CBL-interacting Protein Kinases Research Articles

Evolution, gene expression, and protein‒protein interaction analyses identify candidate CBL-CIPK signalling networks implicated in stress responses to cold and bacterial infection in citrus

BackgroundCold is a major abiotic stress and Huanglongbing and citrus canker disease are two devastating bacterial diseases for citrus. The Ca2+-CBL-CIPK network is known to regulate different types of stress signalling in plants. How do CBL–CIPK signalling networks function in response to cold and infection by CLas or Xcc in citrus?ResultsEight calcineurin B-like proteins (CBLs) and seventeen CBL-interacting protein kinases (CIPKs) were identified from the cold-tolerant satsuma mandarin ‘Guijing2501’ (Citrus. unshiu) and CLas/Xcc-sensitive sweet orange (C. sinensis). Phylogenetic analysis revealed that both CBL and CIPK family members in citrus were classified into an ancient and a recent clade according to their conserved domain characteristics and/or intron/exon structures. Genome duplication analysis suggested that both tandem and segmental duplications contributed to the amplification of the CBL and CIPK gene families in citrus under intense purifying selection, and the duplication events only existed in the recent clades. Expression comparison of the duplicated gene pairs indicated that the duplicated CBL and CIPK genes underwent functional differentiation. Further expression analysis identified that CBL1, 5, 6, and 8 and CIPK2, 8, 12, 15, 16, and 17 were significantly regulated by multiple stresses, including cold, Xcc infection and/or CLas infection, in citrus, whereas CBL2/7 and CIPK1/4/5/11/13/14 were independently highly regulated by cold and CIPK3 was uniquely responsive to Xcc infection. The combination analyses of targeted Y2H assay and expression analysis revealed that CBL6-CIPK8 was the common signalling network in response to cold and Xcc infection, while CBL6/CBL8-CIPK14 was uniquely responsive to cold in citrus. Further stable transformation and cold tolerance assay indicated that overexpression of CuCIPK16 enhanced the cold tolerance of transgenic Arabidopsis with higher POD activity and lower MDA content.ConclusionsIn this study, evolution, gene expression and protein‒protein interaction analyses of citrus CBLs and CIPKs were comprehensively conducted over a genome-wide range. The results will facilitate future functional characterization of individual citrus CBLs and CIPKs under specific stresses and provide clues for the clarification of cold tolerance and disease susceptibility mechanisms in corresponding citrus cultivars.

Characterization of CBL-Interacting Protein Kinases’ Gene Family and Expression Pattern Reveal Their Important Roles in Response to Salt Stress in Poplar

The CBL-interacting protein kinases’ (CIPKs) gene family plays an important role in plants under salt stress. In this study, a total of 31 PtrCIPK genes were identified in poplar. CIPKs’ gene family was divided into two categories, few intron classes and multi-intron classes. They all have the core components of the kinase domain and regulatory domain unique to the CIPK gene family and share most of the same motifs. PtrCIPKs have 17 fragment repeat events and have high homology with Arabidopsis thaliana and Betula platyphylla, and partial homology with Zea mays. Prediction of cis-acting elements found that the PtrCIPK gene family has the most elements in terms of stress. Under NaCl stress, all members of poplar CIPKs’ gene family were significantly expressed. There were fourteen up-regulated genes and four down-regulated genes. Candidate gene expression was significantly higher in the phloem than in other tissues. In this study, characterization of CBL-interacting protein kinases’ gene family and expression pattern reveal their important roles in response to salt stress in poplar.

Halophyte Nitraria billardieri CIPK25 mitigates salinity-induced cell damage by alleviating H2O2 accumulation.

