Processed Pseudogenes, Processed Genes, and Spontaneous Mutations in the Arabidopsis Genome

We identified 411 processed sequences in the Arabidopsis thaliana genome based on the fact that they have lost their intron(s) and have a length that is at least 95% of the length of the gene that gave rise to them. These sequences were generated by 230 different genes and clearly originated from retrotranspositons events because most of them (91%) have a poly(A)-tail. They are composed of 376 sequences with frame shifts and/or premature stop codons (processed pseudogenes) and 35 sequences without disablements (processed genes). Eleven of these processed genes are likely functional retrotransposed genes because they have low Ka/Ks ratios and high Ks values, and their sequences match numerous Arabidopsis ESTs. Processed sequences are mostly randomly distributed in the Arabidopsis genome and their rate of accumulation has steadily been decreasing since it peaked some 50 MYA. In contrast with the situation observed in mammals, the processed sequences found in the Arabidopsis genome originate from genes with high copy numbers and not from highly expressed genes. The patterns of spontaneous mutations in Arabidopsis are slightly different than those of mammals but are similar to those observed in Drosophila. This suggests that methylated cytosine deamination is less frequent in Arabidopsis than in mammals. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editor: Dr. Juergen Brosius]     

Identification of ERF genes in peanuts and functional analysis of AhERF008 and AhERF019 in abiotic stress response

Ethylene-responsive factor (ERF) play an important role in regulating gene expression in plant development and response to stresses. In peanuts (Arachis hypogaea L.), which produce flowers aerially and pods underground, only a few ERF genes have been identified so far. This study identifies 63 ERF unigenes from 247,313 peanut EST sequences available in the NCBI database. The phylogeny, gene structures, and putative conserved motifs in the peanut ERF proteins were analysed. Comparative analysis revealed the absence of two subgroups (A1 and A3) of the ERF family in peanuts; only 10 subgroups were identified in peanuts compared to 12 subgroups in Arabidopsis and soybeans. AP2/ERF domains were found to be conserved among peanuts, Arabidopsis, and soybeans. Outside the AP2/ERF domain, many soybean-specific conserved motifs were also detected in peanuts. The expression analysis of ERF family genes representing each clade revealed differential expression patterns in response to biotic and abiotic stresses. Overexpression of AhERF008 influenced the root gravity of Arabidopsis, whereas overexpression of AhERF019 enhanced tolerance to drought, heat, and salt stresses in Arabidopsis. The information generated in this study will be helpful to further investigate the function of ERFs in plant development and stress response.     

Expression and DNA methylation pattern of reproduction-related genes in partially fertile triploid Pacific oysters <Emphasis Type="Italic">Crassostrea gigas</Emphasis>

Partial or complete sterility is an obvious feature in triploid Pacific oyster (Crassostrea gigas) which contributes to improving rearing performances. Despite the significance of sterility, the molecular mechanism behind it remains elusive and related research was limited. This study focused on six reproduction-related genes and compared their different behavior in gene expression and DNA methylation pattern between triploid and diploid oysters in order to provide more molecular information. The gonadal development of triploid oyster was examined by histology before molecular analysis. Gametogenesis disturbance was observed in triploid oysters at different development stages (stage II and III) with more serious impairment in females. QPCR showed significant gene expression difference between diploid and triploid in two genes: putative Vg and cgER. Gene expression of putative Vg was delayed in triploids while for cgER triploid oyster showed higher expression and the difference was significant at stage III. DNA methylation pattern of these two genes were further investigated by bisulfite sequencing. Between diploid and triploid oysters, no difference was observed in total methylation level but some individual loci showed different patterns: significantly high methylation rate of loci 2284 in cgER was observed in triploid oyster which has a higher expression of this gene. This study indicated that putative Vg and cgER might play a role in partial sterile in triploid C. gigas. Gene expression could be regulated by the methylation pattern at specific individual locus, which deserves equivalent attention as well as total DNA methylation level.     

Genome-wide analysis of GDSL-type esterases/lipases in <Emphasis Type="Italic">Arabidopsis</Emphasis>

Key message

In this present study, we introduce a fundamental framework and provide information regarding the possible roles of GDSL-type esterase/lipase gene family in Arabidopsis.

