Fish growth hormone genes: Divergence of coding sequences in salmonid fishes

Comparison of coding nucleotide sequences of the paralogous GH1 and GH2 genes, as well as of the growth hormone amino acid sequences, in the species of closely related salmonid genera Salvelinus, Oncorhynchus, and Salmo was performed. It was demonstrated that, in different groups of salmonids, the amino acid substitution rates were considerably different. In some cases, an obvious discrepancy between the divergence of growth hormone genes and phylogenetic schemes based on other methods and approaches was revealed. These findings suggest that the reason may be multidirectional selection at duplicated genes at different stages of evolution.     

Unique configuration of genes of silicon transporter in the freshwater pennate diatom <Emphasis Type="Italic">Synedra acus</Emphasis> subsp. <Emphasis Type="Italic">radians</Emphasis>

The existence of the cluster of duplicated sit silicon transporter genes in the chromosome of the diatom Synedra acus subsp. radians was shown for the first time. Earlier, the localization of sit genes in the same chromosome and cluster formation caused by gene duplication was shown only for the marine raphid pennate diatom P. tricornutum. Only non-clustered sit genes were found in the genomes of other diatoms. It is reasonable to assume that sit tandem (sit-td) and sit triplet (sit-tri) genes of S. acus subsp. radians occurred as a result of gene duplication followed by divergence of gene copies.     

Molecular evolution of glutathione peroxidase (<Emphasis Type="Italic">Gpx</Emphasis>) family genes in desert poplar (<Emphasis Type="Italic">Populus euphratica</Emphasis> Oliv.)

Populus euphratica Oliv. is a poplar species that is distributed mainly in deserts, making it an interesting model in which to investigate molecular mechanisms underlying different stress responses. Here, we used molecular population genetic methods to detect potential selection in candidate genes belonging to the P. euphratica glutathione (GSH) peroxidase (Gpx) family, which are associated with an enzymatic mechanism that combats oxidative damage caused by reactive oxygen species (ROS) in plant cells; earlier studies have shown that Gpx proteins play important roles in coping with increased ROS levels during biotic and abiotic stresses in plants. We analyzed the nucleotide diversity and divergence patterns of five loci encoding Gpx genes, and 16 reference loci used as controls, in order to detect departures from the neutral expectation. Gpx1 has an excess of mid-frequency alleles; high intraspecific nucleotide diversity, distributed in the upper tail of the simulated neutral model; and extensive LD, showing strong evidence of balancing selection/local adaptation. The Gpx3.2 gene exhibits very low nucleotide diversity and divergence, suggesting that it has evolved under strong purifying selection. We failed to detect any evidence for natural selection at the other loci (Gpx2, Gpx4, and Gpx5) compared with the reference loci. The results show that nucleotide diversity and/or divergence differ greatly between members of the Gpx gene family, resulting from differential selective pressure acting on genes, and that adaptive evolution influenced the distribution of P. euphratica in desert regions.     

Tandem duplications of receptor-like kinase genes reshaped the homologous chromosome 1 regions in <Emphasis Type="Italic">Oryza sativa</Emphasis> ssp. <Emphasis Type="Italic">indica</Emphasis> and <Emphasis Type="Italic">japonica</Emphasis> and their possible ancestral genomes

Plant receptor-like kinase (Rlk) genes form a large family, each encoding a protein with a signal motif, a single transmembrane region, and a cytoplasmic kinase domain. Various gene duplications have contributed to the establishment and expansion of the family. Here, we characterized the formation and evolution of the Rlk gene family in cultivated rice and their possible progenitors. Using wheat Rlk gene sequences, we identified orthologs from the genomes of domesticated rice subspecies Oryza sativa ssp. japonica and ssp. indica and their putative progenitors O. glaberrima and O. rufipogon. The four chromosome 1 orthologous regions ranged from 103 to 281 kb comprising 181 syntenic blocks with 75 to 100% sequence identity. These regions contained 11–19 Triticum aestivum kinases (Taks) and 10–15 Lr10 receptor-like kinases (Lrks) organized in clusters and 3–12 transposable elements (TEs). Dot plot analyses showed that the 4 regions had 21–37 conserved catalytic domains, mainly in protein kinases (PKs) and tyrosine kinases (TyrKs) in coupling state. Over 50% of the sequences of glaberrima/rufipogon and japonica/indica pairs were colinear, while japonica/indica displayed a marked sequence expansion with duplicated genes and TEs. A total of 2312 single nucleotide polymorphisms (SNPs) and insertion-deletions (INDELs) were identified between japonica and indica. Duplication of the Rlk genes in O. glaberrima and O. rufipogon occurred after the grass species radiation and before the divergence of O. rufipogon from O. glaberrima; the orthologous Rlk genes from O. japonica and O. indica duplicated after O. sativa separated from O. rufipogon; paralogs, obtained through extensive duplication, happened after the separation of rice from maize. Tandem duplication was the major factor contributing to the gene copy number variation and genome size expansion.     

