Fasciclin-like Arabinogalactan Proteins Research Articles

Functional study of the soybean stamen-preferentially expressed gene GmFLA22a in regulating male fertility.

China has a high dependence on soybean imports, yield increase at a faster rate is an urgent problem that need to be solved at present. The application of heterosis is one of the effective ways to significantly increase crop yield. In recent years, the development of an intelligent male sterility system based on recessive nuclear sterile genes has provided a potential solution for rapidly harnessing the heterosis in soybean. However, research on male sterility genes in soybean has been lagged behind. Based on transcriptome data of soybean floral organs in our research group, a soybean stamen-preferentially expressed gene GmFLA22a was identified. It encodes a fasciclin-like arabinogalactan protein with the FAS1 domain, and subcellular localization studies revealed that it may play roles in the endoplasmic reticulum. Take advantage of the gene editing technology, the Gmfla22a mutant was generated in this study. However, there was a significant reduction in the seed-setting rate in the mutant plants at the reproductive growth stage. The pollen viability and germination rate of Gmfla22a mutant plants showed no apparent abnormalities. Histological staining demonstrated that the release of pollen grains in the mutant plants was delayed and incomplete, which may due to the locule wall thickening in the anther development. This could be the reason of the reduced seed-setting rate in Gmfla22a mutants. In summary, our study has preliminarily revealed that GmFLA22a may be involved in regulating soybean male fertility. It provides crucial genetic materials for further uncovering its molecular function and gene resources and theoretical basis for the utilization of heterosis in soybean.

Functional analysis of fasciclin-like arabinogalactan in carotenoid synthesis during tomato fruit ripening

Carotenoids are important pigmented nutrients synthesized by tomato fruits during ripening. To reveal the molecular mechanism underlying carotenoid synthesis during tomato fruit ripening, we analyzed carotenoid metabolites and transcriptomes in six development stages of tomato fruits. A total of thirty different carotenoids were detected and quantified in tomato fruits from 10 to 60 DPA. Based on differential gene expression profiles and WGCNA, we explored several genes that were highly significant and negatively correlated with lycopene, all of which encode fasciclin-like arabinogalactan proteins (FLAs). The FLAs are involved in plant signal transduction, however the functional role of these proteins has not been studied in tomato. Genome-wide analysis revealed that cultivated and wild tomato species contained 18 to 22 FLA family members, clustered into four groups, and mainly evolved by means of segmental duplication. The functional characterization of FLAs showed that silencing of SlFLA1, 5, and 13 were found to contribute to the early coloration of tomato fruits, and the expression of carotenoid synthesis-related genes was up-regulated in fruits that changed phenotypically, especially in SlFLA13-silenced plants. Furthermore, the content of multiple carotenoids (including (E/Z)-phytoene, lycopene, γ-carotene, and α-carotene) was significantly increased in SlFLA13-silenced fruits, suggesting that SlFLA13 has a potential inhibitory function in regulating carotenoid synthesis in tomato fruits. The results of the present study broaden the idea of analyzing the biological functions of tomato FLAs and preliminary evidence for the inhibitory role of SlFLA13 in carotenoid synthesis in fruit, providing the theoretical basis and a candidate for improving tomato fruit quality.

The fasciclin-like arabinogalactan protein FLA11 of Ostrya rehderiana impacts wood formation and salt stress in Populus

Wood is an abundant source of biomass and an important biomaterial for renewable resources and industrial products worldwide. Fasciclin-like arabinogalactan proteins (FLAs) are essential for plant growth, development, and responses to various abiotic stresses and are also closely associated with wood formation. However, the biological roles of FLAs in trees remain largely unclear. Here, we identified the FLA family in the ironwood tree (Ostrya rehderiana) and elucidated its possible function in poplar. Our results showed that OreFLAs underwent significant gene expansion due to multiple duplication events, leading to apparent functional redundancy. Among these genes, OreFLA11 was highly expressed in the xylem and roots. Overexpression of OreFLA11 in Populus alba var. pyramidalis altered xylem structure and cell wall composition, such as thickening the xylem and secondary cell wall (SCW), decreasing lignin contents and increasing cellulose contents, resulting in increased stem tensile strength but decreased salt tolerance, suggesting that OreFLA11 plays a negative role in the salt stress response. The mutant Palfla11 generated using CRIPSR-Cas9 methods showed the opposite phenotype compared to that of OreFLA11-OE poplars, indicating that the orthologues function similarly in wood formation. Furthermore, the transcriptome analysis showed that genes such as PME, BGAL and BGLU, related to cell wall biosynthesis, were up-regulated while the genes associated with salt stress resistance (CYSB, TSPO, RCI2B and CIPK6) were down-regulated in OreFLA11-OE poplars. These results indicated that OreFLA11 promotes wood formation and modulates stem mechanical properties but decreases tolerance to salt stress by decreasing the transcript abundance of salt-responsive genes in trees.