The plant-specific module of calcineurin B-like proteins (CBLs) and CBL-interacting protein kinases (CIPKs) play a crucial role in plant adaptation to different biotic and abiotic stresses in various plant species. Despite the importance of the CBL-CIPK module in regulating plant salt tolerance, few halophyte CIPK orthologs have been studied. We identified NbCIPK25 in the halophyte Nitraria billardieri as a salt-responsive gene that may improve salt tolerance in glycophytes. Sequence analyses indicated that NbCIPK25 is a typical CIPK family member with a conserved NAF motif, which contains the amino acids: asparagine, alanine, and phenylalanine. NbCIPK25 overexpression in salt-stressed transgenic Arabidopsis seedlings resulted in enhanced tolerance to salinity, a higher survival rate, longer newly grown roots, more root meristem cells, and less damaged root cells in comparison to wild-type (WT) plants. H2O2 accumulation and malondialdehyde (MDA) content were both deceased in NbCIPK25-transgenic plants under salt treatment. Furthermore, their proline content, an important factor for scavenging reactive oxygen species, accumulated at a significantly higher level. In concordance, the transcription of genes related to proline accumulation was positively regulated in transgenic plants under salt condition. Finally, we observed a stronger auxin response in salt-treated transgenic roots. These results provide evidence for NbCIPK25 improving salt tolerance by mediating scavenging of reactive oxygen species, thereby protecting cells from oxidation and maintaining plant development under salt stress. These findings suggest the potential application of salt-responsive NbCIPK25 for cultivating glycophytes with a higher salt tolerance through genetic engineering.

CBL-Interacting Protein Kinase 2 Improves Salt Tolerance in Soybean (Glycine max L.)

Salt stress severely limits soybean production worldwide. Calcineurin B-like protein-interacting protein kinases (CIPKs) play a pivotal role in a plant’s adaption to salt stress. However, their biological roles in soybean adaption to salt stress remain poorly understood. Here, the GmCIPK2 expression was increased by NaCl and hydrogen peroxide (H2O2). GmCIPK2-overexpression Arabidopsis and soybean hairy roots displayed improved salt tolerance, whereas the RNA interference of hairy roots exhibited enhanced salt sensitivity. Further analyses demonstrated that, upon salt stress, GmCIPK2 enhanced the proline content and antioxidant enzyme activity and decreased the H2O2 content, malondialdehyde (MDA) content, and Na+/K+ ratios in soybean. Moreover, GmCIPK2 promoted the expression of salt- and antioxidant-related genes in response to salt stress. Moreover, the GmCIPK2-interacting sensor, GmCBL4, increased the salt tolerance of soybean hairy roots. Overall, these results suggest that GmCIPK2 functions positively in soybean adaption to salt stress.

Abscisic acid influences ammonium transport via regulation of kinase CIPK23 and ammonium transporters.

Ammonium uptake at plant roots is regulated at the transcriptional, posttranscriptional, and posttranslational levels. Phosphorylation by the protein kinase calcineurin B-like protein (CBL)-interacting protein kinase 23 (CIPK23) transiently inactivates ammonium transporters (AMT1s), but the phosphatases activating AMT1s remain unknown. Here, we identified the PP2C phosphatase abscisic acid (ABA) insensitive 1 (ABI1) as an activator of AMT1s in Arabidopsis (Arabidopsis thaliana). We showed that high external ammonium concentrations elevate the level of the stress phytohormone ABA, possibly by de-glycosylation. Active ABA was sensed by ABI1-PYR1-like () complexes followed by the inactivation of ABI1, in turn activating CIPK23. Under favorable growth conditions, ABI1 reduced AMT1;1 and AMT1;2 phosphorylation, both by binding and inactivating CIPK23. ABI1 further directly interacted with AMT1;1 and AMT1;2, which would be a prerequisite for dephosphorylation of the transporter by ABI1. Thus, ABI1 is a positive regulator of ammonium uptake, coupling nutrient acquisition to abiotic stress signaling. Elevated ABA reduces ammonium uptake during stress situations, such as ammonium toxicity, whereas ABI1 reactivates AMT1s under favorable growth conditions.

Genome-Wide Identification of the Salvia miltiorrhiza SmCIPK Gene Family and Revealing the Salt Resistance Characteristic of SmCIPK13

Members of the CIPK (CBL-interacting protein kinases) gene family play important roles in calcium (Ca2+) signaling pathway-regulated plant resistance to abiotic stresses. Salvia miltiorrhiza, which is widely planted and grown in complex and diverse environments, is mainly focused on the transcriptional regulation of enzyme genes related to the biosynthesis of its bioactive components. However, the excavation of the genes related to the resistance of S.miltiorrhiza and the involved signaling pathways have not been deeply studied. In this study, 20 SmCIPK genes were identified and classified into two families and five subfamilies by biochemical means. Sequence characteristics and conserved motif analysis revealed the conservation and difference of SmCIPK protein in plants. Expression pattern analysis showed that SmCIPKs were mainly expressed in flowers and roots, and more than 90% of gene expression was induced by SA (salicylic acid), and MeJA (methyl jasmonate). Furthermore, the expression level of SmCIPK13 could be significantly increased after stress treatment with NaCl. SmCIPK13 expression in yeast reduces sensitivity to salt, while overexpression of it in Arabidopsis has the same effect and was localized in the cytoplasm, cell membrane and nucleus. In conclusion, the identification of the SmCIPK gene family and the functional characterization of the SmCIPK13 gene provides the basis for clarification of key genes in the Ca2+ signaling pathway and abiotic stress in S.miltiorrhiza.