Gabaculine selection using bacterial and plant marker genes (GSA-AT) in durum wheat transformation

Selectable marker genes are widely used for the efficient transformation of crop plants. In most cases, selection is based on antibiotic or herbicide resistance genes because they tend to be most efficient. The Synechococcus hemL gene has been successfully employed as a selectable marker for tobacco and alfalfa genetic transformation, by using gabaculine as the selective agent. The gene conferring gabaculine resistance is a mutant form of the hemL gene from Synechococcus PCC6301, strain GR6, encoding a gabaculine insensitive form of the glutamate1-semialdehyde aminotransferase (GSA) enzyme. In the present study we compared the transformation and selection efficiency of the common selection method based on the Streptomyces hygroscopicus bar gene conferring resistance to Bialaphos^®, with both the Synechococcus hemL gene and a Medicago sativa mutated GSA gene (MsGSAgr) conferring resistance to phytotoxin gabaculine. Callus derived from immature embryos of the durum wheat cultivar Varano were simultaneously co-bombarded with bar/hemL and bar/MsGSAgr genes. After gene delivery, the marker genes were individually evaluated through all the selection phases from callus regeneration to adult plant formation, and compared for their transformation and selection efficiency. The integration of the three genes in the T₀ generation was confirmed by PCR analysis with specific primers for each gene and southern blot analysis. Both Synechococcus hemL and MsGSA were more efficient than bar for biolistic transformation (2.8% vs. 1.8% and 1.1% vs. 0.5%) and selection (79% vs. 43% and 87% vs. 50%). Thus, an efficient selection method for durum wheat transformation was established that obviates the use of herbicide resistance genes.     

The complete chloroplast genome sequence of <Emphasis Type="Italic">Pseudoroegneria libanotica</Emphasis>, genomic features,and phylogenetic relationship with <Emphasis Type="Italic">Triticeae</Emphasis> species

Pseudoroegneria libanotica is an important herbage diploid species possessing the St genome. The St genome participates in the formation of nine perennial genera in Triticeae (Poaceae). The whole chloroplast (cp) genome of P. libanotica is 135 026 bp in length. The typical quadripartite structure consists of one large single copy of 80 634 bp, one small single copy of 12 766 bp and a pair of inverted regions (20 813 bp each). The cp genome contains 76 coding genes, four ribosomal RNA and 30 transfer RNA genes. Comparative sequence analysis suggested that: 1) the 737 bp deletion in the cp of P. libanotica was specific in Triticeae species and might transfer into its nuclear genome; 2) hot-spot regions, indels in intergenic regions and protein coding sequences mainly led to the length variation in Triticeae; 3) highly divergence regions combined with negative selection in rpl2, rps12, ccsA, rps8, ndhH, petD, ndhK, psbM, rps3, rps18, and ndhA were identified as effective molecular markers and could be considered in future phylogenetic studies of Triticeae species; and 4) ycf3 gene with rich cpSSRs was suitable for phylogeny analysis or could be used for DNA barcoding at low taxonomic levels. The cpSSRs distribution in the coding regions of diploid Triticeae species was shown for the first time and provided a valuable source for developing primers to study specific simple sequence repeat loci.     

Genome-wide identification and resistance expression analysis of the <Emphasis Type="Italic">NBS</Emphasis> gene family in <Emphasis Type="Italic">Triticum urartu</Emphasis>

As the largest class of resistant genes, the nucleotide binding site (NBS) has been studied extensively at a genome-wide level in rice, sorghum, maize, barley and hexaploid wheat. However, no such comprehensive analysis has been conducted of the NBS gene family in Triticum urartu, the donor of the A genome to the common wheat. Using a bioinformatics method, 463 NBS genes were isolated from the whole genome of T. urartu, of which 461 had location information. The expansion pattern and evolution of the 461 NBS candidate proteins were analyzed, and 118 of them were duplicated. By calculating the lengths of the copies, it was inferred that the NBS resistance gene family of T. urartu has experienced at least two duplication events. Expression analysis based on RNA-seq data found that 6 genes were differentially expressed among Tu38, Tu138 and Tu158 in response to Blumeria graminis f.sp.tritici (Bgt). Following Bgt infection, the expression levels of these genes were up-regulated. These results provide critical references for further identification and analysis of NBS family genes with important functions.     