Promoter and domain structures regulate FLA12 function during Arabidopsis secondary wall development.

Fasciclin-like arabinogalactan-proteins (FLAs) are a family of multi-domain glycoproteins present at the cell surface and walls of plants. Arabidopsis thaliana FLA12 and homologs in cotton, Populus, and flax have been shown to play important functions regulating secondary cell wall (SCW) development. FLA12 has been shown to have distinct roles from the closely related FLA11 that also functions during SCW development. The promoter and domain features of FLA12 that regulate functional specificity have not been well characterized. In this study, promoter swap experiments of FLA11 and FLA12 were investigated. Mutation of proposed functional regions within FLA12 were used to investigate the role of post-translational modifications on sub-cellular location and trafficking. Domain swap experiments between FLA11 and FLA12 were performed to identify regions of functional specificity. Promote swap experiments showed that FLA12 is differentially expressed in both stem and rosette leaves compared to FLA11. Post-translational modifications, in particular addition of the glycosylphosphatidylinositol-anchor (GPI-anchor), were shown to be important for FLA12 location at the plasma membrane (PM)/cell wall interface. Domain swap experiments between FLA11 and FLA12 showed that the C-terminal arabinogalactan (AG) glycan motif acts as a key regulatory region differentiating FLA12 functions from FLA11. Understanding of FLA12 promoter and functional domains has provided new insights into the regulation of SCW development and functional specificity of FLAs for plant growth and development.

Drought-responsive genes in tomato: meta-analysis of gene expression using machine learning

Plants have diverse molecular mechanisms to protect themselves from biotic and abiotic stressors and adapt to changing environments. To uncover the genetic potential of plants, it is crucial to understand how they adapt to adverse conditions by analyzing their genomic data. We analyzed RNA-Seq data from different tomato genotypes, tissue types, and drought durations. We used a time series scale to identify early and late drought-responsive gene modules and applied a machine learning method to identify the best responsive genes to drought. We demonstrated six candidate genes of tomato viz. Fasciclin-like arabinogalactan protein 2 (FLA2), Amino acid transporter family protein (ASCT), Arginine decarboxylase 1 (ADC1), Protein NRT1/PTR family 7.3 (NPF7.3), BAG family molecular chaperone regulator 5 (BAG5) and Dicer-like 2b (DCL2b) were responsive to drought. We constructed gene association networks to identify their potential interactors and found them drought-responsive. The identified candidate genes can help to explore the adaptation of tomato plants to drought. Furthermore, these candidate genes can have far-reaching implications for molecular breeding and genome editing in tomatoes, providing insights into the molecular mechanisms that underlie drought adaptation. This research underscores the importance of the genetic basis of plant adaptation, particularly in changing climates and growing populations.

Characterization of the FLA Gene Family in Tomato (Solanum lycopersicum L.) and the Expression Analysis of SlFLAs in Response to Hormone and Abiotic Stresses.

Fasciclin-like arabinogalactan proteins (FLAs), a subclass of arabinogalactan proteins (AGPs), participate in mediating plant growth, development, and response to abiotic stress. However, the characterization and function of FLAs in tomato are currently unknown. In this study, members of the tomato FLA family are characterized and analyzed in relation to their response to phytohormonal and abiotic stresses. The results show that a total of 24 FLA members were characterized in tomato. The structural domain analysis showed that these members have a high protein similarity. The expression profiles of different tissues indicated that the genes of most members of the tomato FLA gene family are highly expressed in roots, but to a lower extent in fruits. qRT-PCR analysis revealed that all 24 tomato FLA genes are responsive to ABA and MeJA. SlFLAs showed a positive response to salt and cold stress. SlFLA1, SlFLA12, and SlFLA14 are significantly induced under darkness. SlFLA1 and SlFLA3 are significantly induced under drought stress. This study provides a basis for a further understanding of the role of tomato FLA homologous genes in plant response to abiotic stress and lays the foundation for further research on the function of FLAs in tomato.