Identification of the CIPK-CBL family gene and functional characterization of CqCIPK14 gene under drought stress in quinoa

BackgroundCalcineurin-like Protein (CBL) and CBL interacting protein kinase (CIPK) play a key role in plant signal transduction and response to various environmental stimuli. Quinoa, as an important plant with high nutritional value, can meet the basic nutritional needs of human Cash crop, is also susceptible to abiotic stress. However, CBL-CIPK in quinoa have not been reported.ResultsIn this study, 16 CBL and 41 CIPK genes were identified in quinoa. CBL-CIPK gene shows different intron-exon gene structure and motif, they participate in different biological processes, and form a complex regulatory network between CBL-CIPK proteins. Many cis-regulatory element associated with ABA and drought have been found. The expression patterns of CBL-CIPK showed different expression patterns in various abiotic stresses and tissues. RT-qPCR showed that most members of these two gene families were involved in drought regulation of quinoa, in particular, the expression levels of CqCIPK11, CqCIPK15, CqCIPK37 and CqCBL13 increased significantly under drought stress.ConclusionsThe structures and functions of the CBL-CIPK family in quinoa were systematically explored. Many CBL-CIPK may play vital roles in the regulation of organ development, growth, and responses to abiotic stresses. This research has great significance for the functional characterisation of the quinoa CBL-CIPK family and our understanding of the CBL-CIPK family in higher plants.

GmCIPK21, a CBL-interacting protein kinase confers salt tolerance in soybean (Glycine max. L)

Salt stress severely affects plant development and yield. Calcineurin B-like protein interacting protein kinases (CIPKs) play a crucial role in plant adaptation to environmental challenges. However, the biological functions of CIPKs in soybean remain poorly understood. Here, we identified GmCIPK21, a salt-responsive CIPK gene from soybean. Overexpression of GmCIPK21 in Arabidopsis and soybean hairy roots led to increased salt tolerance. The hairy roots with GmCIPK21 suppression by RNA interference exhibited salt-sensitive phenotypes. Further physiological analysis revealed that GmCIPK21 reduced the content of hydrogen peroxide (H2O2) and malondialdehyde (MDA) and increased the activity of the antioxidant enzymes under salt stress. Additionally, GmCIPK21 was found to enhance the ABA sensitivity of transgenic plants. GmCIPK21 was also implicated in increasing the activation of antioxidant-, salt-, and ABA-related genes upon salt stress. Interestingly, GmCIPK21 interacted with GmCBL4, promoting the scavenging salt-induced reactive oxygen species (ROS). These results collectively suggested that GmCIPK21 affects ROS homeostasis and ABA response to improve salt tolerance in soybean.

CBL-Interacting Protein Kinase OsCIPK18 Regulates the Response of Ammonium Toxicity in Rice Roots.

Ammonium () is one of the major nitrogen sources for plants. However, excessive ammonium can cause serious harm to the growth and development of plants, i.e., ammonium toxicity. The primary regulatory mechanisms behind ammonium toxicity are still poorly characterized. In this study, we showed that OsCIPK18, a CBL-interacting protein kinase, plays an important role in response to ammonium toxicity by comparative analysis of the physiological and whole transcriptome of the T-DNA insertion mutant (cipk18) and the wild-type (WT). Root biomass and length of cipk18 are less inhibited by excess compared with WT, indicating increased resistance to ammonium toxicity. Transcriptome analysis reveals that OsCIPK18 affects the uptake by regulating the expression of OsAMT1;2 and other transporters, but does not affect ammonium assimilation. Differentially expressed genes induced by excess in WT and cipk18 were associated with functions, such as ion transport, metabolism, cell wall formation, and phytohormones signaling, suggesting a fundamental role for OsCIPK18 in ammonium toxicity. We further identified a transcriptional regulatory network downstream of OsCIPK18 under stress that is centered on several core transcription factors. Moreover, OsCIPK18 might function as a transmitter in the auxin and abscisic acid (ABA) signaling pathways affected by excess ammonium. These data allowed us to define an OsCIPK18-regulated/dependent transcriptomic network for the response of ammonium toxicity and provide new insights into the mechanisms underlying ammonium toxicity.