Abstract

GDSL-type esterases/lipases are hydrolytic enzymes with multifunctional properties such as broad substrate specificity, regiospecificity, and stereoselectivity. In this study, we identified 105 GDSL-type esterase/lipase genes in Arabidopsis thaliana by conducting a comprehensive computational analysis. Expression studies indicated that GDSL-type lipase proteins showed varied expression patterns. Phylogenetic tree analysis indicated that AtGELP (Arabidopsis thaliana GDSL-type esterase/lipase protein) gene family was divided into four clades. The phylogenetic analysis, combined with protein motif architectures, and expression profiling were used to predict the roles AtGELP genes. To investigate the physical roles of the AtGELP gene family, we successfully screened 88 AtGELP T-DNA knockout lines for 54 AtGELP genes from 199 putative SALK T-DNA mutants. Transgenic plants of AtGELP genes were used to elucidate the phenotypic characteristics in various developmental stages or stress conditions. Our results suggest that the AtGELP genes have diverse physical functions such as affecting the germination rate and early growth of seedlings subjected to high concentrations of glucose, or being involved in biotic stress responses.

     

Genomics and expression analysis of DHHC-cysteine-rich domain S-acyl transferase protein family in apple

S-acylation is one of a group of lipid modifications that occurs on eukaryotic proteins, mediated by DHHC-CRD-containing proteins, which plays an important role in regulating the membrane association, trafficking and function of target proteins. Although genome-wide identification of PAT family has been carried out in yeast, mice, humans and Arabidopsis, little is known about apple PAT genes. In this study, a total of 33 putative apple PAT proteins, containing DHHC-CRD by domain analysis, have been identified, and were classified into three groups according to the phylogenetic analysis of PAT proteins in apple and Arabidopsis. More complex TMDs in the most MdPATs revealed the PM location of the gene family. Gene structure, gene chromosomal location and paralogs analysis of MdPAT genes within the apple genome demonstrated that tandem and segmental duplications, as well as whole genome duplications, have likely contributed to the expansion and evolution of the PAT gene family in apple. According to the microarray and expressed sequence tag (ESTs) analysis, the different expression patterns indicate that they may play different roles during fruit development and rootstock-scion interactions process. Moreover, PATs were performed expression profile analyses in different tissues, indicating that the PATs are involved in various aspects of physiological and developmental processes of apple. To our knowledge, this is the first report of a genome-wide analysis of the apple PAT gene family, and this genomic analysis of apple DHHC-CRD PAT genes provides the first step towards a functional study of this gene family in apple.     

Characterization of an HSP70 Cognate Gene Family in Arabidopsis

Analysis of the polypeptide composition of extracts from heat-shocked leaves of Arabidopsis indicated the presence of at least 12 HSP70-related polypeptides, most of which were constitutively expressed. In vitro translation of mRNA from heat-shocked and control leaves indicated that the amount of mRNA encoding four HSP70 polypeptides was increased strongly by heat-shock. Three Arabidopsis genes which exhibit homology to a Drosophila HSP70 gene were cloned. Two of the three genes are arranged in direct orientation approximately 1.5 kilobases apart. The third gene is not closely linked to the other two. Nucleotide sequence analysis of the 5′ regions of the two linked genes revealed that both contain a TATA box, the CAAT motif, and several short sequences which are homologous to the Drosophila heat-shock consensus sequence. The deduced partial amino acid sequence of the open reading frames were 79 and 72% homologous to the corresponding regions of the Drosophila HSP70-cognate and HSP70 sequences, respectively. As with the two maize HSP70 genes which have been characterized, and the Drosophila HSP70-cognate genes, the Arabidopsis genes contained a putative intron in the codon specifying amino acid 72. Analysis of mRNA levels with gene-specific oligonucleotide probes indicated that two of the genes were not expressed or were expressed at very low levels in leaves during normal growth or after heat-shock, whereas the other gene was constitutively expressed. By analogy with the results of similar studies of other organisms, it appears that the three cloned genes are members of a small family which are most closely related to the HSP70-cognate genes found in other species.     

Molecular characterization of 5-chlorophyll a/b-binding protein genes from Panax ginseng Meyer and their expression analysis during abiotic stresses

The chlorophyll _a/b-binding protein (CAB) serves in both photosystems (PS), I and II, as a coordinator of antenna pigments in the light-harvesting complex (LHC). The CABs constitute abundant and important proteins in the thylakoid membrane of higher plants. In our study, five CAB genes, which contained full-length cDNA sequences from the 4-year-old ginseng leaves (Panax ginseng Meyer), were isolated and named PgCAB. Phylogenetic comparison of the members of the subfamily between ginseng and higher plants, including Arabidopsis, revealed that the putative functions of these ginseng CAB proteins were clustered into the different family of Arabidopsis CABs; two PgCABs in LHCII family and three PgCABs in LHCI family. The expression analysis of PgCABs consistently showed dark-dependent inhibition in leaves. Expression analysis during abiotic stress identified that PgCAB genes responded to heavy metal, salinity, chilling, and UV stresses differently, suggesting their specific function during photosynthesis. This is the first comprehensive study of the CAB gene family in P. ginseng.