernel smut, one of the most significant diseases in hybrid rice-growing areas worldwide. Little is known about the pathogenic mechanisms and functions of effectors in T. horrida. Here, we performed functional studies of the effectors in T. horrida and found that, of six putative effectors tested, only ThSCSP_14 caused the cell death phenotype in epidermal cells of Nicotiana benthamiana leaves. ThSCSP_14 was upregulated early on during the infection process, and the encoded protein was secreted. The predicted signal peptide (SP) of ThSCSP_14 was required for its ability to induce the necrosis phenotype. Furthermore, the ability of ThSCSP_14 to trigger cell death in N. benthamiana depended on suppressing the G2 allele of Skp1 (SGT1), required for Mla12 resistance (RAR1), heat-shock protein 90 (HSP90), and somatic embryogenesis receptor-like kinase (SERK3). It is important to note that ThSCSP_14 induced a plant defense response in N. benthamiana leaves. Hence, these results demonstrate that ThSCSP_14 is a possible effector that plays an essential role in T. horrida-host interactions.

ThSCSP_12: Novel Effector in Tilletia horrida That Induces Cell Death and Defense Responses in Non-Host Plants

The basidiomycete fungus Tilletia horrida causes rice kernel smut (RKS), a crucial disease afflicting hybrid-rice-growing areas worldwide, which results in significant economic losses. However, few studies have investigated the pathogenic mechanisms and functions of effectors in T. horrida. In this study, we found that the candidate effector ThSCSP_12 caused cell necrosis in the leaves of Nicotiana benthamiana. The predicted signal peptide (SP) of this protein has a secreting function, which is required for ThSCSP_12 to induce cell death. The 1- 189 amino acid (aa) sequences of ThSCSP_12 are sufficient to confer it the ability to trigger cell death in N. benthamiana. The expression of ThSCSP_12 was induced and up-regulated during T. horrida infection. In addition, we also found that ThSCSP_12 localized in both the cytoplasm and nucleus of plant cells and that nuclear localization of this protein is required to induce cell death. Furthermore, the ability of ThSCSP_12 to trigger cell death in N. benthamiana depends on the (RAR1) protein required for Mla12 resistance but not on the suppressor of the G2 allele of Skp1 (SGT1), heat shock protein 90 (HSP90), or somatic embryogenesis receptor-like kinase (SERK3). Crucially, however, ThSCSP_12 induced a defense response in N. benthamiana leaves; yet, the expression of multiple defense-related genes was suppressed in response to heterologous expression in host plants. To sum up, these results strongly suggest that ThSCSP_12 operates as an effector in T. horrida-host interactions.

Functional Analysis of Glucose-6-Phosphate Translocator PoGPT1 in Embryogenic Callus Formation of Tree Peony

Tree peony (Paeonia ostii) is an ornamental flowering plant that is generally recalcitrant to establishment of a mature somatic embryo regeneration system in vitro. Glucose-6-phosphate translocator (GPT) plays an important regulatory role in embryogenesis of plants. In this study, PoGPT1 was cloned, and a bioinformatic analysis and functional verification of the gene were performed. The results showed that PoGPT1 encoded a polypeptide of 392 amino acids, which was a basic non-secreted hydrophobic transmembrane protein, and was mainly localized in the plastids. PoGPT1 was highly expressed in tree peony leaves, and its transcript abundance increased with the progression of zygotic embryo development. Overexpression of PoGPT1 caused up-regulation of leafy cotyledon 1 (PoLEC1), somatic embryogenesis receptor-like kinase (PoSERK), and agamous-like15 (PoAGL15) in tree peony callus. In addition, PoGPT1 overexpression promoted the increase in indole-3-acetic acid (IAA), 5-deoxystrigol (5DS), and brassinolide (BL) contents, especially of IAA, but reduced the contents of abscisic acid (ABA), 6-benzyladenosine (BARP), and 1-aminocyclopropanecarboxylic acid (ACC). The present research showed that PoGPT1 synergistically regulated the contents of endogenous hormones and expression levels of embryogenesis-related genes to promote the embryonic development of tree peony. The results provide theoretical and technical support for the establishment of a tree peony embryogenic callus formation and subsequent research on somatic embryogenesis.

