Overexpression of miR5505 enhanced drought and salt resistance in rice (Orayza sativa)

Rice is one of the most important crops in the world and half of the world population consumes it as their staple food. The abiotic stresses caused by drought, salt and other stresses have severely impacted rice production. MicroRNAs (miRNAs) are a type of small non-coding RNAs which widely reported as gene regulators, suppressing genes expression by degradation mRNA or translation inhibition. Previously, high-throughput sequencing has found a conserved miRNA miR5505 responding to drought stress in Dongxiang wild rice (DXWR). Several other studies also revealed that miR5505 was involved in rice stress responses. We further studied the effect of miRNA in drought and salt tolerance by overexpression it in rice. 2 in 18 successfully transformed transgenic lines with higher miR5505 expression were selected and then drought and salt resistance ability were evaluated. Both transgenic lines showed stronger drought and salt tolerance than wild-type (WT). Putative targets of miR5505 were identified by psRNATarget and several of them were found stress-related. RNA-seq found 1,980 differentially expressed genes (DEGs) in transgenic lines. Among them, 978 genes were down-regulated. Three genes were predicted by psRNATarget and two of them might be stress-related. We also found various environmental stress cis-acting elements in upstream of miR5505 promoter through Software PlantCARE. In all, we improved rice drought and salt tolerance by overexpressing miR5505, and the generated putative targets and cis-acting elements also suggested miR5505 might play important roles in the regulation of drought and salt responses. Keywords: rice, overexpression line , drought and salt stress, miR5505

Genome Wide Identification of Respiratory Burst Oxidase Homolog (Rboh) Genes in Citrus sinensis and Functional Analysis of CsRbohD in Cold Tolerance

Respiratory burst oxidase homologs (Rbohs) are critical enzymes involved in the generation of reactive oxygen species (ROS) that play an important role in plant growth and development as well as various biotic and abiotic stresses in plants. Thus far, there have been few reports on the characterization of the Rboh gene family in Citrus. In this study, seven Rboh genes (CsRbohA~CsRbohG) were identified in the Citrus sinensis genome. The CsRboh proteins were predicted to localize to the cell membrane. Most CsRbohs contained four conserved domains, an EF-hand domain, and a transmembrane region. Phylogenetic analysis demonstrated that the CsRbohs were divided into five groups, suggesting potential distinct functions and evolution. The expression profiles revealed that these seven CsRboh genes displayed tissue-specific expression patterns, and five CsRboh genes were responsive to cold stress. Fourteen putative cis-acting elements related to stress response, hormone response, and development regulation were present within the promoters of CsRboh genes. The in-silico microRNA target transcript analyses indicated that CsRbohE might be targeted by csi-miR164. Further functional and physiological analyses showed that the knockdown of CsRbohD in trifoliate orange impaired resistance to cold stress. As a whole, our results provide valuable information for further functional studies of the CsRboh genes in response to cold stress.

Genome-Wide Identification and Characterization of the RCI2 Gene Family in Allotetraploid Brassica napus Compared with Its Diploid Progenitors

Brassica napus and its diploid progenitors (B. rapa and B. oleracea) are suitable for studying the problems associated with polyploidization. As an important anti-stress protein, RCI2 proteins widely exist in various tissues of plants, and are crucial to plant growth, development, and stress response. In this study, the RCI2 gene family was comprehensively identified and analyzed, and 9, 9, and 24 RCI2 genes were identified in B. rapa, B. oleracea, and B. napus, respectively. Phylogenetic analysis showed that all of the identified RCI2 genes were divided into two groups, and further divided into three subgroups. Ka/Ks analysis showed that most of the identified RCI2 genes underwent a purifying selection after the duplication events. Moreover, gene structure analysis showed that the structure of RCI2 genes is largely conserved during polyploidization. The promoters of the RCI2 genes in B. napus contained more cis-acting elements, which were mainly involved in plant development and growth, plant hormone response, and stress responses. Thus, B. napus might have potential advantages in some biological aspects. In addition, the changes of RCI2 genes during polyploidization were also discussed from the aspects of gene number, gene structure, gene relative location, and gene expression, which can provide reference for future polyploidization analysis.