Insight into the CBL and CIPK gene families in pecan (Carya illinoinensis): identification, evolution and expression patterns in drought response

BackgroundCalcium (Ca2+) serves as a ubiquitous second messenger and plays a pivotal role in signal transduction. Calcineurin B-like proteins (CBLs) are plant-specific Ca2+ sensors that interact with CBL-interacting protein kinases (CIPKs) to transmit Ca2+ signals. CBL-CIPK complexes have been reported to play pivotal roles in plant development and response to drought stress; however, limited information is available about the CBL and CIPK genes in pecan, an important nut crop.ResultsIn the present study, a total of 9 CBL and 30 CIPK genes were identified from the pecan genome and divided into four and five clades based on phylogeny, respectively. Gene structure and distribution of conserved sequence motif analysis suggested that family members in the same clade commonly exhibited similar exon-intron structures and motif compositions. The segmental duplication events contributed largely to the expansion of pecan CBL and CIPK gene families, and Ka/Ks values revealed that all of them experienced strong negative selection. Phylogenetic analysis of CIPK proteins from 14 plant species revealed that CIPKs in the intron-poor clade originated in seed plants. Tissue-specific expression profiles of CiCBLs and CiCIPKs were analysed, presenting functional diversity. Expression profiles derived from RNA-Seq revealed distinct expression patterns of CiCBLs and CiCIPKs under drought treatment in pecan. Moreover, coexpression network analysis helped to elucidate the relationships between these genes and identify potential candidates for the regulation of drought response, which were verified by qRT–PCR analysis.ConclusionsThe characterization and analysis of CBL and CIPK genes in pecan genome could provide a basis for further functional analysis of CiCBLs and CiCIPKs in the drought stress response of pecan.

Genome-Wide Identification and Functional Analysis of the Calcineurin B-like Protein and Calcineurin B-like Protein-Interacting Protein Kinase Gene Families in Chinese Cabbage (Brassica rapa ssp. pekinensis).

In plants, calcineurin B-like proteins (CBL) are a unique set of calcium sensors that decode calcium signals by activating a plant-specific protein kinase family called CBL-interacting protein kinases (CIPKs). The CBL–CIPK family and its interacting complexes regulate plant responses to various environmental stimuli. Chinese cabbage (Brassica rapa ssp. pekinensis) is an important vegetable crop in Asia; however, there are no reports on the role of the CBLs–CIPKs’ signaling system in response to abiotic stress during cabbage growth. In this study, 18 CBL genes and 47 CIPK genes were identified from the Chinese cabbage genome. Expansion of the gene families was mainly due to tandem repeats and segmental duplication. An analysis of gene expression patterns showed that different duplicate genes exhibited different expression patterns in response to treatment with Mg2+, K+, and low temperature. In addition, differences in the structural domain sequences of NAF/FISL and interaction profiles in yeast two-hybrid assays suggested a functional divergence of the duplicate genes during the long-term evolution of Chinese cabbage, a result further validated by potassium deficiency treatment using trans-BraCIPK23.1/23.2/23.3 Arabidopsis thaliana. Our results provide a basis for studies related to the functional divergence of duplicate genes and in-depth studies of BraCBL–BraCIPK functions in Chinese cabbage.

Deciphering the Molecular Mechanisms of Chilling Tolerance in Lsi1-Overexpressing Rice.