Distinct functions of FASCILIN-LIKE ARABINOGALACTAN PROTEINS relate to domain structure.

The role of glycoproteins as key cell surface molecules during development and stress is well established; yet, the relationship between their structural features and functional mechanisms is poorly defined. FASCICLIN-LIKE ARABINOGALACTAN PROTEINs (FLAs), which impact plant growth and development, are an excellent example of a glycoprotein family with a complex multidomain structure. FLAs combine globular fasciclin-like (FAS1) domains with regions that are intrinsically disordered and contain glycomotifs for directing the addition of O-linked arabinogalactan (AG) glycans. Additional posttranslational modifications on FLAs include N-linked glycans in the FAS1 domains, a cleaved signal peptide at the N terminus, and often a glycosylphosphatidylinositol (GPI) anchor signal sequence at the C terminus. The roles of glycosylation, the GPI anchor, and FAS1 domain functions in the polysaccharide-rich extracellular matrix of plants remain unclear, as do the relationships between them. In this study, we examined sequence-structure-function relationships of Arabidopsis (Arabidopsis thaliana) FLA11, demonstrated to have roles in secondary cell wall (SCW) development, by introducing domain mutations and functional specialization through domain swaps with FLA3 and FLA12. We identified FAS1 domains as essential for FLA function, differentiating FLA11/FLA12, with roles in SCW development, from FLA3, specific to flowers and involved in pollen development. The GPI anchor and AG glycosylation co-regulate the cell surface location and release of FLAs into cell walls. The AG glycomotif sequence closest to the GPI anchor (AG2) is a major feature differentiating FLA11 from FLA12. The results of our study show that the multidomain structure of different FLAs influences their subcellular location and biological functions during plant development.

Characterization of the Fasciclin-like arabinogalactan gene family in Brassica napus and the negative regulatory role of BnFLA39 in response to clubroot disease stress

Fasciclin-like arabinogalactan proteins (FLAs) are critical for regulation of plant development and environmental adaptation. Although FLAs have been reported in numerous plants, the FLAs family members in Brassica napus and their responses to plant pathogens are unknown. In this study, 59 BnFLAs identified from a genome-wide study were classified into 4 groups based on conserved motif compositions, gene structure analyses, and phylogenetic relationships. Furthermore, a number of stress-responsive and phytohormone responsive hormonal cis-regulatory elements were identified from BnFLAs, promoters, and may be the targets of transcription factors like ERF that are important in stress responses. Subcellular localization showed that most BnFLAs are localized in the cell membrane. Moreover, expression patterns at different growth stages showed that the BnFLAs are actively involved in B. napus growth and developmental processes. Expression analyses also revealed that BnFLAs respond significantly to salt and Plasmodiophora brassicae (Pb) stresses. In addition, a BnFLA39 homolog, AtFLA7, was shown to be a negative regulator of the defense responses of Arabidopsis to Pb stress. The protein interaction analysis revealed that BnFLA39 exerted biological functions by interacting with BnFLA15/43/26/53. This study provides the basis for further research on BnFLAs functions and regulatory mechanisms and provides insights for a better understanding of the roles of FLAs in Brassisca-Pb interactions.

Genome-Wide Comparative Analysis of the Fasciclin-like Arabinogalactan Proteins (FLAs) in Salicacea and Identification of Secondary Tissue Development-Related Genes.

