Genome-wide analysis of the KNOX gene family in Moso bamboo: insights into their role in promoting the rapid shoot growth.

BACKGROUND: KNOTTED1-like homeobox (KNOX) genes, plant-specific homologous box transcription factors (TFs), play a central role in regulating plant growth, development, organ formation, and response to biotic and abiotic stresses. However, a comprehensive genome-wide identification of the KNOX genes in Moso bamboo (Phyllostachys edulis), the fastest growing plant, has not yet been conducted, and the specific biological functions of this family remain unknown. RESULTS: The expression profiles of 24 KNOX genes, divided into two subfamilies, were determined by integrating Moso bamboo genome and its transcriptional data. The KNOX gene promoters were found to contain several light and stress-related cis-acting elements. Synteny analysis revealed stronger similarity with rice KNOX genes than with Arabidopsis KNOX genes. Additionally, several conserved structural domains and motifs were identified in the KNOX proteins. The expansion of the KNOX gene family was primarily regulated by tandem duplications. Furthermore, the KNOX genes were responsive to naphthaleneacetic acid (NAA) and gibberellin (GA) hormones, exhibiting distinct temporal expression patterns in four different organs of Moso bamboo. Short Time-series Expression Miner (STEM) analysis and quantitative real-time PCR (qRT-PCR) assays demonstrated that PeKNOX genes may play a role in promoting rapid shoot growth. Additionally, Gene Ontology (GO) and Protein-Protein Interaction (PPI) network enrichment analyses revealed several functional annotations for PeKNOXs. By regulating downstream target genes, PeKNOXs are involved in the synthesis of AUX /IAA, ultimately affecting cell division and elongation. CONCLUSIONS: In the present study, we identified and characterized a total of 24 KNOX genes in Moso bamboo and investigated their physiological properties and conserved structural domains. To understand their functional roles, we conducted an analysis of gene expression profiles using STEM and RNA-seq data. This analysis successfully revealed regulatory networks of the KNOX genes, involving both upstream and downstream genes. Furthermore, the KNOX genes are involved in the AUX/IAA metabolic pathway, which accelerates shoot growth by influencing downstream target genes. These results provide a theoretical foundation for studying the molecular mechanisms underlying the rapid growth and establish the groundwork for future research into the functions and transcriptional regulatory networks of the KNOX gene family.

MrERF039 transcription factor plays an active role in the cold response of Medicago ruthenica as a sugar molecular switch.

Cold stress severely restricts plant development, causing significant agricultural losses. We found a critical transcription factor network in Medicago ruthenica was involved in plant adaptation to low-temperature. APETALA2/ethylene responsive factor (AP2/ERF) transcription factor MrERF039 was transcriptionally induced by cold stress in M. ruthenica. Overexpression of MrERF039 significantly increased the glucose and maltose content, thereby improving the tolerance of M. ruthenica. MrERF039 could bind to the DRE cis-acting element in the MrCAS15A promoter. Additionally, the methyl group of the 14th amino acid in MrERF039 was required for binding. Transcriptome analysis showed that MrERF039 acted as a sugar molecular switch, regulating numerous sugar transporters and sugar metabolism-related genes. In addition, we found that MrERF039 could directly regulate ß-amylase gene, UDP glycosyltransferase gene, and C2H2 zinc finger protein gene expression. In conclusion, these findings suggest that high expression of MrERF039 can significantly improve the cold tolerance of M. ruthenica root tissues during cold acclimation. Our results provide a new theoretical basis and candidate genes for breeding new legume forage varieties with high resistance.

Genome-wide identification and expression pattern analysis of the GRF transcription factor family in Astragalus mongholicus.