Improving tolerance to low-temperature stress during the rice seedling stage is of great significance in agricultural science. In this study, using the low silicon gene 1 (Lsi1)-overexpressing (Dular-OE) and wild-type rice (Dular-WT), we showed that Lsi1 overexpression enhances chilling tolerance in Dular-OE. The overexpression of the Lsi1 increases silicon absorption, but it was not the main reason for chilling tolerance in Dular-OE. Instead, our data suggest that the overexpression of a Lsi1-encoding NIP and its interaction with key proteins lead to chilling tolerance in Dular-OE. Additionally, we show that the high-mobility group protein (HMG1) binds to the promoter of Lsi1, positively regulating its expression. Moreover, Nod26-like major intrinsic protein (NIP)’s interaction with α and β subunits of ATP synthase and the 14-3-3f protein was validated by co-immunoprecipitation (Co-IP), bimolecular fluorescent complementary (BiFC), and GST-pulldown assays. Western blotting revealed that the overexpression of NIP positively regulates the ATP-synthase subunits that subsequently upregulate calcineurin B-like interacting protein kinases (CIPK) negatively regulating 14-3-3f. Overall, these NIP-mediated changes trigger corresponding pathways in an orderly manner, enhancing chilling tolerance in Dular-OE.

AcCIPK5, a pineapple CBL-interacting protein kinase, confers salt, osmotic and cold stress tolerance in transgenic Arabidopsis

Plant-specific calcineurin B-like proteins (CBLs) and their interacting kinases, CBL-interacting protein kinases (CIPKs) module, are essential for dealing with various biotic and abiotic stress. The kinases (CIPKs) of this module have been well studied in several plants; however, the information about pineapple CIPKs remains limited. To understand how CIPKs function against environmental cues in pineapple, the CIPK5 gene of pineapple was cloned and characterized. The phylogenetic analyses revealed that AcCIPK5 is homologous to the CIPK12 of Arabidopsis and other plant species. Quantitative real-time PCR (qRT-PCR) analysis revealed that AcCIPK5 responds to multiple stresses, including osmotic, salt stress, heat and cold. Under optimal conditions, AcCIPK5 gets localized to the cytoplasm and cell membrane. The ectopic expression of AcCIPK5 in Arabidopsis improved the germination under osmotic and salt stress. Furthermore, AcCIPK5 positively regulated osmotic, drought, salt and cold tolerance and negatively regulated heat and fungal stress in Arabidopsis. Besides, the expression of AcCIPK impacted ABA-related genes and ROS homeostasis. Overall, the present study demonstrates that AcCIPK5 contributes to multiple stress tolerance and has the potential to be utilized in the development of stress-tolerant crops.

Signal Transduction in Cereal Plants Struggling with Environmental Stresses: From Perception to Response.

Cereal plants under abiotic or biotic stressors to survive unfavourable conditions and continue growth and development, rapidly and precisely identify external stimuli and activate complex molecular, biochemical, and physiological responses. To elicit a response to the stress factors, interactions between reactive oxygen and nitrogen species, calcium ions, mitogen-activated protein kinases, calcium-dependent protein kinases, calcineurin B-like interacting protein kinase, phytohormones and transcription factors occur. The integration of all these elements enables the change of gene expression, and the release of the antioxidant defence and protein repair systems. There are still numerous gaps in knowledge on these subjects in the literature caused by the multitude of signalling cascade components, simultaneous activation of multiple pathways and the intersection of their individual elements in response to both single and multiple stresses. Here, signal transduction pathways in cereal plants under drought, salinity, heavy metal stress, pathogen, and pest attack, as well as the crosstalk between the reactions during double stress responses are discussed. This article is a summary of the latest discoveries on signal transduction pathways and it integrates the available information to better outline the whole research problem for future research challenges as well as for the creative breeding of stress-tolerant cultivars of cereals.

Overexpression of <italic>ZmCIPKHT</italic> enhances heat tolerance in plant

<p id="C3">The effects of high temperature stress on the normal growth and yield of plants is more and more serious. To adapt the changes of external environment, plants have evolved a series of molecular genetic mechanisms to respond to high temperature stress. The calcineurin B-like protein (CBL) interacting protein kinase (CIPK) is actively involved in response to high temperature stress depended on ABA signal transduction pathway in plants. Based on previous genome-wide association analysis, a candidate gene <italic>ZmCIPKHT</italic> related to maize high temperature tolerance was cloned in this study. Real-time quantitative PCR results showed that <italic>ZmCIPKHT</italic> gene was significantly induced by high temperature stress. Transient transformation of maize protoplasts revealed that ZmCIPKHT was localized in the nucleus. Overexpressing <italic>ZmCIPKHT</italic> plants of transgenic <italic>Arabidopsis thaliana</italic> had significantly higher survival rate and better growth status than wild type under high temperature stress. The yeast two-hybrid experiment confirmed that the interaction between ZmCIPKHT protein and ZmCBL4 protein in maize CBLs family. The relative expression levels of genes related to abscisic acid (ABA) pathway in transgenic <italic>Arabidopsis thaliana</italic> with <italic>ZmCIPKHT</italic> under high temperature stress were changed accordingly, indicating that the regulations of <italic>ZmCIPKHT</italic> genes under high temperature stress were in the ABA-dependent pathway. These results provide a new experimental basis for elucidating the molecular mechanism of maize CBL-CIPK signaling pathway dependent on ABA pathway to abiotic stress in plants.