Genome-wide identification and expression analysis of the GRAS family proteins in <Emphasis Type="Italic">Medicago truncatula</Emphasis>

The GRAS gene family performs a variety of functions in plant growth and development processes, and they also play essential roles in plant response to environmental stresses. Medicago truncatula is a diploid plant with a small genome used as a model organism. Despite the vital role of GRAS genes in plant growth regulation, few studies on these genes in M. truncatula have been conducted to date. Using the M. truncatula reference genome data, we identified 68 MtGRAS genes, which were classified into 16 groups by phylogenetic analysis, located on eight chromosomes. The structure analysis indicated that MtGRAS genes retained a relatively constant exon–intron composition during the evolution of the M. truncatula genome. Most of the closely related members in the phylogenetic tree had similar motif compositions. Different motifs distributed in different groups of the MtGRAS genes were the sources of their functional divergence. Twenty-eight MtGRAS genes were expressed in six tissues, namely root, bud, blade, seedpod, nodule, and flower tissues, suggesting their putative function in many aspects of plant growth and development. Nine MtGRAS genes were upregulated under cold, freezing, drought, ABA, and salt stress treatments, indicating that they play vital roles in the response to abiotic stress in M. truncatula. Our study provides valuable information that can be utilized to improve the quality and agronomic benefits of M. truncatula and other plants.     

Genome-wide identification,classification, and expression analysis of the phytocyanin gene family in <Emphasis Type="Italic">Phalaenopsis equestris</Emphasis>

Phytocyanins (PCs) are ancient blue copper-binding proteins in plants that bind to single type I copper atoms and function as electron transporters. PCs play an important role in plant development and stress resistance. Many PCs are considered to be chimeric arabinogalactan proteins (AGPs). Previously, 38, 62, and 84 PC genes were identified in Arabidopsis thaliana, Oryza sativa, and Brassica rapa, respectively. In this study, we identified 30 putative PC genes in the orchid Phalaenopsis equestris through comprehensive bioinformatics analysis. Based on phylogeny and motif constitution, the P. equestris phytocyanins (PePCs) were divided into five subclasses: 10 early nodulin-like proteins, 10 uclacyanin-like proteins, five stellacyanin-like proteins, four plantacyanin-like proteins, and one unknown protein. Structural and glycosylation predictions suggested that 16 PePCs were glycosylphosphatidylinositol-anchored proteins localized to the plasma membrane, 22 PePCs contain N-glycosylation sites, and 14 are chimeric AGPs. Phylogenetic analysis indicated that each subfamily was derived from a common ancestor before the divergence of monocot and dicot lineages and that the expansion of the PC subfamilies occurred after the divergence of orchids and Arabidopsis. The number of exons in PC genes was conserved. Expression analysis in four tissues revealed that nine PC genes were highly expressed in flowers, stems, and roots, suggesting that these genes play important roles in growth and development in P. equestris. The results of this study lay the foundation for further analysis of the functions of this gene family in plants.     

Genome-wide analysis of the PHB gene family in <Emphasis Type="Italic">Glycine max</Emphasis> (L.) Merr.

Prohibitins (PHBs) have one SPFH domain in common and present in species ranging from prokaryotes to eukaryotes. Although a number of researches on PHBs were performed in different plant species, a systematic analysis of the PHB family in soybean is still remains uncharacterized. In the present study, 24 putative PHB genes have been first systemically identified in soybean. According to phylogenetic analysis, these GmPHBs could be classified into four groups. Gene structures and motif patterns showed high levels of conservation within the phylogenetic subgroups. Several members of this family have undergone purifying selection based on Ka/Ks analysis on duplicated PHB genes in soybean. We performed microsynteny analysis across four legume species based on the comparisons among the specific regions contained in PHB genes. As a result, numerous microsyntenic gene pairs among soybean, Medicago, Lotus and Phaseolus were identified. Most soybean PHB genes exhibited different expression levels in various tissues and developmental stages through expression analysis using publicly available RNA-seq datasets. The 11 GmPHB genes from III_B subgroup were examined by qPCR for their expression in two soybean cultivar after infection by Phytophthora sojae. Besides three GmPHB genes previous reported by us, here other four genes also were rapidly induced by P. sojae infection in the resistant genotype, while induction was very weak in the susceptible genotype. The comprehensive overview of the PHB gene family in soybean genome will provide useful information for further functional analysis of the PHB gene family in soybean.     