BACKGROUND: Astragalus membranaceus is a plant of the Astragalus genus, which is used as a traditional Chinese herbal medicine with extremely high medicinal and edible value. Astragalus mongholicus, as one of the representative medicinal materials with the same origin of medicine and food, has a rising market demand for its raw materials, but the quality is different in different production areas. Growth-regulating factors (GRF) are transcription factors unique to plants that play important roles in plant growth and development. Up to now, there is no report about GRF in A. mongholicus. METHODS AND RESULTS: This study conducted a genome-wide analysis of the AmGRF gene family, identifying a total of nine AmGRF genes that were classified into subfamily V based on phylogenetic relationships. In the promoter region of the AmGRF gene, we successfully predicted cis-elements that respond to abiotic stress, growth, development, and hormone production in plants. Based on transcriptomic data and real-time quantitative polymerase chain reaction (qPCR) validation, the results showed that AmGRFs were expressed in the roots, stems, and leaves, with overall higher expression in leaves, higher expression of AmGRF1 and AmGRF8 in roots, and high expression levels of AmGRF1 and AmGRF9 in stems. CONCLUSIONS: The results of this study provide a theoretical basis for the further exploration of the functions of AmGRFs in plant growth and development.

Detection of Transversions and Transitions in HBG2 Cis-Elements Associated with Sickle Cell Allele in Ghanaians.

Short tandem repeats located 5' prime to the ß-globin gene, have been observed to be in linkage disequilibrium with the HbS allele, and thought to affect the severity of sickle cell disease. Here, we report on new mutants within the HBG2 region that may impact sickle cell disease. To determine the cis-acting elements microsatellites, indels and single nucleotide polymorphisms (SNPs), within the HBG2 region by sequencing, in subjects with sickle cell disease. The case-control study was located at the Center for Clinical Genetics, Sickle cell unit, Korle-Bu Teaching Hospital. A questionnaire was used for demographic data and clinical information. Hematological profile (red blood cell, white blood cell, platelet, hemoglobin and mean corpuscular volume) were assessed in 83 subjects. A set of 45 samples comprising amplified DNA on the HBG2 gene from HbSS (22), HbSC (17) and 6 controls (HbAA) were sequenced. Differences in the microsatellite region between sickle cell disease (SCD) (HbSS and HbSC) genotypes and control subjects were identified by counting and assessed by Chi-square analysis. Red blood cells, hematocrit, platelets, white blood cells and hemoglobin indices differed in genotypic groups. HbSS subjects were affirmed to have severer hemolytic anemia than HbSC subjects. Two indels (T1824 and C905) were seen in both SS and SC genotypes. Two peculiar SNPs: G:T1860 (transition) and A:G1872 transversions were found within the HBG2 gene that were significantly associated with the HbSS genotype (Fisher's exact test, p = 0.006) and HbS allele respectively (Fisher's exact test, p = 0.006). Cis-acting elements in HbSS and HbSC were different and may contribute to the phenotype seen in the disease state.

Whole-genome identification and expression profiling of growth-regulating factor (GRF) and GRF-interacting factor (GIF) gene families in Panax ginseng.

BACKGROUND: Panax ginseng is a perennial herb and one of the most widely used traditional medicines in China. During its long growth period, it is affected by various environmental factors. Past studies have shown that growth-regulating factors (GRFs) and GRF-interacting factors (GIFs) are involved in regulating plant growth and development, responding to environmental stress, and responding to the induction of exogenous hormones. However, GRF and GIF transcription factors in ginseng have not been reported. RESULTS: In this study, 20 GRF gene members of ginseng were systematically identified and found to be distributed on 13 chromosomes. The ginseng GIF gene family has only ten members, which are distributed on ten chromosomes. Phylogenetic analysis divided these PgGRFs into six clades and PgGIFs into two clades. In total, 18 of the 20 PgGRFs and eight of the ten PgGIFs are segmental duplications. Most PgGRF and PgGIF gene promoters contain some hormone- and stress- related cis-regulatory elements. Based on the available public RNA-Seq data, the expression patterns of PgGRF and PgGIF genes were analysed from 14 different tissues. The responses of the PgGRF gene to different hormones (6-BA, ABA, GA3, IAA) and abiotic stresses (cold, heat, drought, and salt) were studied. The expression of the PgGRF gene was significantly upregulated under GA3 induction and three weeks of heat treatment. The expression level of the PgGIF gene changed only slightly after one week of heat treatment. CONCLUSIONS: The results of this study may be helpful for further study of the function of PgGRF and PgGIF genes and lay a foundation for further study of their role in the growth and development of Panax ginseng.