Fasciclin-like arabinogalactan proteins (FLAs) are a subclass of arabinogalactan proteins (AGPs) containing both AGP-like glycated domains and fasciclin (FAS) domains, which are involved in plant growth and development and synthesis of the cell wall. However, these proteins have not been identified or analyzed in willow, Salix, the sister genus of Populus. In this study, we performed a whole genome study of the FLA gene family of Salix suchowensis and compared it with the FLA gene family of Populus deltoides. The results showed the presence of 40 and 46 FLA genes in P. deltoides and S. suchowensis, distributed on 17 and 16 chromosomes, respectively. Four pairs of tandem repeat genes were found in willow, while poplar had no tandem repeat genes. Twelve and thirteen pairs of duplicated gene fragments were identified in poplar and willow, respectively. The multispecies phylogenetic tree showed that the FLA gene family could be divided into four groups (I-IV), with Group 1 showing significant expansion in woody plants. A gene expression analysis showed that PdeFLA19/27 in Group I of poplar was highly expressed, specifically during the secondary growth period of the stem and the rapid elongation of seed hairs. In the Group I genes of S. suchowensis, SsuFLA25/26/28 was also highly expressed during the secondary growth period, whereas increased expression of SsuFLA35 was associated with seed hair tissue. These results provide important clues about the differences in the FLA gene family during the evolution of herbs and woody plants, and suggest that the FLA gene family may play an essential role in regulating the secondary growth of woody plants. It also provides a reference for further studies on the regulation of secondary growth and seed hair development by FLA genes in poplar and willow.

Cas9/gRNA-Mediated Mutations in PtrFLA40 and PtrFLA45 Reveal Redundant Roles in Modulating Wood Cell Size and SCW Synthesis in Poplar

Fasciclin-like arabinogalactan proteins (FLAs) play an important role in plant development and adaptation to the environment. However, the roles of FLAs in wood formation remain poorly understood. Here, we identified a total of 50 PtrFLA genes in poplar. They were classified into four groups: A to D, among which group A was the largest group with 28 members clustered into four branches. Most PtrFLAs of group A were dominantly expressed in developing xylem based on microarray and RT-qPCR data. The roles of PtrFLA40 and PtrFLA45 in group A were investigated via the Cas9/gRNA-induced mutation lines. Loss of PtrFLA40 and PtrFLA45 increased stem length and diameter in ptrfla40ptrfla45 double mutants, but not in ptrfla40 or ptrfla45 single mutants. Further, our findings indicated that the ptrfla40ptrfla45 mutants enlarged the cell size of xylem fibers and vessels, suggesting a negative modulation in stem xylem cell size. In addition, wood lignin content in the ptrfla40fla45 mutants was increased by nearly 9%, and the lignin biosynthesis-related genes were significantly up-regulated in the ptrfla40fla45 mutants, in agreement with the increase in wood lignin content. Overall, Cas9/gRNA-mediated mutations in PtrFLA40 and PtrFLA45 reveal redundant roles in modulating wood cell size and secondary cell wall (SCW) synthesis in poplar.

Transcriptomic Evidence Reveals Low Gelatinous Layer Biosynthesis in Neolamarckia cadamba after Gravistimulation.

Trees can control their shape and resist gravity by producing tension wood (TW), which is a special wood that results from trees being put under stress. TW is characterized by the presence of a gelatinous layer (G layer) and the differential distribution of cell wall polymers. In this study, we investigated whether or not gravistimulation in N. cadamba resulted in TW with an obvious G layer. The results revealed an absence of an obvious G layer in samples of the upper side of a leaning stem (UW), as well as an accumulation of cellulose and a decrease in lignin content. A negligible change in the content of these polymers was recorded and compared to untreated plant (NW) samples, revealing the presence of a G layer either in much lower concentrations or in a lignified form. A transcriptomic investigation demonstrated a higher expression of cell wall esterase- and hydrolase-related genes in the UW, suggesting an accumulation of noncellulosic sugars in the UW, similar to the spectroscopy results. Furthermore, several G-layer-specific genes were also downregulated, including fasciclin-like arabinogalactan proteins (FLA), beta-galactosidase (BGAL) and chitinase-like proteins (CTL). The gene coexpression network revealed a strong correlation between cell-wall-synthesis-related genes and G-layer-synthesis-specific genes, suggesting their probable antagonistic role during G layer formation. In brief, the G layer in N. cadamba was either synthesized in a very low amount or was lignified during an early stage of growth; further experimental validation is required to understand the exact mechanism and stage of G layer formation in N. cadamba during gravistimulation.

Plasma membrane proteomic changes of Arabidopsis DRP1E during cold acclimation in association with the enhancement of freezing tolerance.