CmNF-YB8 affects drought resistance in chrysanthemum by altering stomatal status and leaf cuticle thickness.

Drought is a major abiotic stress that limits plant growth and development. Adaptive mechanisms have evolved to mitigate drought stress, including the capacity to adjust water loss rate and to modify the morphology and structure of the epidermis. Here, we show that the expression of CmNF-YB8, encoding a nuclear factor Y (NF-Y) B-type subunit, is lower under drought conditions in chrysanthemum (Chrysanthemum morifolium). Transgenic chrysanthemum lines in which transcript levels of CmNF-YB8 were reduced by RNA interference (CmNF-YB8-RNAi) exhibited enhanced drought resistance relative to control lines, whereas lines overexpressing CmNF-YB8 (CmNF-YB8-OX) were less tolerant to drought. Compared to wild type (WT), CmNF-YB8-RNAi plants showed reduced stomatal opening and a thicker epidermal cuticle that correlated with their water loss rate. We also identified genes involved in stomatal adjustment (CBL-interacting protein kinase 6, CmCIPK6) and cuticle biosynthesis (CmSHN3) that are more highly expressed in CmNF-YB8-RNAi lines than in WT, CmCIPK6 being a direct downstream target of CmNF-YB8. Virus-induced gene silencing of CmCIPK6 or CmSHN3 in the CmNF-YB8-RNAi background abolished the effects of CmNF-YB8-RNAi on stomatal closure and cuticle deposition, respectively. CmNF-YB8 thus regulates CmCIPK6 and CmSHN3 expression to alter stomatal movement and cuticle thickness in the leaf epidermis, thereby affecting drought resistance.

The Calcium Sensor Calcineurin B-Like Proteins -Calcineurin B-Like Interacting Protein Kinases Is Involved in Leaf Development and Stress Responses Related to Latex Flow in Hevea brasiliensis.

Latex flow in Hevea brasiliensis (the Para rubber tree), the sole commercial source of natural rubber (cis-1,4-polyisoprene, NR), renders it uniquely suited for the study of plant stress responses. Calcineurin B-like interacting protein kinases (CIPK) serving as calcium-sensor protein kinases react with calcineurin B-like proteins (CBL) to play crucial roles in hormone signaling transduction and response to abiotic stress in plant developmental processes. However, little is known about their functions in Hevea. In this study, a total of twelve CBL (HbCBL) and thirty CIPK (HbCIPK) genes were identified from the Hevea genome. Structure and phylogenetic analysis assigned these CIPKs to five groups and CBLs to four groups, and mapped onto fourteen of the eighteen Hevea chromosomes. RNA-seq and qPCR analysis showed that the expressions of HbCBL and HbCIPK genes varied in the seven Hevea tissues examined, i.e., latex (cytoplasm of rubber-producing laticifers), bark, leaf, root, seed, female flower, and male flower. The expressions of two HbCBL and sixteen HbCIPK genes showed upward trends during leaf development. Following ethylene yield stimulation and the latex tapping treatment, both practices invoking stress, the expression levels of most latex-expressed genes were significantly altered. Yeast two-hybrid test revealed interactions for multiple combinations of HbCBLs and HbCIPKs with substantial gene expression in latex or other Hevea tissues. However, all the HbCBL-HbCIPK complexes examined did not recruit HbSOS1 or AtSOS1 to form functional salt tolerance SOS pathway in yeast cells. Taken together, the results suggested a role of the Hevea CBL-CIPK network as a point of convergence for several different signaling pathways in growth, development, and stress responses in relation to latex production.