Differential Expression of lncRNAs and miRNAs Between Self-Rooting Juvenile and Donor Clones Unveils Novel Insight Into the Molecular Regulation of Rubber Biosynthesis in Hevea brasiliensis

The rubber tree (Hevea brasiliensis Muell. Arg.) is a tropical tree species that produce natural rubber. Self-rooted juvenile clones (SRJCs) are novel rubber tree planting materials developed through primary somatic embryogenesis. SRJCs have a higher rubber yield compared with donor clones (DCs). The molecular basis underlying increased rubber yield in SRJCs remains largely unknown. Here, the latex from SRJCs and DCs were collected for strand-specific and small RNA-seq methods. A total of 196 differentially expressed long noncoding RNAs (DELs), and 11 differentially expressed microRNAs were identified in latex between SRJCs and DCs. Targeted genes of DELs were markedly enriched for various biological pathways related to plant hormone signal transduction, photosynthesis, glutathione metabolism, and amino acids biosynthesis. DELs probably acted as cis-acting regulation was calculated, and these DELs relevant to potentially regulate rubber biosynthesis, reactive oxygen species metabolism, and epigenetic modification. Furthermore, the DELs acting as microRNA targets were studied. The interaction of microRNA and DELs might involve in the regulation of natural rubber biosynthesis.

In silico characterization and comparison of the fruit ripening related beta‐ amylase (BAM) gene family in banana genome A and B

Banana is one of the most important commodities for maintaining global food security. Primary metabolic processes during the ripening of banana greatly affect post‐harvest quality, particularly in starch metabolism. The beta‐ amylase (BAM) gene family is known as a group of genes that plays an important role in starch metabolism regulation. In this study, we focused on the characterization and comparative analysis of the BAM gene family in DH Pahang and Pisang Klutuk Wulung (PKW) varieties, these being the AA and BB genomes, respectively. The sequences of BAM gene family were retrieved from the database of Musa acuminata ’DH Pahang’ and Musa balbisiana ’PKW’ genome, then structural and functional characterization was performed, followed by identification of cis‐acting elements in the BAM promoter regions. The results showed that the BAM gene family structure was relatively conserved in both genomes, and a putative BAM11 gene was found, the function of which has not been studied in other plants. Cis‐acting element analysis showed that they were distinct in the copy number and types of elements that were responsive to various phytohormones. This study suggested that the BAM genes involved in ripening are spatiotemporally regulated. However, further functional genomic analysis is required to describe the specific role and regulation of BAM genes during ripening in banana.

Genome-Wide Identification and Expression Profile of OSCA Gene Family Members in Triticum aestivum L.

Hyperosmolality and various other stimuli can trigger an increase in cytoplasmic-free calcium concentration ([Ca2+]cyt). Members of the Arabidopsis thaliana (L.) reduced hyperosmolality-gated calcium-permeable channels (OSCA) gene family are reported to be involved in sensing extracellular changes to trigger hyperosmolality-induced [Ca2+]cyt increases and controlling stomatal closure during immune signaling. Wheat (Triticum aestivum L.) is a very important food crop, but there are few studies of its OSCA gene family members. In this study, 42 OSCA members were identified in the wheat genome, and phylogenetic analysis can divide them into four clades. The members of each clade have similar gene structures, conserved motifs, and domains. TaOSCA genes were predicted to be regulated by cis-acting elements such as STRE, MBS, DRE1, ABRE, etc. Quantitative PCR results showed that they have different expression patterns in different tissues. The expression profiles of 15 selected TaOSCAs were examined after PEG (polyethylene glycol), NaCl, and ABA (abscisic acid) treatment. All 15 TaOSCA members responded to PEG treatment, while TaOSCA12/-39 responded simultaneously to PEG and ABA. This study informs research into the biological function and evolution of TaOSCA and lays the foundation for the breeding and genetic improvement of wheat.

Effects of homoeologous exchange on gene expression and alternative splicing in a newly formed allotetraploid wheat

Homoeologous exchange (HE) is a major mechanism generating post-polyploidization genetic variation with important evolutionary consequences. However, the direct impacts of HE without entangling with additional evolutionary forces on gene expression remains to be fully understood. Here, we analyzed high-throughput RNA-seq data of young leaves from individuals of a synthetic allotetraploid wheat (AADD), which contain variable numbers of HEs. We aimed to investigate if and to which extent HE directly impacts gene expression and alternative splicing (AS). We found that HE impacts expression of genes located within HE regions primarily via cis-acting dosage effect, which led to significant changes in the total expression of homoeolog pairs, especially for homoeologs whose original expression was biased. In parallel, HE influences expression of a large amount of genes residing in non-HE regions by trans-regulation leading to convergent expression of homoeolog pairs. Intriguingly, when taking into account of the original relative homoeolog expression states, homoeolog pairs under trans-effect are more prone to showing convergent response to HE whereas those under cis-effect trended to show subgenome-specific expression. Moreover, HE induced quantitative, largely individual-specific, changes of alternative splicing (AS) events. Like homoeologs expression, homoeo-AS events which related to trans effect were more responsive to HE. HE therefore exerts multifaceted immediate effects on gene expression and, to a less extent, also transcript diversity in nascent allopolyploidy.

