New insights into evolution and functional diversification of Camellia sinensis LRR-RLKs.

Leucine-rich repeat receptor-like kinases (LRR-RLKs) represent the largest subgroup of receptor-like kinases (RLKs) in plants. While some LRR-RLK members play a role in regulating various plant growth processes related to morphogenesis, disease resistance, and stress response, the functions of most LRR-RLK genes remain unclear. In this study, we identified 397 LRR-RLK genes from the genome of Camellia sinensis and categorized them into 16 subfamilies. Approximately 62% of CsLRR-RLK genes are situated in regions resulting from segmental duplications, suggesting that the expansion of CsLRR-RLK genes is due to segmental duplications. Analysis of gene expression patterns revealed differential expression of CsLRR-RLK genes across different tissues and in response to stress. Furthermore, we demonstrated that CssEMS1 localizes to the cell membrane and can complement Arabidopsis ems1 mutant. This study is the initial in-depth evolutionary examination of LRR-RLKs in tea and provides a basis for future investigations into their functionality. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-024-01458-1.

Lineage-specific gene duplication and expansion of DUF1216 gene family in Brassicaceae.

Proteins containing domain of unknown function (DUF) are prevalent in eukaryotic genome. The DUF1216 proteins possess a conserved DUF1216 domain resembling to the mediator protein of Arabidopsis RNA polymerase II transcriptional subunit-like protein. The DUF1216 family are specifically existed in Brassicaceae, however, no comprehensive evolutionary analysis of DUF1216 genes have been performed. We performed a first comprehensive genome-wide analysis of DUF1216 proteins in Brassicaceae. Totally 284 DUF1216 genes were identified in 27 Brassicaceae species and classified into four subfamilies on the basis of phylogenetic analysis. The analysis of gene structure and conserved motifs revealed that DUF1216 genes within the same subfamily exhibited similar intron/exon patterns and motif composition. The majority members of DUF1216 genes contain a signal peptide in the N-terminal, and the ninth position of the signal peptide in most DUF1216 is cysteine. Synteny analysis revealed that segmental duplication is a major mechanism for expanding of DUF1216 genes in Brassica oleracea, Brassica juncea, Brassica napus, Lepidium meyneii, and Brassica carinata, while in Arabidopsis thaliana and Capsella rubella, tandem duplication plays a major role in the expansion of the DUF1216 gene family. The analysis of Ka/Ks (non-synonymous substitution rate/synonymous substitution rate) ratios for DUF1216 paralogous indicated that most of gene pairs underwent purifying selection. DUF1216 genes displayed a specifically high expression in reproductive tissues in most Brassicaceae species, while its expression in Brassica juncea was specifically high in root. Our studies offered new insights into the phylogenetic relationships, gene structures and expressional patterns of DUF1216 members in Brassicaceae, which provides a foundation for future functional analysis.

Origin, evolution, and diversification of inositol 1,4,5-trisphosphate 3-kinases in plants and animals.

BACKGROUND: In Eukaryotes, inositol polyphosphates (InsPs) represent a large family of secondary messengers and play crucial roes in various cellular processes. InsPs are synthesized through a series of pohophorylation reactions catalyzed by various InsP kinases in a sequential manner. Inositol 1,4,5-trisphosphate 3-kinase (IP3 3-kinase/IP3K), one member of InsP kinase, plays important regulation roles in InsPs metabolism by specifically phosphorylating inositol 1,4,5-trisphosphate (IP3) to inositol 1,3,4,5-tetrakisphosphate (IP4) in animal cells. IP3Ks were widespread in fungi, plants and animals. However, its evolutionary history and patterns have not been examined systematically. RESULTS: A total of 104 and 31 IP3K orthologues were identified across 57 plant genomes and 13 animal genomes, respectively. Phylogenetic analyses indicate that IP3K originated in the common ancestor before the divergence of fungi, plants and animals. In most plants and animals, IP3K maintained low-copy numbers suggesting functional conservation during plant and animal evolution. In Brassicaceae and vertebrate, IP3K underwent one and two duplication events, respectively, resulting in multiple gene copies. Whole-genome duplication (WGD) was the main mechanism for IP3K duplications, and the IP3K duplicates have experienced functional divergence. Finally, a hypothetical evolutionary model for the IP3K proteins is proposed based on phylogenetic theory. CONCLUSION: Our study reveals the evolutionary history of IP3K proteins and guides the future functions of animal, plant, and fungal IP3K proteins.