In situ deletions reveal regulatory components for expression of an intracellular immune receptor gene and its co-expressed genes in Arabidopsis.

Intracellular immune receptor nucleotide-binding leucine-rich repeats (NLRs) are highly regulated transcriptionally and post-transcriptionally for balanced plant defence and growth. NLR genes often exist in gene clusters and are usually co-expressed under various conditions. Despite of intensive studies of regulation of NLR proteins, cis-acting elements for NLR gene induction, repression or co-expression are largely unknown due to a larger than usual cis-region for their expression regulation. Here we used the CRISPR/Cas9 genome editing technology to generate a series of in situ deletions at the endogenous location of a NLR gene SNC1 residing in the RPP5 gene cluster. These deletions that made in the wild type and the SNC1 constitutive expressing autoimmune mutant bon1 revealed both positive and negative cis-acting elements for SNC1 expression. Two transcription factors that could bind to these elements were found to have an impact on the expression of SNC1. In addition, co-expression of two genes with SNC1 in the same cluster is found to be mostly dependent on the SNC1 function. Therefore, SNC1 expression is under complex local regulation involving multiple cis elements and SNC1 itself is a critical regulator of gene expression of other NLR genes in the same gene cluster.

Genome-wide analysis of glutamate receptor gene family in allopolyploid Brassica napus and its diploid progenitors.

Ionotropic glutamate receptors are ligand-gated nonselective cation channels that mediate neurotransmission in the central nervous system of animals. Plants possess homologous proteins called glutamate receptor-like channels (GLRs) which are involved in vital physiological processes including seed germination, long-distance signaling, chemotaxis, Ca2+ signaling etc. Till now, a comprehensive genome-wide analysis of the GLR gene family members in different economically important species of Brassica is missing. Considering the origin of allotetraploid Brassica napus from the hybridization between the diploid Brassica oleracea and Brassica rapa, we have identified 11, 27 and 65 GLR genes in B. oleracea, B. rapa and B. napus, respectively showing an expansion of this gene family in B. napus. Chromosomal locations revealed several tandemly duplicated GLR genes in all the three species. Moreover, the gene family expanded in B. napus after allopolyploidization. The phylogenetic analysis showed that the 103 GLRs are classified into three main groups. The exon-intron structures of these genes are not very conserved and showed wide variation in intron numbers. However, protein sequences are much conserved as shown by the presence of ten short amino acid sequence motifs. Predicted cis-acting elements in 1 kb promoters of GLR genes are mainly involved in light, stress and hormone responses. RNA-seq analysis showed that in B. oleracea and B. rapa, some GLRs are more tissue specific than others. In B. napus, some GLRs are downregulated under cold stress, while others are upregulated. In summary, this bioinformatic study of the GLR gene family of the three Brassica species provides evidence for the expansion of this gene family in B. napus and also provided useful information for in-depth studies of their biological functions in Brassica.

Genome wide identification and evolutionary analysis of vat like NBS-LRR genes potentially associated with resistance to aphids in cotton.