Phytocytokine signalling reopens stomata in plant immunity and water loss.

Stomata exert considerable effects on global carbon and water cycles by mediating gas exchange and water vapour1,2. Stomatal closure prevents water loss in response to dehydration and limits pathogen entry3,4. However, prolonged stomatal closure reduces photosynthesis and transpiration and creates aqueous apoplasts that promote colonization by pathogens. How plants dynamically regulate stomatal reopening in a changing climate is unclear. Here we show that the secreted peptides SMALL PHYTOCYTOKINES REGULATING DEFENSE AND WATER LOSS (SCREWs) and the cognate receptor kinase PLANT SCREW UNRESPONSIVE RECEPTOR (NUT) counter-regulate phytohormone abscisic acid (ABA)- and microbe-associated molecular pattern (MAMP)-induced stomatal closure. SCREWs sensed by NUT function as immunomodulatory phytocytokines and recruit SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE (SERK) co-receptors to relay immune signalling. SCREWs trigger the NUT-dependent phosphorylation of ABA INSENSITIVE 1 (ABI1) and ABI2, which leads to an increase in the activity of ABI phosphatases towards OPEN STOMATA 1 (OST1)-a key kinase that mediates ABA- and MAMP-induced stomatal closure5,6-and a reduction in the activity of S-type anion channels. After induction by dehydration and pathogen infection, SCREW-NUT signalling promotes apoplastic water loss and disrupts microorganism-rich aqueous habitats to limit pathogen colonization. The SCREW-NUT system is widely distributed across land plants, which suggests that it has an important role in preventing uncontrolled stomatal closure caused by abiotic and biotic stresses to optimize plant fitness.

Cloning and functional identification of setaria italica somatic embryogenesis receptor-like kinase1 gene (SiSERK1).

Plant somatic embryogenesis receptor-like kinases (SERK), members of leucine-rich repeat receptor-like kinases (LRR-RLKs) subfamily, are widely involved in plant growth, development and innate immunity. In this study, the setaria italica somatic embryogenesis receptor-like kinase1 gene (SiSERK1) was cloned by gateway technology, and transferred into a brasssinosteroid (BR) receptor mutant of Arabidopsis thaliana WS2 (bri1-5). After BL treatment, the transgenic plants could partially restore the phenotype of bri1-5. After Pst DC3000 treatment, the CFU value of SiSERK1 overexpression plant pathogen was between WS2 and bri1-5. Stomatal opening and plant height were also between them. Therefore, it is speculated that SiSERK1 gene is involved in BR signaling pathway and can improve the resistance of bri1-5 to Pst DC3000 through SA and NHP mediated systemic acquired resistance (SAR).

Essential roles of SERKs in the ROOT MERISTEM GROWTH FACTOR-mediated signaling pathway.

ROOT MERISTEM GROWTH FACTORs (RGFs), a group of peptide hormones, play key roles in root apical meristem development. In Arabidopsis (Arabidopsis thaliana), there are 11 members of RGFs, in which at least RGF1, RGF2, and RGF3 are expressed at the root tip and are involved in root stem cell niche maintenance. RGFs are perceived by five functionally redundant receptor-like protein kinases, RGF1 INSENSITIVE 1 (RGI1) to RGI5, to maintain the expression of two downstream APETALA 2 (AP2) transcription factor genes, PLETHORA 1 (PLT1) and PLT2, and to stabilize PLT2. RGI1 to RGI3 were also named RGF RECEPTOR 1 (RGFR1) to RGFR3, respectively. Although previous studies have suggested that BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1) and its paralogs, SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASEs (SERKs), may act as coreceptors of RGIs, comprehensive genetic and biochemical analyses have not been well documented. Here, we report that single, double, and triple mutants of SERKs show various degrees of short root phenotypes and insensitivity to exogenously applied RGF1. The interaction between RGIs and BAK1 and their mutual phosphorylation are RGF1 dependent. We also found that RGF1-induced MAPK activation relies on both RGIs and SERKs. We demonstrate that RGIs play redundant roles in regulating root apical meristem development. Therefore, we genetically and biochemically substantiated that SERKs, as coreceptors, play essential roles in the RGF1-mediated signaling pathway.