ALLENE OXIDE SYNTHASE (AOS) induces petal senescence through a novel JA-associated regulatory pathway in Arabidopsis.

Flowers are crucial for the reproduction of flowering plants and their senescence has drastic effects on plant-animal interactions as well as pollination. Petal senescence is the final phase of flower development which is regulated by hormones and genes. Among these, jasmonic acid (JA) has emerged as a major contributor to petal senescence, but its molecular mechanisms remain elusive. Here, the role of JA in petal senescence in Arabidopsis was investigated. We showed that petal senescence in aos mutant was significantly delayed, which also affected petal cell size and proliferation. Similar significant delays in petal senescence were observed in dad1 and coi1 mutants. However, MYB21/24 and MYC2/3/4, known downstream regulators of JA in flower development, played no role in petal senescence. This indicated that JA regulates petal senescence by modulating other unknown transcription factors. Transcriptomic analysis revealed that AOS altered the expression of 3681 genes associated, and identified groups of differentially expressed transcription factors, highlighting the potential involvement of AP-2, WRKY and NAC. Furthermore, bHLH13, bHLH17 and URH2 were identified as potential new regulators of JA-mediated petal senescence. In conclusion, our findings suggest a novel genetic pathway through which JA regulates petal senescence in Arabidopsis. This pathway operates independently of stamen development and leaf senescence, suggesting the evolution of specialized mechanisms for petal senescence. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-024-01425-w.

Development and genetic regulation of pollen intine in Arabidopsis and rice.

Pollen intine serves as a protective layer situated between the pollen exine and the plasma membrane. It performs essential functions during pollen development, including maintaining the morphological structure of the pollen, preventing the loss of pollen contents, and facilitating pollen germination. The formation of the intine layer commences at the bicellular pollen stage. Pectin, cellulose, hemicellulose and structural proteins are the key constituents of the pollen intine. In Arabidopsis and rice, numerous regulatory factors associated with polysaccharide metabolism and material transport have been identified, which regulate intine development. In this review, we elucidate the developmental processes of the pollen wall and provide a concise summary of the research advancements in the development and genetic regulation of the pollen intine in Arabidopsis and rice. A comprehensive understanding of intine development and regulation is crucial for unraveling the genetic network underlying intine development in higher plants.

Origin, evolution and diversification of plant mechanosensitive channel of small conductance-like (MSL) proteins.

Mechanosensitive (MS) ion channels provide efficient molecular mechanism for transducing mechanical forces into intracellular ion fluxes in all kingdoms of life. The mechanosensitive channel of small conductance (MscS) was one of the best-studied MS channels and its homologs (MSL, MscS-like) were widely distributed in cell-walled organisms. However, the origin, evolution and expansion of MSL proteins in plants are still not clear. Here, we identified more than 2100 MSL proteins from 176 plants and conducted a broad-scale phylogenetic analysis. The phylogenetic tree showed that plant MSL proteins were divided into three groups (I, II and III) prior to the emergence of chlorophytae algae, consistent with their specific subcellular localization. MSL proteins were distributed unevenly into each of plant species, and four parallel expansion was identified in angiosperms. In Brassicaceae, most MSL duplicates were derived by whole-genome duplication (WGD)/segmental duplications. Finally, a hypothetical evolutionary model of MSL proteins in plants was proposed based on phylogeny. Our studies illustrate the evolutionary history of the MSL proteins and provide a guide for future functional diversity analyses of these proteins in plants.

Cell Biological Analyses of Anther Morphogenesis and Pollen Viability in Arabidopsis and Rice.

Major advances have been made in our understanding of anther developmental processes in flowering plants through a combination of genetic studies, cell biological technologies, biochemical analyses, microarray and high-throughput sequencing-based approaches. In this chapter, we summarize widely used protocols for pollen viability staining, investigation of anther morphogenesis by scanning electron microscopy (SEM), light microscopy of semi-thin sections, ultrathin section-based transmission electron microscopy (TEM), TUNEL (terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate (dUTP) nick end labeling) assay for tapetum programmed cell death, and laser microdissection procedures to obtain specific cells or cell layers for transcriptome analysis.

Understanding the Origin and Evolution of Tea (Camellia sinensis [L.]): Genomic Advances in Tea.