Nucleotide Binding Site - Leucine Rich Repeat (NBS-LRR) genes play a significant role in plant defense against biotic stresses and are an integral part of signal transduction pathways. Vat gene has been well reported for their role in resistance to Aphis gossypii and viruses transmitted by them. Despite their importance, Vat like NBS-LRR resistance genes have not yet been identified and studied in cotton species. This study report hundreds of orthologous Vat like NBS-LRR genes from the genomes of 18 cotton species through homology searches and the distribution of those identified genes were tend to be clustered on different chromosome. Especially, in a majority of the cases, Vat like genes were located on chromosome number 13 and they all shared two conserved NBS-LRR domains, one disease resistant domain and several repeats of LRR on the investigated cotton Vat like proteins. Gene ontology study on Vat like NBS-LRR genes revealed the molecular functions viz., ADP and protein binding. Phylogenetic analysis also revealed that Vat like sequences of two diploid species, viz., G. arboreum and G. anomalum, were closely related to the sequences of the tetraploids than all other diploids. The Vat like genes of G. aridum and G. schwendimanii were distantly related among diploids and tetraploids species. Various hormones and defense related cis-acting regulatory elements were identified from the 2 kb upstream sequences of the Vat like genes implying their defensive response towards the biotic stresses. Interestingly, G. arboreum and G. trilobum were found to have more regulatory elements than larger genomes of tetraploid cotton species. Thus, the present study provides the evidence for the evolution of Vat like genes in defense mechanisms against aphids infestation in cotton genomes and allows further characterization of candidate genes for developing aphid and aphid transmitted viruses resistant crops through cotton breeding.

Epigenetic modifications and miRNAs determine the transition of somatic cells into somatic embryos.

KEY MESSAGE: This review discusses the epigenetic changes during somatic embryo (SE) development, highlights the genes and miRNAs involved in the transition of somatic cells into SEs as a result of epigenetic changes, and draws insights on biotechnological opportunities to study SE development. Somatic embryogenesis from somatic cells occurs in a series of steps. The transition of somatic cells into somatic embryos (SEs) is the most critical step under genetic and epigenetic regulations. Major regulatory genes such as SERK, WUS, BBM, FUS3/FUSA3, AGL15, and PKL, control SE steps and development by turning on and off other regulatory genes. Gene transcription profiles of somatic cells during SE development is the result of epigenetic changes, such as DNA and histone protein modifications, that control and decide the fate of SE formation. Depending on the type of somatic cells and the treatment with plant growth regulators, epigenetic changes take place dynamically. Either hypermethylation or hypomethylation of SE-related genes promotes the transition of somatic cells. For example, the reduced levels of DNA methylation of SERK and WUS promotes SE initiation. Histone modifications also promote SE induction by regulating SE-related genes in somatic cells. In addition, miRNAs contribute to the various stages of SE by regulating the expression of auxin signaling pathway genes (TIR1, AFB2, ARF6, and ARF8), transcription factors (CUC1 and CUC2), and growth-regulating factors (GRFs) involved in SE formation. These epigenetic and miRNA functions are unique and have the potential to regenerate bipolar structures from somatic cells when a pluripotent state is induced. However, an integrated overview of the key regulators involved in SE development and downstream processes is lacking. Therefore, this review discusses epigenetic modifications involved in SE development, SE-related genes and miRNAs associated with epigenetics, and common cis-regulatory elements in the promoters of SE-related genes. Finally, we highlight future biotechnological opportunities to alter epigenetic pathways using the genome editing tool and to study the transition mechanism of somatic cells.

Bioinformatics analysis and function prediction of NBS-LRR gene family in Broussonetia papyrifera.