The cardiac lncRNA Chantico directly regulates Cxcl1 chemokine transcription

In local gene regulation, long noncoding RNA (lncRNA) loci can function at three distinct, but non-mutually exclusive levels: the RNA molecule itself, the process of its transcription, and/or the underlying DNA element. Yet for cis-acting lncRNA, these distinctions are particularly challenging with present tools. To address this problem, we developed Omegazymes, catalytic nucleic acid enzymes to specifically target lncRNA for degradation without triggering premature termination of their transcription. We use Omegazymes to selectively target one of a highly refined set of lncRNAs in the mouse cardiomyocytes, demonstrating that the Chantico lncRNA molecule directly potentiates the transcription of Cxcl1, a neighboring chemokine gene. We find that the Chantico locus also acts at the DNA-level, as the binding of an essential cardiac transcription factor to the Chantico promoter is necessary for Cxcl1 transcription in mature cardiomyocytes. This Chantico regulation of Cxcl1 impacts cardiomyocyte signaling to immune cells, potentially regulating tissue-residence and inflammation.

A tandem duplication in Drosophila melanogaster shows enhanced expression beyond the gene copy number

Tandem duplicated genes are common features of genomes, but the phenotypic consequences of their origins are not well understood. It is not known whether a simple doubling of gene expression should be expected, or else some other expression outcome. This study describes an experimental framework using engineered deletions to assess any contribution of locally-acting cis- and globally-acting trans-regulatory factors to expression interactions of particular tandem duplicated genes. Acsx1L (CG6300) and Acsx1R (CG11659) are tandem duplicates of a putative acyl-CoA synthetase gene found in D. melanogaster. Experimental deletions of the duplicated segments were used to investigate whether the presence of one tandem duplicated block influences the expression of its neighbor. Acsx1L, the gene in the left block, shows much higher expression than either its duplicate Acsx1R or the single Acsx1 in D. simulans. Acsx1L expression decreases drastically upon deleting the right-hand duplicated block. Crosses among wildtype and deletion strains show that high tandem expression is primarily due to cis-acting interactions between the duplicated blocks. No effect of these genes on cuticular hydrocarbons was detected. Sequence and phylogenetic analysis suggest that the duplication rose to fixation in D. melanogaster and has been subject to extensive gene conversion. Some strains actually carry three tandem copies, yet strains with three Acsx1s do not have higher expression levels than strains with two. Surveys of tandem duplicate expression have typically not found the expected twofold increase in expression. This study suggests that cis-regulatory interactions between duplicated blocks could be responsible for this trend.

Comprehensive Analysis of Jumonji Domain C Family from Citrus grandis and Expression Profilings in the Exocarps of “Huajuhong” (Citrus grandis “Tomentosa”) during Various Development Stages

Citrus grandis “Tomentosa” (“Huajuhong”) is a famous Traditional Chinese Medicine. In this study, a total of 18 jumonji C (JMJC) domain-containing proteins were identified from C. grandis. The 18 CgJMJCs were unevenly located on six chromosomes of C. grandis. Phylogenetic analysis revealed that they could be classified into five groups, namely KDM3, KDM4, KDM5, JMJC, and JMJD6. The domain structures and motif architectures in the five groups were diversified. Cis-acting elements on the promoters of 18 CgJMJC genes were also investigated, and the abscisic acid-responsive element (ABRE) was distributed on 15 CgJMJC genes. Furthermore, the expression profiles of 18 CgJMJCs members in the exocarps of three varieties of “Huajuhong”, for different developmental stages, were examined. The results were validated by quantitative real-time PCR (qRT-PCR). The present study provides a comprehensive characterization of JMJC domain-containing proteins in C. grandis and their expression patterns in the exocarps of C. grandis “Tomentosa” for three varieties with various development stages.