Analysis of gene expression in early seed germination of rice: landscape and genetic regulation

BackgroundSeed germination is a crucial process, which determines the initiation of seed plant life cycle. The early events during this important life cycle transition that called early seed germination is defined as initially water uptake plus radicle growing out of the covering seed layers. However, a specific genome-wide analysis of early seed germination in rice is still obscure.ResultsIn this study, the physiological characteristics of rice seed during seed germination are determined to define key points of early seed germination. Transcriptome analyses of early phase of seed germination provided deeper insight into the genetic regulation landscape. Many genes involved in starch-to-sucrose transition were differentially expressed, especially alpha-amylase 1b and beta-amylase 2, which were predominantly expressed. Differential exon usage (DEU) genes were identified, which were significantly enriched in the pathway of starch and sucrose metabolism, indicating that DEU events were critical for starch-to-sucrose transition at early seed germination. Transcription factors (TFs) were also dramatic expressed, including the abscisic acid (ABA) responsive gene, OsABI5, and gibberellic acid (GA) responsive genes, GAI. Moreover, GAI transactivated GA responsive gene, GAMYB in vivo, indicating a potential pathway involved in early seed germination process. In addition, CBL-interacting protein kinase (CIPK) genes, such as CIPK13, CIPK14 and CIPK17 were potentially interacted with other proteins, indicating its pivotal role at early seed germination.ConclusionTaken together, gene regulation of early seed germination in rice was complex and protein-to-gene or protein-to-protein interactions were indispensable.

GR24-mediated enhancement of salt tolerance and roles of H2O2 and Ca2+ in regulating this enhancement in cucumber

This study investigated the regulation of the exogenous strigolactone (SL) analog GR24 in enhancing the salt tolerance and the effects of calcium ion (Ca2+) and hydrogen peroxide (H2O2) on GR24's regulation effects in cucumber. The seedlings were sprayed with (1) distilled water (CK), (2) NaCl, (3) GR24, then NaCl, (4) GR24, then H2O2 scavenger, then NaCl, and (5) GR24, then Ca2+ blocker, then NaCl. The second true leaf was selected for biochemical assays. Under the salt stress, the exogenous GR24 maintained the ion balance, increased the activity of antioxidant enzymes, reduced the membrane lipid peroxidation, and increased the activities of catalase (CAT), superoxide dismutase (SOD), peroxidase (POD), and ascorbate peroxidase (APX), accompanied by a decrease in relative conductivity, an increase in the proline content, and elevated gene expression levels of antioxidant enzymes, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, calcium-dependent protein kinases (CDPKs), salt overly sensitive SOS1, CBL-interacting protein kinase 2 (CIPK2), and calcineurin B-like protein 3 (CBL3). Such protective effects triggered by GR24 were attenuated or almost abolished by ethylene glycol tetraacetic acid (EGTA), lanthanum chloride (LaCl3, Ca2+ channel blocker), diphenyleneiodonium (DPI, NADPH oxidase inhibitor), and dimethylthiourea (DMTU, hydroxyl radical scavenger). Our data suggest that exogenous GR24 is highly effective in alleviating salt-induced damages via modulating antioxidant capabilities and improving ionic homeostasis and osmotic balance and that H2O2 and Ca2+ are required for GR24-mediated enhancement of salt tolerance.

A CBL-interacting protein kinase, AcCIPK18, from Ananas comosus regulates tolerance to salt, drought, heat stress and Sclerotinia sclerotiorum infection in Arabidopsis

In plants, calcineurin B-like proteins (CBLs) and their interacting kinase, CBL-interacting protein kinases (CIPKs), perform essential roles in mediating responses to various extracellular cues, including biotic and abiotic stresses. Several CIPK genes have been functionally characterized in different plant species. However, to date, the functions of pineapple CIPKs have not been investigated, which limits their potential use in molecular breeding. Here we functionally characterized an AcCIPK18 of pineapple and verified its role in salt, drought, heat stresses and immunity. Bioinformatics studies and phylogenetic analysis showed that AcCIPK18 is homologous to CIPK6 of Arabidopsis. Expression analysis shows that AcCIPK18 gets induced by salt, osmotic, heat and ABA. Besides, the ectopic expression of AcCIPK18 increased tolerance to salt, drought and heat stress but displayed sensitivity against fungal infection in Arabidopsis. We found that overexpression of AcCIPK18 changes the ROS homeostasis by altering the expression of several ABA signaling genes. These results provide evidence for the involvement of AcCIPK18 in multiple stress responses and advocates that AcCIPK18 could be used as a potential candidate gene for improving tolerance against multiple abiotic stress.