Most of the currently available disease resistance (R) genes have NBS (nucleotide-binding site) and LRR (leucine-rich-repeat) domain which belongs to the NBS-LRR gene family. The whole genome sequencing of Broussonetia papyrifera provides an important bioinformatics database for the study of the NBS-LRR gene family. In this study, 328 NBS-LRR family genes were identified and classified in B. papyrifera according to different classification schemes, where there are 92 N types, 47 CN type, 54 CNL type, 29 NL types, 55 TN type, and 51 TNL type. Subsequently, we conducted bioinformatics analysis of the NBS-LRR gene family. Classification, motif analysis of protein sequences, and phylogenetic tree studies of the NBS-LRR genes in B. papyrifera provide important basis for the functional study of NBS-LRR family genes. Additionally, we performed structural analysis of the chromosomal location, physicochemical properties, and sequences identified by genetic characterization. In addition, through the analysis of GO enrichment, it was found that NBS-LRR genes were involved in defense responses and were significantly enriched in biological stimulation, immune response, and abiotic stress. In addition, we found that Bp06g0955 was the most sensitive to low temperature and encoded the RPM1 protein by analyzing the low temperature transcriptome data of B. papyrifera. Quantitative results of gene expression after 48 h of Fusarium infection showed that Bp01g3293 increased 14 times after infection, which encodes RPM1 protein. The potential of NBS-LRR gene responsive to biotic and abiotic stresses can be exploited to improve the resistance of B. papyrifera.

Characterization of myo-inositol oxygenase from rice (OsMIOX): influence of salinity stress in different indica rice cultivars.

Myo-inositol oxygenase (MIOX), the only catabolic enzyme of the inositol pathway, catalyzes conversion of myo-inositol to D-GlcA (glucuronic acid). The present study encompasses bioinformatic analysis of MIOX gene across phylogenetically related plant lineages and representative animal groups. Comparative motif analysis of the MIOX gene(s) across various plant groups suggested existence of abiotic- stress related cis-acting elements such as, DRE, MYB, MYC, STRE, MeJa among others. A detailed analysis revealed a single isoform of MIOX gene, located in chromosome 6 of indica rice (Oryza sativa) with an open reading frame of 938 bp coding for 308 amino acids producing a protein of ~ 35 kD. Secondary structure prediction of the protein gave the predicted number of 144 alpha helices and 154 random coils. The three-dimensional structure suggested it to be a monomeric protein with a single domain. Bacterial overexpression of the protein, purification and enzyme assay showed optimal catalytic activity at pH 7.5-8 at an optimal temperature of 37 °C with Michaelis constant of 40.92 mM. The range of Km was determined as 22.74-28.7 mM and the range of Vmax was calculated as 3.51-3.6 µM/min, respectively. Four salt-tolerant and salt-sensitive rice cultivars displayed differential gene expression of OsMIOX at different time points in different tissues under salinity and drought stress as observed from qRT-PCR data, microarray results and protein expression profile in immunoblot analysis. Gel volumetric analysis confirmed a very high expression of MIOX in roots and leaves on 7th day following germination. Microarray data showed high expression of MIOX at all developmental stages including seedling growth and reproduction. These data suggest that OsMIOX might have a role to play in rice abiotic stress responses mediated through the myo-inositol oxidation pathway. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01340-6.

RAD gene family analysis in cotton provides some key genes for flowering and stress tolerance in upland cotton G. hirsutum.

BACKGROUND: RADIALIS (RAD), belongs to the MYB gene family and regulates a variety of functions including floral dorsoventral asymmetry in Antirrhinum majus and development of fruit proteins in Solanum lycopersicum. RAD genes contain an SNF2_N superfamily domain. Here, we comprehensively identified 68 RAD genes from six different species including Arabidopsis and five species of cotton. RESULTS: Phylogenetic analysis classified RAD genes into five groups. Gene structure, protein motifs and conserved amino acid residues indicated that GhRAD genes were highly conserved during the evolutionary process. Chromosomal location information showed that GhRAD genes were distributed unevenly on different chromosomes. Collinearity and selection pressure analysis indicated RAD gene family expansion in G. hirsutum and G. barbadense with purifying selection pressure. Further, various growth and stress related promotor cis-acting elements were observed. Tissue specific expression level indicated that most GhRAD genes were highly expressed in roots and flowers (GhRAD2, GhRAD3, GhRAD4 and GhRAD11). Next, GhRAD genes were regulated by phytohormonal stresses (JA, BL and IAA). Moreover, Ghi-miRN1496, Ghi-miR1440, Ghi-miR2111b, Ghi-miR2950a, Ghi-miR390a, Ghi-miR390b and Ghi-miR7495 were the miRNAs targeting most of GhRAD genes. CONCLUSIONS: Our study revealed that RAD genes are evolutionary conserved and might be involved in different developmental processes and hormonal stress response. Data presented in our study could be used as the basis for future studies of RAD genes in cotton.