Tea, which is processed by the tender shoots or leaves of tea plant (Camellia sinensis), is one of the most popular nonalcoholic beverages in the world and has numerous health benefits for humans. Along with new progress in biotechnologies, the refined chromosome-scale reference tea genomes have been achieved, which facilitates great promise for the understanding of fundamental genomic architecture and evolution of the tea plants. Here, we summarize recent achievements in genome sequencing in tea plants and review the new progress in origin and evolution of tea plants by population sequencing analysis. Understanding the genomic characterization of tea plants is import to improve tea quality and accelerate breeding in tea plants.

Peptide signaling in anther development and pollen-stigma interactions.

Polypeptides play irreplaceable roles in cell-cell communication by binding to receptor-like kinases. Various types of peptide-receptor-like kinase-mediated signaling have been identified in anther development and male-female interactions in flowering plants. Here, we provide a comprehensive summary of the biological functions and signaling pathways of peptides and receptors involved in anther development, self-incompatibility, pollen tube growth and pollen tube guidance.

The lineage-specific evolution of the oleosin family in Theaceae.

Oleosins play essential roles in stabilization of lipid droplets (LDs) and seed oil production. However, evolution of this gene family has not been reported in Theaceae, a large plant family that contains many important tea and oil tea species. In this study, a total of 65 oleosin genes were identified in nine genome-sequenced Theaceae species. Among these genomes, the gene number of oleosin showed significant difference, with Camellia sinensis var. sinensis cv. Shuchazao and Camellia lanceoleosa displayed more oleosin numbers than other species. Phylogenetic analyses revealed that Theaceae oleosin genes were classified into three clades (U, SL, SH) respectively. Proteins within the same clade had similar gene structure and motif composition. Segmental duplication was the primary driving force for the evolution of oleosin genes in Shuchazao (SCZ), Huangdan (HD), C.lanceoleosa (Cla), and wild tea (DASZ). Synteny analysis showed that most oleosin genes displayed inter-species synteny among tea and oil tea species. Expression analysis demonstrated that oleosin genes were specifically expressed in seed and kernel of Huangdan (HD) and C.lanceoleosa. Moreover, expression divergence was observed in paralogous pairs and ∼1-2 oleosin genes in each clade have become activate. This study leads to a comprehensive understanding of evolution of oleosin family in Theaceae, and provides a rich resource to further address the functions of oleosin in tea and oil tea species.

Effect of prior drought and heat stress on Camellia sinensis transcriptome changes to Ectropis oblique (Lepidoptera: Geometridae) resistance.

Tea plants are continuously confronted with a wide range of biotic and abiotic stressors in the field, which can occur concurrently or sequentially. To elucidate the molecular mechanisms in responses to such individual and combined stresses, we used RNAseq to compare the temporal changes in the transcriptome of Camellia sinensis to Ectropis oblique Prout alone or in combination with exposure to drought and heat. Compared with the individual stress, tea plants exhibit significant differences in transcriptome profiles under the combined stresses. Additionally, many unique genes exhibited significant differences in expression in individual and combined stress conditions. Our research showed novel insights into the molecular mechanisms of E. oblique Prout resistance in tea plants and provided a valuable resource for developing tea varieties with broad spectrum stress tolerance.

Expansion and Diversification of the 14-3-3 Gene Family in Camellia sinensis.

14-3-3 proteins are signal moderators in sensing various stresses and play essential functions in plant growth and development. Although, 14-3-3 gene families have been identified and characterized in many plant species, its evolution has not been studied systematically. In this study, the plant 14-3-3 family was comprehensively analyzed from green algae to angiosperm. Our result indicated that plant 14-3-3 originated during the early evolutionary history of green algae and expanded in terricolous plants. Twenty-six 14-3-3 genes were identified in the tea genome. RNA-seq analysis showed that tea 14-3-3 genes display different expression patterns in different organs. Moreover, the expression of most tea 14-3-3 genes displayed variable expression patterns under different abiotic and biotic stresses. In conclusion, our results elucidate the evolutionary origin of plant 14-3-3 genes, and beneficial for understanding their biological functions and improving tea agricultural traits in the future.

Tapetal 3-Ketoacyl-Coenzyme A Synthases Are Involved in Pollen Coat Lipid Accumulation for Pollen-Stigma Interaction in Arabidopsis.