The freezing tolerance of plants that live in cold regions increases after exposure to low temperature, a process termed cold acclimation (CA). During CA, restructuring of the plasma membrane (PM) is important to enhance freezing tolerance. We have previously shown that the function of DYNAMIN-RELATED PROTEIN 1 E (DRP1E), which regulates endocytosis by pinching vesicles from the PM, is associated with the enhancement of freezing tolerance during CA in Arabidopsis. DRP1E is predicted to play a role in reconstituting the PM composition during CA. In this study, to test the validity of this hypothesis, we studied the changes in PM proteome patterns induced by drp1e mutation. In a detailed physiological analysis, after 3 days of CA, only young leaves showed significantly less increase in freezing tolerance in the mutant than in the wild type (WT). Using nano-liquid chromatography-tandem mass spectrometry, 496 PM proteins were identified. Among these proteins, 81 or 71 proteins were specifically altered in the WT or the mutant, respectively, in response to CA. Principal component analysis showed that the proteomic pattern differed between the WT and the mutant upon cold acclimation (CA), suggesting that DRP1E contributes to reconstruction of the PM during CA. Cluster analysis revealed that proteins that were significantly increased in the mutant after CA were biased toward glycosylphosphatidylinositol-anchored proteins, such as fasciclin-like arabinogalactan proteins. Thus, a primary target of DRP1E-associated PM reconstruction during CA is considered to be glycosylphosphatidylinositol-anchored proteins, which may be removed from the PM by DRP1E in young leaves after 3 days of CA.

The fasciclin-like arabinogalactan proteins of Camellia oil tree are involved in pollen tube growth

Fasciclin-like arabinogalactan proteins (FLAs) are a class of highly glycosylated glycoproteins that perform crucial functions in plant growth and development. This study was carried out to further explore their roles in pollen tube growth. The results showed that seven members (CoFLA1/2/3/4/7/8/17) of the CoFLAs family were identified by sequence characteristics, and they all possessed the fasciclin 1 (FAS1) domain and H1 and H2 conserved domains. They were all located on the plasma membranes of tobacco epidermal cells, and the GPI-anchor sequences of CoFLA1/2/3/4 determined the membrane localization. In flower tissues, CoFLA2 and CoFLA8 were not expressed in the pollen tube but were expressed in the unpollinated style and ovary; the others were all expressed in the pollen tube. In the pollination-compatible style and ovary, they exhibited different expression patterns. Furthermore, all CoFLAs promoted pollen germination in vitro, while only CoFLA7 significantly promoted pollen tube elongation, and the expression of CoFLA1/3/4/7/17 in pollen tubes was regulated by CoFLA proteins. The ABA and ABA synthetic inhibitor (sodium tungstate, ST) both inhibited pollen tube elongation; however, only ST downregulated the expression of CoFLA1/7/17 and upregulated the expression of CoFLA4. Taken together, these results demonstrate that CoFLAs may be significant in pollen tube growth in C. oleifera and that some CoFLAs may participate in the regulation of ABA signaling.

Comparative Transcriptome Analysis Reveals the Mechanisms Underlying Differential Seed Vigor in Two Contrasting Peanut Genotypes

Seed vigor is an important agronomic trait, and wide variation exists among peanut accessions. However, the detailed regulatory mechanisms underlying differences in seed vigor between varieties are not known in peanut yet. Here, we performed a comparative transcriptome analysis of germinating seeds in two contrasting peanut accessions, namely A86 (high-vigor variety) and A279 (low-vigor variety). A total of 583 and 860 differentially expressed genes (DEGs) were identified at two imbibition stages between A86 and A279, respectively. Pathway enrichment tests highlighted the cell wall remodeling-, hormone signaling-, transcriptional regulation-, and oxidative stress-related DEGs, which may explain to a certain extent the difference in seed vigor between the two cultivars. Among them, the largest number of cell wall remodeling-related DEGs were extensions followed by cellulose synthases, fasciclin-like arabinogalactan proteins, polygalacturonases, expansins, and pectinesterases and the hormone signaling-related DEGs belonged mainly to the auxin and ethylene signaling pathway. The majority of transcriptional regulation-related DEGs were MYB, FAR1, and bHLH transcription factors, and the oxidative stress-related DEGs were mainly peroxidases. Further physiological analyses indicated that differences in seed vigor between A86 and A279 may be associated with differences in the ROS-scavenging abilities mediated by peroxidases. Moreover, we identified 16 DEGs homologous to known Arabidopsis regulators of seed dormancy and germination, suggesting that these DEGs would play similar functional roles during peanut seed germination. Our results not only provide important insights into the difference in seed vigor between varieties, but offer candidate genes that are worth investigating in future studies.