In situ deletions reveal regulatory components for expression of an intracellular immune receptor gene and its co-expressed genes in Arabidopsis.

Intracellular immune receptor nucleotide-binding leucine-rich repeats (NLRs) are highly regulated transcriptionally and post-transcriptionally for balanced plant defense and growth. NLR genes often exist in gene clusters and are usually co-expressed under various conditions. Despite intensive studies of the regulation of NLR proteins, cis-acting elements for NLR gene induction, repression or co-expression are largely unknown due to a larger than usual cis-region for their expression regulation. Here we used the CRISPR/Cas9 genome editing technology to generate a series of in situ deletions at the endogenous location of an NLR gene SNC1 residing in the RPP5 gene cluster. These deletions that made in the wild type and the SNC1 constitutive expressing autoimmune mutant bon1 revealed both positive and negative cis-acting elements for SNC1 expression. Two transcription factors that could bind to these elements were found to have an impact on the expression of SNC1. In addition, co-expression of two genes with SNC1 in the same cluster is found to be mostly dependent on the SNC1 function. Therefore, SNC1 expression is under complex local regulation involving multiple cis-elements and SNC1 itself is a critical regulator of gene expression of other NLR genes in the same gene cluster.

Comprehensive analysis and expression profiles of cassava UDP-glycosyltransferases (UGT) family reveal their involvement in development and stress responses in cassava.

UDP-glycosyltransferases (UGTs) are widely involved in plant growth and stress responses. However, UGT family are not well understood in cassava. Here, we identified 121 MeUGT genes and classified them into 14 subfamilies by phylogenetic analysis. All MeUGT proteins have typical feature of the UGTs family. Tandem duplications are the crucial driving force for the expansion of MeUGT family. Cis-Acting elements analysis uncovered those 14 kinds of cis-elements associated with biotic and abiotic stress responses. Transcriptomic and qRT-PCR analyses indicated that MeUGT genes participate in postharvest physiological deterioration of storage root and the responses of biotic and abiotic stresses. Of which, MeUGT-14/41 were significantly induced after Xam treatment. Silencing of MeUGT-14 or MeUGT-41 reduced cassava resistance to Xam, verifying the accuracy of transcriptomic data for function prediction. Together, this study characterized the MeUGTs family and revealed their potential functions, which build a solid foundation for MeUGTs associated genetic improvement of cassava.

Molecular characterization of miRNA genes and their expression in Dimocarpus longan Lour.

MAIN CONCLUSION: A genome-wide analysis of longan miRNA genes was conducted, and full-length pri-miRNA transcripts were cloned. Bioinformatics and expression analyses contributed to the functional characterization of longan miRNA genes. MicroRNAs are important for the post-transcriptional regulation of target genes. However, little is known about the transcription and regulation of miRNA genes in longan (Dimocarpus longan Lour.). In this study, 80 miRNA precursors (pre-miRNA) were predicted, and their secondary structure, size, conservation, and diversity were analyzed. Furthermore, the full-length cDNA sequences of 13 longan primary miRNAs (pri-miRNAs) were amplified by RLM-RACE and SMART-RACE and analyzed, which revealed that longan pri-miRNA transcripts have multiple transcription start sites (TSSs) and the downstream pre-miRNAs are polymorphic. Accordingly, the longan pri-miRNAs and protein-encoding genes may have similar transcriptional specificities. An analysis of the longan miRNA gene promoter elements indicated that the three most abundant cis-acting elements were light-responsive, stress-responsive, and hormone-responsive elements. A quantitative real-time PCR assay elucidated the potential spatial and temporal expression patterns of longan pre-miRNAs during the early stages of somatic embryogenesis (SE) and in different longan organs/tissues. This is the first report regarding the molecular characterization of miRNA genes and their expression profiles in longan. The generated data may serve as a foundation for future research aimed at clarifying the longan miRNA gene functions.