Pollen coat lipids form an outer barrier to protect pollen itself and play essential roles in pollen-stigma interaction. However, the precise molecular mechanisms underlying the production, deposition, regulation, and function of pollen coat lipids during anther development remain largely elusive. In lipid metabolism, 3-ketoacyl-coenzyme A synthases (KCS) are involved in fatty acid elongation or very-long-chain fatty acid (VLCFA) synthesis. In this study, we identified six members of the Arabidopsis KCS family expressed in anther. Among them, KCS7, KCS15, and KCS21 were expressed in tapetal cells at anther stages 8-10. Further analysis demonstrated that they act downstream of male sterility 1 (MS1), a regulator of late tapetum development. The kcs7/15/21 triple mutant is fertile. Both cellular observation and lipid staining showed pollen coat lipid was decreased in kcs7/15/21 triple mutant. After landing on stigma, the wild-type pollen grains were hydrated for about 5 min while the kcs7/15/21 triple mutant pollen took about 10 min to hydrate. Pollen tube growth of the triple mutant was also delayed. These results demonstrate that the tapetum-localized KCS proteins are involved in the accumulation of pollen coat lipid and reveal the roles of tapetal-derived pollen coat lipid for pollen-stigma interaction.

Genome-wide identification and expression analysis of the MADS-box transcription factor family in Camellia sinensis.

The MADS-box genes are an important class of transcription factors and play critical roles in flower development. However, the functions of these genes in the economically important drinking plant, Camellia sinensis, are still not reported. Here, an evolutionary analysis of tea MADS-box genes was performed at whole genome level. A total of 83 MADS-box genes were identified in tea, and their gene structures and expression patterns were further analyzed. The tea MADS-box genes were classified into Mα (26), Mß (12), Mγ (9), MIKC* (7), and MIKCC (29) clade according to their phylogenetic relationship with Arabidopsis thaliana. Several cis-elements were identified in the promoter regions of the CsMADS genes that are important in regulating growth, development, light responses, and the response to several stresses. Most CsMADS genes display clear different expression patterns in different organs and different species of tea plant. The expression of CsMADS genes can be regulated by abiotic stresses and phytohormone treatment. Our results lay the foundation for future research on the function of CsMADS genes and beneficial for improving tea agricultural traits in the future.

Understanding the molecular mechanism of anther development under abiotic stresses.

KEY MESSAGE: The developmental stage of anther development is generally more sensitive to abiotic stress than other stages of growth. Specific ROS levels, plant hormones and carbohydrate metabolism are disturbed in anthers subjected to abiotic stresses. As sessile organisms, plants are often challenged to multiple extreme abiotic stresses, such as drought, heat, cold, salinity and metal stresses in the field, which reduce plant growth, productivity and yield. The development of reproductive stage is more susceptible to abiotic stresses than the vegetative stage. Anther, the male reproductive organ that generate pollen grains, is more sensitive to abiotic stresses than female organs. Abiotic stresses affect all the processes of anther development, including tapetum development and degradation, microsporogenesis and pollen development, anther dehiscence, and filament elongation. In addition, abiotic stresses significantly interrupt phytohormone, lipid and carbohydrate metabolism, alter reactive oxygen species (ROS) homeostasis in anthers, which are strongly responsible for the loss of pollen fertility. At present, the precise molecular mechanisms of anther development under adverse abiotic stresses are still not fully understood. Therefore, more emphasis should be given to understand molecular control of anther development during abiotic stresses to engineer crops with better crop yield.

Phenylpropanoid Derivatives Are Essential Components of Sporopollenin in Vascular Plants.

The outer wall of pollen and spores, namely the exine, is composed of sporopollenin, which is highly resistant to chemical reagents and enzymes. In this study, we demonstrated that phenylpropanoid pathway derivatives are essential components of sporopollenin in seed plants. Spectral analyses showed that the autofluorescence of Lilium and Arabidopsis sporopollenin is similar to that of lignin. Thioacidolysis and NMR analyses of pollen from Lilium and Cryptomeria further revealed that the sporopollenin of seed plants contains phenylpropanoid derivatives, including p-hydroxybenzoate (p-BA), p-coumarate (p-CA), ferulate (FA), and lignin guaiacyl (G) units. The phenylpropanoid pathway is expressed in the tapetum in Arabidopsis, consistent with the fact that the sporopollenin precursor originates from the tapetum. Further germination and comet assays showed that this pathway plays an important role in protection of pollen against UV radiation. In the pteridophyte plant species Ophioglossum vulgatum and Lycopodium clavata, phenylpropanoid derivatives including p-BA and p-CA were also detected, but G units were not. Taken together, our results indicate that phenylpropanoid derivatives are essential for sporopollenin synthesis in vascular plants. In addition, sporopollenin autofluorescence spectra of bryophytes, such as Physcomitrella and Haplocladium, exhibit distinct characteristics compared with those of vascular plants, indicating the diversity of sporopollenin among land plants.