DEAP1 encodes a fasciclin-like arabinogalactan protein required for male fertility in rice.

Arabinogalactan proteins (AGPs) are widely distributed in plant cells. Fasciclin-like AGPs (FLAs) belong to a subclass of AGPs that play important roles in plant growth and development. However, little is known about the biological functions of rice FLA. Herein, we report the identification of a male-sterile mutant of DEFECTIVE EXINE AND APERTURE PATTERNING1 (DEAP1) in rice. The deap1 mutant anthers produced aberrant pollen grains with defective exine formation and a flattened aperture annulus and exhibited slightly delayed tapetum degradation. DEAP1 encodes a plasma membrane-associated member of group III plant FLAs and is specifically and temporally expressed in reproductive cells and the tapetum layer during male development. Gene expression studies revealed reduced transcript accumulation of genes related to exine formation, aperture patterning, and tapetum development in deap1 mutants. Moreover, DEAP1 may interact with two rice D6 PROTEIN KINASE-LIKE3s (OsD6PKL3s), homologs of a known Arabidopsis aperture protein, to affect rice pollen aperture development. Our findings suggested that DEAP1 is involved in male reproductive development and may affect exine formation and aperture patterning, thereby providing new insights into the molecular functions of plant FLAs in male fertility.

Integrated multi-omics analysis provides insights into genome evolution and phosphorus deficiency adaptation in pigeonpea (Cajanus cajan).

Pigeonpea (Cajanus cajan) is an important legume food crop and plays a crucial role in a secure food supply in many developing countries. Several previous studies have suggested that pigeonpea has great potential for phosphorus (P) deficiency tolerance, but little is known about the underlying mechanism. In this study, the physiological and molecular responses of pigeonpea roots to phosphate (Pi) starvation were investigated through integrating phenotypic, genomic, transcriptomic, metabolomic, and lipidomic analyses. The results showed that low-Pi treatment increased total root length, root surface area, and root acid phosphatase activity, and promoted the secretion of organic acids (e.g. citric acids, piscidic acids, and protocatechuic acids) and the degradation of phospholipids and other P-containing metabolites in the roots of pigeonpea. Consistent with the morphological, physiological, and biochemical changes, a large number of genes involved in these Pi-starvation responses were significantly upregulated in Pi-deficient pigeonpea roots. Among these Pi-starvation response genes upregulated by low-Pi treatment, four gene families were expanded through recent tandem duplication in the pigeonpea genome, namely phosphate transporter 1 (PHT1), phosphoethanolamine/phosphocholine phosphatase (PECP), fasciclin-like arabinogalactan protein (FLA), and glutamate decarboxylase (GAD). These gene families may be associated with Pi uptake from the soil, phospholipid recycling, root morphological remodeling, and regulation of organic acid exudation. Taken together, our results suggest that pigeonpea employs complex Pi-starvation responses to strengthen P acquisition and utilization during low-Pi stress. This study provides new insights into the genome evolution and P deficiency adaptation mechanism of pigeonpea.

OsFLA1 encodes a fasciclin-like arabinogalactan protein and affects pollen exine development in rice.

OsFLA1 positively regulates pollen exine development, and locates in the cellular membrane. Arabinogalactan proteins are a type of hydroxyproline-rich glycoprotein that are present in all plant tissues and cells and play important roles in plant growth and development. Little information is available on the participation of fasciclin-like arabinogalactan proteins in sexual reproduction in rice. In this study, a rice male-sterile mutant, osfla1, was isolated from an ethylmethanesulfonate-induced mutant library. The osfla1 mutant produced withered, shrunken, and abortive pollen. The gene OsFLA1 encoded a FLA protein and was expressed strongly in the anthers in rice. Subcellular localization showed that OsFLA1 was located in the cellular membrane. In the osfla1 mutant, abnormal Ubisch bodies and a discontinuous nexine layer of the microspore wall were observed, which resulted in pollen abortion and ultimately in male sterility. The results show the important role that OsFLA1 plays in male reproductive development in rice.