Expression analysis of OsSERK, OsLEC1 and OsWOX4 genes in rice (Oryza sativa L.) callus during somatic embryo development

Somatic embryogenesis is an asexual reproduction process that occurs in many plant species, including rice. This process contains several totipotency markers such as Somatic Embryogenesis Receptor-like Kinase (SERK), Leafy Cotyledon1 (LEC1) and WUSCHEL-Related Homeobox4 (WOX4) and also a helpful model for embryo development and clones and transformations. Here, we report the gene expression during somatic embryo development correlates with regeneration frequency in 14 Javanica rice (pigmented and non-pigmented) using modifified N6 media supplemented with Kinetin (2.0 mg/L) and NAA (1.0 mg/L). Although there have been advances in understanding the genetic basis of somatic embryogenesis in other varieties, rice is still unexplored, especially during somatic embryo development. Moreover, for the formation of callus induction from immature embryos, 2,4-D (2.0 mg/L, 3.0 mg/L) was used. This study analysed the gene expression of OsSERK, OsWOX4 and OsLEC1 genes through RT-PCR analysis. Higher expression of the OsLEC1 gene indicates that their function may correlate in the in vitro with the high response of rice after transfer to regeneration media. This study found that rice varieties of pigmented rice (MS Pendek and Gogoniti II) and non-pigmented rice (Pandan Ungu) showed high regeneration frequency, showing higher OsLEC1 expression than other varieties because OsLEC1 promotes the maturation of somatic embryos in plant regeneration on day 14. However, the contrast with Genjah nganjuk may be effective because of other regulatory genes. RT-PCR analysis showed OsSERK had less expression level than OsLEC1 and OsWOX4 in the varieties, which correlate with the percentage of plant regeneration, but not for Gogoniti II. In conclusion, the higher percentage of plant regeneration correlates with the higher expression level of OsLEC1 at day 14 of media regeneration of rice.

Synthetic Strigolactone GR24 Improves Arabidopsis Somatic Embryogenesis through Changes in Auxin Responses.

Somatic embryogenesis in Arabidopsis encompasses an induction phase requiring auxin as the inductive signal to promote cellular dedifferentiation and formation of the embryogenic tissue, and a developmental phase favoring the maturation of the embryos. Strigolactones (SLs) have been categorized as a novel group of plant hormones based on their ability to affect physiological phenomena in plants. The study analyzed the effects of synthetic strigolactone GR24, applied during the induction phase, on auxin response and formation of somatic embryos. The expression level of two SL biosynthetic genes, MORE AXILLARY GROWTH 3 and 4 (MAX3 and MAX4), which are responsible for the conversion of carotene to carotenal, increased during the induction phase of embryogenesis. Arabidopsis mutant studies indicated that the somatic embryo number was inhibited in max3 and max4 mutants, and this effect was reversed by applications of GR24, a synthetic strigolactone, and exacerbated by TIS108, a SL biosynthetic inhibitor. The transcriptional studies revealed that the regulation of GR24 and TIS108 on somatic embryogenesis correlated with changes in expression of AUXIN RESPONSIVE FACTORs 5, 8, 10, and 16, known to be required for the production of the embryogenic tissue, as well as the expression of WUSCHEL (WUS) and Somatic Embryogenesis Receptor-like Kinase 1 (SERK1), which are markers of cell dedifferentiation and embryogenic tissue formation. Collectively, this work demonstrated the novel role of SL in enhancing the embryogenic process in Arabidopsis and its requirement for inducing the expression of genes related to auxin signaling and production of embryogenic tissue.

SERKs regulate embryonic cuticle integrity through the TWS1-GSO1/2 signaling pathway in Arabidopsis.