Genome-wide identification and expression analysis of GL2-interacting-repressor (GIR) genes during cotton fiber and fuzz development.

MAIN CONCLUSION: GL2-interacting-repressor (GIR) family members may contribute to fiber/fuzz formation via a newly discovered unique pathway in Gossypium arboreum. There are similarities between cotton fiber development and the formation of trichomes and root hairs. The GL2-interacting-repressors (GIRs) are crucial regulators of root hair and trichome formation. The GaFzl gene, annotated as GaGIR1, is negatively associated with trichome development and fuzz initiation. However, there is relatively little available information regarding the other GIR genes in cotton, especially regarding their effects on cotton fiber development. In this study, 21 GIR family genes were identified in the diploid cotton species Gossypium arboreum; these genes were divided into three groups. The GIR genes were characterized in terms of their phylogenetic relationships, structures, chromosomal distribution and evolutionary dynamics. These GIR genes were revealed to be unequally distributed on 12 chromosomes in the diploid cotton genome, with no GIR gene detected on Ga06. The cis-acting elements in the promoter regions were predicted to be responsive to light, phytohormones, defense activities and stress. The transcriptomic data and qRT-PCR results revealed that most GIR genes were not differentially expressed between the wild-type control and the fuzzless mutant line. Moreover, 14 of 21 family genes were expressed at high levels, indicating these genes may play important roles during fiber development and fuzz formation. Furthermore, Ga01G0231 was predominantly expressed in root samples, suggestive of a role in root hair formation rather than in fuzz initiation and development. The results of this study have enhanced our understanding of the GIR genes and their potential utility for improving cotton fiber through breeding.

Genome-Wide Investigation and Expression Analysis of K+-Transport-Related Gene Families in Chinese Cabbage (Brassica rapa ssp. pekinensis).

Potassium (K+) transport and channel systems play vital roles in plant growth, development and responses to various stresses. In this study, 44 putative K+-transport-related genes (18K+ uptake permease (KUP)/high-affinity K+ (HAK)/K+ transporter (KT) family genes and 26 channel genes, including 18 Shaker family genes and 8K+ channel outward (KCO) family genes) were identified in the genome of Chinese cabbage (Brassica rapa ssp. pekinensis). To clarify the molecular evolution of each family in Chinese cabbage, phylogenetic analysis and assessments of the gene structures, conserved motifs, chromosomal locations, gene duplications, expression patterns and cis-acting elements of the 44 putative K+-transport-related genes were performed. The phylogenetic analysis showed that these genes could be classified into five clades [KUP/HAK/KTs, KCOs, Kout, Kin (KAT) and Kin (AKT)] and that the members of a given clade shared conserved exon-intron distributions and motif compositions. These K+-transport-related genes were unevenly distributed over all ten chromosomes, including four duplicated gene pairs that implied an expansion of K+-transport-related genes in Chinese cabbage. Analyses of Illumina RNA-seq data for these 44K+-transport-related genes indicated tissue-/organ-specific expression patterns. In addition, an overall evaluation showed that the expression levels of KUP/HAK/KT genes were significantly higher than those of potassium channel genes in six tissues. Promoter cis-acting element analysis revealed that these 44K+-transport-related genes may be associated with responses to 10 abiotic stresses, primarily light, methyl jasmonate (MeJA) and abscisic acid (ABA). Our results provide a systematic and comprehensive overview of K+-transport-related gene families in Chinese cabbage and establish a foundation for further research on K+ absorption and transport functions.

Genome-Wide Identification and Characterization of Calcium-Dependent Protein Kinase (CDPK) and CDPK-Related Kinase (CRK) Gene Families in Medicago truncatula.