Comprehensive Analysis of Arabinogalactan Protein-Encoding Genes Reveals the Involvement of Three BrFLA Genes in Pollen Germination in Brassica rapa.

Arabinogalactan proteins (AGPs) are a superfamily of hydroxyproline-rich glycoproteins that are massively glycosylated, widely implicated in plant growth and development. No comprehensive analysis of the AGP gene family has been performed in Chinese cabbage (Brassica rapa ssp. chinensis). Here, we identified a total of 293 putative AGP-encoding genes in B. rapa, including 25 classical AGPs, three lysine-rich AGPs, 30 AG-peptides, 36 fasciclin-like AGPs (FLAs), 59 phytocyanin-like AGPs, 33 xylogen-like AGPs, 102 other chimeric AGPs, two non-classical AGPs and three AGP/extensin hybrids. Their protein structures, phylogenetic relationships, chromosomal location and gene duplication status were comprehensively analyzed. Based on RNA sequencing data, we found that 73 AGP genes were differentially expressed in the floral buds of the sterile and fertile plants at least at one developmental stage in B. rapa, suggesting a potential role of AGPs in male reproductive development. We further characterized BrFLA2, BrFLA28 and BrFLA32, three FLA members especially expressed in anthers, pollen grains and pollen tubes. BrFLA2, BrFLA28 and BrFLA32 are indispensable for the proper timing of pollen germination under high relative humidity. Our study greatly extends the repertoire of AGPs in B. rapa and reveals a role for three members of the FLA subfamily in pollen germination.

Proteins, Small Peptides and Other Signaling Molecules Identified as Inconspicuous but Possibly Important Players in Microspores Reprogramming Toward Embryogenesis

In this review, we describe and integrate the latest knowledge on the signaling role of proteins and peptides in the stress-induced microspore embryogenesis (ME) in some crop plants with agricultural importance (i.e., oilseed rape, tobacco, barley, wheat, rice, triticale, rye). Based on the results received from the most advanced omix analyses, we have selected some inconspicuous but possibly important players in microspores reprogramming toward embryogenic development. We provide an overview of the roles and downstream effect of stress-related proteins (e.g., β-1,3-glucanases, chitinases) and small signaling peptides, especially cysteine—(e.g., glutathione, γ-thionins, rapid alkalinization factor, lipid transfer, phytosulfokine) and glycine-rich peptides and other proteins (e.g., fasciclin-like arabinogalactan protein) on acclimation ability of microspores and the cell wall reconstruction in a context of ME induction and haploids/doubled haploids (DHs) production. Application of these molecules, stimulating the induction and proper development of embryo-like structures and green plant regeneration, brings significant improvement of the effectiveness of DHs procedures and could result in its wider incorporation on a commercial scale. Recent advances in the design and construction of synthetic peptides–mainly cysteine-rich peptides and their derivatives–have accelerated the development of new DNA-free genome-editing techniques. These new systems are evolving incredibly fast and soon will find application in many areas of plant science and breeding.

The Arabinogalactan Protein Family of Centaurium erythraea Rafn.

Centaurium erythraea (centaury) is a medicinal plant with exceptional developmental plasticity in vitro and vigorous, often spontaneous, regeneration via shoot organogenesis and somatic embryogenesis, during which arabinogalactan proteins (AGPs) play an important role. AGPs are highly glycosylated proteins belonging to the super family of O-glycosylated plant cell surface hydroxyproline-rich glycoproteins (HRGPs). HRGPs/AGPs are intrinsically disordered and not well conserved, making their homology-based mining ineffective. We have applied a recently developed pipeline for HRGP/AGP mining, ragp, which is based on machine learning prediction of proline hydroxylation, to identify HRGP sequences in centaury transcriptome and to classify them into motif and amino acid bias (MAAB) classes. AGP sequences with low AG glycomotif representation were also identified. Six members of each of the three AGP subclasses, fasciclin-like AGPs, receptor kinase-like AGPs and AG peptides, were selected for phylogenetic and expression analyses. The expression of these 18 genes was recorded over 48 h following leaf mechanical wounding, as well as in 16 tissue samples representing plants from nature, plants cultivated in vitro, and developmental stages during shoot organogenesis and somatic embryogenesis. None of the selected genes were upregulated during both wounding recovery and regeneration. Possible functions of AGPs with the most interesting expression profiles are discussed.