Three <Emphasis Type="Italic">P5CS</Emphasis> genes including a novel one from <Emphasis Type="Italic">Lilium regale</Emphasis> play distinct roles in osmotic,drought and salt stress tolerance

Proline accumulations in abiotically stressed plants is generally considered to benefit their stress tolerance. The Δ¹-Pyrroline-5-carboxylate synthetase (P5CS) gene family, which encodes the rate-limiting enzyme in proline biosynthesis pathway, usually contains two duplicated genes in most plants. However, three P5CS genes including LrP5CS1, LrP5CS2 as well as a third one, LrP5CS3, were isolated from Lilium regale. LrP5CS3 is highly identical to LrP5CS1 in amino acid sequences, indicating they could come from a paralogous duplication. The phylogenetic tree suggested that the duplication of LrP5CS occurred independently after the divergence of Liliales and commelinoids. The expression of LrP5CS1 was strongly induced in leaves and roots both under drought and salinity, while that of LrP5CS3 was upregulated more moderately. LrP5CS2 stayed almost constitutive under stress. LrP5CS1 exhibited the highest activity after expressed in E. coli. Overexpression of LrP5CS genes conferred enhanced osmotic, drought and salt tolerance on transgenic Arabidopsis without negative effects in unstressed condition. Under salt stress, lines LrP5CS2 accumulated fewer proline than others, and lines LrP5CS1 grew better in root elongation. The roots of lines LrP5CS3 grew better than all others under unstressed condition and osmotic stress. Our study suggests that the three LrP5CS genes play distinct roles respectively in proline accumulation and abiotic stress tolerance.     

Plastomes of Mimosoideae: structural and size variation,sequence divergence,and phylogenetic implication

Plastomes of Fabaceae show both significant structural and size variation; however, most published plastomes are from subfamily Papilionoideae and only a few are from the other two subfamilies. In order to address the plastome structural and size variation of subfamily Mimosoideae, we integrated 11 newly sequenced plastomes from representing genera with three previously published ones. Each mimosoid plastome presented a typical quadripartite structure and contained 111 unique genes. Their inverted repeats (IRs) experienced multiple expansion/contraction; a ca. 13-kb IR expansion into small single copy (SSC) was detected in plastomes of a clade formed by tribe Ingeae and Acacia sensu stricto (s.s.), and a ca. 1.7-kb IR expansion into and a ca. 1.9-kb contraction out of large single copy (LSC) were found in Pithecellobium flexicaule and Acacia dealbata, respectively. Linear regression analysis showed decreased synonymous substitution rates of genes relocating from SSC into IR. A loss of both introns of clpP occurred in A. dealbata and Faidherbia albida, and a duplicated clpP copy was detected in A. dealbata. Furthermore, a 421-bp inversion that containing rps18 was found in A. dealbata. The size of mimosoid plastomes was found significantly affected by a IR-SC boundary shift, and also associated with repeat content. Plastome coding and noncoding regions with variable sequence divergence may supply valuable markers for molecular evolutionary and phylogenetic studies at different taxonomic levels. Plastid phylogenomics well resolved relationships among sampled mimosoid species.     

Molecular characterization of <Emphasis Type="Italic">NBS</Emphasis>–<Emphasis Type="Italic">LRR</Emphasis> genes in the soybean <Emphasis Type="Italic">Rsv3</Emphasis> locus reveals several divergent alleles that likely confer resistance to the soybean mosaic virus

Key message

The divergence patterns of NBS – LRR genes in soybean Rsv3 locus were deciphered and several divergent alleles ( NBS_C, NBS_D and Columbia NBS_E ) were identified as the likely functional candidates of Rsv3.

Abstract

The soybean Rsv3 locus, which confers resistance to the soybean mosaic virus (SMV), has been previously mapped to a region containing five nucleotide binding site–leucine-rich repeats (NBS–LRR) genes (referred to as nbs_A–E) in Williams 82. In resistant cultivars, however, the number of NBS–LRR genes in this region and their divergence from susceptible alleles remain unclear. In the present study, we constructed and screened a bacterial artificial chromosome (BAC) library for an Rsv3-possessing cultivar, Zaoshu 18. Sequencing two positive BAC inserts on the Rsv3 locus revealed that Zaoshu 18 possesses the same gene content and order as Williams 82, but two of the NBS–LRR genes, NBS_C and NBS_D, exhibit distinct features that were not observed in the Williams 82 alleles. Obtaining these NBS-LRR genes from eight additional cultivars demonstrated that the NBS_A–D genes diverged into two different alleles: the nbs_A–D alleles were associated with the rsv3-type cultivars, whereas the NBS_A–D alleles were associated with the Rsv3-possessing cultivars. For the NBS_E gene, the cultivar Columbia possesses an allele (NBS_E) that differed from that in Zaoshu 18 and rsv3-type cultivars (nbs_E). Exchanged fragments were further detected on alleles of the NBS_C–E genes, suggesting that recombination is a major force responsible for allele divergence. Also, the LRR domains of the NBS_C–E genes exhibited extremely strong signals of positive selection. Overall, the divergence patterns of the NBS–LRR genes in Rsv3 locus elucidated by this study indicate that not only NBS_C but also NBS_D and Columbia NBS_E are likely functional alleles that confer resistance to SMV.