Calcium-dependent protein kinase (CDPK or CPK) and CDPK-related kinase (CRK) play an important role in plant growth, development, and adaptation to environmental stresses. However, their gene families had been yet inadequately investigated in Medicago truncatula. In this study, six MtCRK genes were computationally identified, they were classified into five groups with MtCDPKs based on phylogenetic relationships. Six pairs of segmental duplications were observed in MtCDPK and MtCRK genes and the Ka/Ks ratio, an indicator of selection pressure, was below 0.310, indicating that these gene pairs underwent strong purifying selection. Cis-acting elements of morphogenesis, multiple hormone responses, and abiotic stresses were predicted in the promoter region. The spatial expression of MtCDPKs and MtCRKs displays diversity. The expression of MtCDPKs and MtCRKs could be regulated by various stresses. MtCDPK4, 14, 16, 22, and MtCRK6 harbor both N-myristoylation site and palmitoylation site and were anchored on plasma membrane, while MtCDPK7, 9, and 15 contain no or only one N-acylation site and were distributed in cytosol and nucleus, suggesting that the N-terminal acylation sites play a key role in subcellular localization of MtCDPKs and MtCRKs. In summary, comprehensive characterization of MtCDPKs and MtCRKs provide a subset of candidate genes for further functional analysis and genetic improvement against drought, cold, salt and biotic stress.

Genome-Wide Identification and Expression Analysis of the Aux/IAA and Auxin Response Factor Gene Family in Medicago truncatula.

Aux/IAA and auxin response transcription factor (ARF) genes are key regulators of auxin responses in plants. A total of 25 MtIAA and 40 MtARF genes were identified based on the latest updated Medicago truncatula reference genome sequence. They were clustered into 10 and 8 major groups, respectively. The homologs among M. truncatula, soybean, and Arabidopsis thaliana shared close relationships based on phylogenetic analysis. Gene structure analysis revealed that MtIAA and MtARF genes contained one to four concern motifs and they are localized to eight chromosomes, except chromosome 6 without MtARFs. In addition, some MtIAA and MtARF genes were expressed in all tissues, while others were specifically expressed in specific tissues. Analysis of cis-acting elements in promoter region and expression profiles revealed the potential response of MtIAA and MtARF genes to hormones and abiotic stresses. The prediction protein-protein interaction network showed that some ARF proteins could interact with multiple Aux/IAA proteins, and the reverse is also true. The investigation provides valuable, basic information for further studies on the biological functions of MtIAA and MtARF genes in the regulation of auxin-related pathways in M. truncatula.

Genome-Wide Identification and Characterization of Short-Chain Dehydrogenase/Reductase (SDR) Gene Family in Medicago truncatula.

Short-chain dehydrogenase/reductase (SDR) belongs to the NAD(P)(H)-dependent oxidoreductase superfamily. Limited investigations reveal that SDRs participate in diverse metabolisms. A genome-wide identification of the SDR gene family in M. truncatula was conducted. A total of 213 MtSDR genes were identified, and they were distributed on all chromosomes unevenly. MtSDR proteins were categorized into seven subgroups based on phylogenetic analysis and three types including 'classic', 'extended', and 'atypical', depending on the cofactor-binding site and active site. Analysis of the data from M. truncatula Gene Expression Atlas (MtGEA) showed that above half of MtSDRs were expressed in at least one organ, and lots of MtSDRs had a preference in a tissue-specific expression. The cis-acting element responsive to plant hormones (salicylic acid, ABA, auxin, MeJA, and gibberellin) and stresses were found in the promoter of some MtSDRs. Many genes of MtSDR7C,MtSDR65C, MtSDR110C, MtSDR114C, and MtSDR108E families were responsive to drought, salt, and cold. The study provides useful information for further investigation on biological functions of MtSDRs, especially in abiotic stress adaptation, in the future.