Genome-wide systematic characterization and expression analysis of the phosphatidylinositol 4-phosphate 5-kinases in plants.

Phosphatidylinositol 4-phosphate 5-kinases (PIP5Ks) are key enzymes in the process of phosphatidylinositol signaling pathway and have essential functions in growth, development, and biotic and abiotic stresses responses in plants. However, the evolutionary history and patterns of PIP5K genes in plants have not been examined systematically. Here, we use whole-genome sequences from eight plant species of land plants and algae to define the evolutionary relationships between these proteins in plants. 85 PIP5K genes were identified and divided into two subfamilies based on phylogenetic analyses. PIP5K members in subfamily II underwent several duplication events in land plants, resulting in multiple gene copies in angiosperms, while PIP5K members in subfamily I displayed low-copy numbers and lost in eudicots. Furthermore, PIP5K genes within the same subfamily had similar motifs and intron/exon features. Nine duplicated soybean gene pairs, four duplicated Arabidopsis gene pairs and two rice duplicated gene pairs were identified and many of them localized in synteny genomic regions. These duplicate events were formed by Whole-genome duplication (WGD)/segmental duplications. In addition, the ratios of non-synonymous to synonymous substitutions (Ka/Ks) showed that the PIP5K family had undergone purifying selection in higher plants. Expression analysis showed that PIP5K genes had complex and variable expression patterns in different developmental stages. The specificity of these genes is utilized to provide evidence for selective expression in the evolutionary process.

How to Study the Proteomes and Phosphoproteomes of Anther and Pollen.

Proteomics analysis was a powerful technology for characterizing proteins and protein posttranslational modification (PTMs). Recently, many anther and pollen-related proteomic analyses have been reported, which have expanded our understanding of anther and pollen development and regulation. In this chapter, we describe a detailed, optimized protocol for the separation, digestion, tagging, and subsequent mass spectrometry-based identification and quantification of proteins and phosphoproteins from anther and pollen.

Advances in research on functional genes of tea plant.

Tea plant (Camellia sinensis) is an important leaf-type woody crop used to produce non-alcoholic beverages all over the world. Tea is one of the oldest and most popular non-alcoholic beverages in the world, and long-term tea drinking has numerous healthful for humans due to many of the important secondary metabolites, such as polyphenols and theanine. Theanine and polyphenols are also closely related to tea flavor and tea aroma, which is usually as the standard for judging tea quality. The growth of tea plants and quality of teas are susceptible to adversity abiotic and biotic stresses, such as low temperatures and pests. Consequently, this review focus on the research progress of key genes related to the stress resistance and material metabolism of tea plants in recent years. We aim at comprehensively understanding the growth and metabolism of tea plants and their relationship with the external environment, so as to provide an in-depth and broad theoretical support for the breeding of excellent tea plant varieties.

Expansion and Functional Divergence of Inositol Polyphosphate 5-Phosphatases in Angiosperms.

Inositol polyphosphate 5-phosphatase (5PTase), a key enzyme that hydrolyzes the 5` position of the inositol ring, has essential functions in growth, development, and stress responses in plants, yeasts, and animals. However, the evolutionary history and patterns of 5PTases have not been examined systematically. Here, we report a comprehensive molecular evolutionary analysis of the 5PTase gene family and define four groups. These four groups are different from former classifications, which were based on in vitro substrate specificity. Most orthologous groups appear to be conserved as single or low-copy genes in all lineages in Groups II-IV, whereas 5PTase genes in Group I underwent several duplication events in angiosperm, resulting in multiple gene copies. Whole-genome duplication (WGD) was the main mechanism for 5PTase duplications in angiosperm. Plant 5PTases have more members than that of animals, and most plant 5PTase genes appear to have evolved under strong purifying selection. The paralogs have diverged in substrate specificity and expression pattern, showing evidence of selection pressure. Meanwhile, the increase in 5PTases and divergences in sequence, expression, and substrate might have contributed to the divergent functions of 5PTase genes, allowing the angiosperms to successfully adapt to a great number of ecological niches.