The embryonic cuticle integrity is critical for the embryo to separate from the neighboring endosperm. The sulfated TWISTED SEED1 (TWS1) peptide precursor generated in the embryo diffuses through gaps of the nascent cuticle to the surrounding endosperm, where it is cleaved by ABNORMAL LEAF SHAPE1 (ALE1) and becomes an active mature form. The active TWS1 is perceived by receptor-like protein kinases GASSHO1 (GSO1) and GSO2 in the embryonic epidermal cells to start the downstream signaling and guide the formation of an intact embryonic cuticle. However, the early signaling events after TWS1 is perceived by GSO1/2 are still unknown. Here, we report that serk1/2/3 embryos show cuticle defects similar to ale1, tws1, and gso1/2. Genetic and biochemical analyses were performed to dissect the signaling pathway mediated by SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASEs (SERKs) during cuticle development. SERKs function with GSO1/2 in a common pathway to monitor the integrity of the embryonic cuticle. SERKs interact with GSO1/2, which can be enhanced dramatically by TWS1. The phosphorylation levels of SERKs and GSO1/2 rely on each other and can respond to and be elevated by TWS1. Our results demonstrate that SERKs may function as coreceptors of GSO1/2 to transduce the TWS1 signal and ultimately regulate embryonic cuticle integrity.

IDA (INFLORESCENCE DEFICIENT IN ABSCISSION)-like peptides and HAE (HAESA)-like receptors regulate corolla abscission in Nicotiana benthamiana flowers

BackgroundAbscission is an active, organized, and highly coordinated cell separation process enabling the detachment of aerial organs through the modification of cell-to-cell adhesion and breakdown of cell walls at specific sites on the plant body known as abscission zones. In Arabidopsis thaliana, abscission of floral organs and cauline leaves is regulated by the interaction of the hormonal peptide INFLORESCENCE DEFICIENT IN ABSCISSION (IDA), a pair of redundant receptor-like protein kinases, HAESA (HAE) and HAESA-LIKE2 (HSL2), and SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE (SERK) co-receptors. However, the functionality of this abscission signaling module has not yet been demonstrated in other plant species.ResultsThe expression of the pair of NbenIDA1 homeologs and the receptor NbenHAE.1 was supressed at the base of the corolla tube by the inoculation of two virus-induced gene silencing (VIGS) constructs in Nicotiana benthamiana. These gene suppression events arrested corolla abscission but did not produce any obvious effect on plant growth. VIGS plants retained a higher number of corollas attached to the flowers than control plants, an observation related to a greater corolla breakstrength. The arrest of corolla abscission was associated with the preservation of the parenchyma tissue at the base of the corolla tube that, in contrast, was virtually collapsed in normal corollas. In contrast, the inoculation of a viral vector construct that increased the expression of NbenIDA1A at the base of the corolla tube negatively affected the growth of the inoculated plants accelerating the timing of both corolla senescence and abscission. However, the heterologous ectopic overexpression of citrus CitIDA3 and Arabidopsis AtIDA in N. benthamiana did not alter the standard plant phenotype suggesting that the proteolytic processing machinery was unable to yield active peptides.ConclusionHere, we demonstrate that the pair of NbenIDA1 homeologs encoding small peptides of the IDA-like family and the receptor NbenHAE.1 control cellular breakdown at the base of the corolla tube awhere an adventitious AZ should be formed and, therefore, corolla abscission in N. benthamiana flowers. Altogether, our results provide the first evidence supporting the notion that the IDA-HAE/HSL2 signaling module is conserved in angiosperms.

A robust workflow for indirect somatic embryogenesis and cormlet production in saffron (Crocus sativus L.) and its wild allies; C. caspius and C. speciosus

Saffron (Crocus sativus L.) and its wild relatives, Crocus caspius and Crocus speciosus are of considerable significance in the pharmaceutical, nutraceutical, and ornamental bulbs industry. Towards the ultimate goal of the conservation of wild Crocus species and establishment of an efficient workflow for in vitro production of Crocuses, efficient protocols were developed for disinfection and in vitro production of cormlets in C. sativus and its wild allies C. caspius and C. speciosus. Moreover, the differential expression of the Somatic Embryogenesis Receptor-like Kinase (SERK) gene was evaluated as a potential molecular marker during embryogenesis between embryogenic and non-embryogenic calli. A highly efficient disinfection recipe and a low-cost TDZ-free protocol have been successfully developed for in vitro cormlet production in three Crocus species. MS medium containing 10.18 μM 2, 4-D + 4.44 μM BAP was most efficiently induced callus and somatic embryo formation. The highest conversion frequency and maximum cormlet weight were achieved in MS containing 5.37 μM NAA + 8.88 μM BAP. The SERK expression was significantly much higher in embryogenic calli than non-embryogenic in all Crocus species. The current low-cost and easy-to-use recipe suggests a promising in vitro propagation workflow for mass production of uniform pathogen-free cormlets of Crocus species, as well as a platform to better conservation of wild Crocus species and effective gene and genome editing using CRISPR-Cas9 in future studies.

Comparative transcriptome analysis of gynoecious and monoecious inflorescences reveals regulators involved in male flower development in the woody perennial plant Jatropha curcas.

ABCE model genes along with genes related to GA biosynthesis and auxin signalling may play significant roles in male flower development in Jatropha curcas. Flowering plants exhibit extreme reproductive diversity. Jatropha curcas, a woody plant that is promising for biofuel production, is monoecious. Here, two gynoecious Jatropha mutants (bearing only female flowers) were used to identify key genes involved in male flower development. Using comparative transcriptome analysis, we identified 17 differentially expressed genes (DEGs) involved in floral organ development between monoecious plants and the two gynoecious mutants. Among these DEGs, five floral organ identity genes, Jatropha AGAMOUS, PISTILLATA, SEPALLATA 2-1 (JcSEP2-1), JcSEP2-2, and JcSEP3, were downregulated in ch mutant inflorescences; two gibberellin (GA) biosynthesis genes, Jatropha GA REQUIRING 1 and GIBBERELLIN 3-OXIDASE 1, were downregulated in both the ch and g mutants; and two genes involved in the auxin signalling pathway, Jatropha NGATHA1 and STYLISH1, were downregulated in the ch mutant. Furthermore, four hub genes involved in male flower development, namely Jatropha SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE 1, CRYPTOCHROME 2, SUPPRESSOR OF OVEREXPRESSION OF CO 1 and JAGGED, were identified using weighted gene correlation network analysis. These results suggest that floral organ identity genes and genes involved in GA biosynthesis and auxin signalling may participate in male flower development in Jatropha. This study will contribute to understanding sex differentiation in woody perennial plants.

SERK genes identification and expression analysis during somatic embryogenesis and sporogenesis of sexual and apomictic Brachiaria brizantha (Syn. Urochloa brizantha).

In Brachiaria brizantha BbrizSERK1, BbrizSERK2 and BbrizSERK3 were identified. SERK expression marks somatic embryogenesis, sexual MMC, and sexual and apomictic PMC. BbrizSERK3 might have a regulatory role in reproductive development. Somatic embryogenesis receptor-like kinase (SERK) consists of plasma membrane receptor genes that have been characterized in various species, associated with several aspects of plant development, including reproduction. SERK genes are involved in anther development and in early embryo development in sexual and asexual seed formation. To comprehend the complexity of the SERK genes and their function in Brachiaria reproduction, we performed a homology-based search in a genomic database of a sexual B. brizantha and identified sequences of three SERK genes, BbrizSERK1, BbrizSERK2, and BbrizSERK3. RNASeq data showed equivalent abundance of BbrizSERK1 and BbrizSERK2 transcripts in ovaries at early megasporogenesis of sexuals and apomicts, while BbrizSERK3 transcripts were more abundant in ovaries of sexuals than in apomicts. BbrizSERK3 results in three coding sequences due to alternative splicing, among them Variant 1 results in a protein with all the predicted domains of a SERK. BbrizSERK transcripts were detected in male reproductive tissues of both sexual and apomictic plants, suggesting a role in controlling anther development. BbrizSERK transcripts were detected early in ovule development, in the integuments, and in the megaspore mother cell of the sexual plant, butnot in the cells that give rise to apomictic embryo sacs, suggesting a role in female reproductive development of sexuals. This paper provides evidences that SERK genes plays a role in the onset and establishment of somatic embryogenesis and in the reproductive development of B. brizantha and suggests a distinct role of BbrizSERK in apomixis initiation.