Temporal and spatial regulation of DROOPING LEAF gene expression that promotes midrib formation in rice

Genes involved in the differentiation and development of tissues and organs are temporally and spatially regulated in plant development. The DROOPING LEAF (DL) gene, a member of the YABBY gene family, promotes midrib formation in the leaf and carpel specification in the flower. Consistent with these functions, DL is initially expressed in the central region of the leaf primordia (presumptive midrib) and in the presumptive carpel primordia in the meristem. To understand the regulatory mechanism underlying DL expression, we tried to identify cis-regulatory regions required for temporal and spatial expression of this gene. We found that the cis region responsible for the presumptive midrib-specific expression in the leaf primordia is located in intron 2. Next, we confined the region to a sequence of about 200bp, which corresponds to a conserved non-coding sequence (CNS) identified by phylogenetic footprinting. In addition, a sequence termed DG1, incorporating a 5' upstream region of about 7.4kb, and introns 1 and 2, was shown to be sufficient to induce DL in the presumptive midrib, and to suppress it in other regions in the leaf primordia. By contrast, the regulatory region required for carpel-specific expression was not included in the DG1 sequence. We modified Oryza sativa (rice) plant architecture by expressing an activated version of DL (DL-VP16) in a precise manner using the DG1 sequence: the resulting transgenic plant produced a midrib in the distal region of the leaf blade, where there is no midrib in wild type, and formed more upright leaves compared with the wild type.     

The YABBY gene DROOPING LEAF regulates carpel specification and midrib development in Oryza sativa

In this article, we report that carpel specification in the Oryza sativa (rice) flower is regulated by the floral homeotic gene DROOPING LEAF (DL) that is distinct from the well-known ABC genes. Severe loss-of-function mutations of DL cause complete homeotic transformation of carpels into stamens. Molecular cloning reveals that DL is a member of the YABBY gene family and is closely related to the CRABS CLAW (CRC) gene of Arabidopsis thaliana. DL is expressed in the presumptive region (carpel anlagen), where carpel primordia would initiate, and in carpel primordia. These results suggest that carpel specification is regulated by DL in rice flower development. Whereas CRC plays only a partial role in carpel identity, DL may have been recruited to have the more essential function of specifying carpels during the evolution of rice. We also show that DL interacts antagonistically with class B genes and controls floral meristem determinacy. In addition, severe and weak dl alleles fail to form a midrib in the leaf. The phenotypic analysis of dl mutants, together with analyses of the spatial expression patterns and ectopic expression of DL, demonstrate that DL regulates midrib formation by promoting cell proliferation in the central region of the rice leaf.     

SUPERWOMAN1 and DROOPING LEAF genes control floral organ identity in rice

We analyzed recessive mutants of two homeotic genes in rice, SUPERWOMAN1 (SPW1) and DROOPING LEAF (DL). The homeotic mutation spw1 transforms stamens and lodicules into carpels and palea-like organs, respectively. Two spw1 alleles, spw1-1 and spw1-2, show the same floral phenotype and did not affect vegetative development. We show that SPW1 is a rice APETALA3 homolog, OsMADS16. In contrast, two strong alleles of the dl locus, drooping leaf-superman1 (dl-sup1) and drooping leaf-superman2 (dl-sup2), cause the complete transformation of the gynoecium into stamens. In these strong mutants, many ectopic stamens are formed in the region where the gynoecium is produced in the wild-type flower and they are arranged in a non-whorled, alternate pattern. The intermediate allele dl-1 (T65), results in an increase in the number of stamens and stigmas, and carpels occasionally show staminoid characteristics. In the weakest mutant, dl-2, most of the flowers are normal. All four dl alleles cause midrib-less drooping leaves. The flower of the double mutant, spw1 dl-sup, produces incompletely differentiated organs indefinitely after palea-like organs are produced in the position where lodicules are formed in the wild-type flower. These incompletely differentiated organs are neither stamens nor carpels, but have partial floral identity. Based on genetic and molecular results, we postulate a model of stamen and carpel specification in rice, with DL as a novel gene controlling carpel identity and acting mutually and antagonistically to the class B gene, SPW1.     

The DROOPING LEAF and OsETTIN2 genes promote awn development in rice

The awn is a long needle‐like appendage that, in some grass species, is formed on the lemma that encloses floral organs together with the palea. In rice, most wild species and most strains of Oryza sativa ssp. indica generate an awn, whereas most strains of O. sativa ssp. japonica do not. In japonica, the long‐awn characteristic appears to have been lost during domestication and breeding programs. Here, we found that the genes DROOPING LEAF (DL) and OsETTIN2 (OsETT2) are involved in awn development in the awned indica strain Kasalath. Genetic analyses and RNA‐silencing experiments indicate that DL and OsETT2 act independently in awn formation, and that either gene alone is not sufficient for awn development. Scanning electron microscopy revealed that the top region of the lemma (a putative awn primordium) is larger in an awned floret than in an awnless floret. OsETT2 is expressed in the awn primordium in the awned indica floret, but not in the awnless japonica floret except in the provascular bundle. DL is expressed underneath the primordium at similar levels in both indica and japonica florets, suggesting non‐cell‐autonomous action. We hypothesize that loss of expression of OsETT2 in the awn primordium is probably associated with the failure of awn formation in japonica strains.     

Genome-wide identification and expression analysis of rice cell cycle genes

Cyclins, cyclin-dependent kinases, and a number of other proteins control the progression of plant cell cycle. Although extensive studies have revealed the roles of some cell cycle regulators and the underlying mechanisms in Arabidopsis, relatively a small number of cell cycle regulators were functionally analyzed in rice. In this study, we describe 41 regulators in the rice genome. Our results indicate that the rice genome contains a less number of the core cell cycle regulators than the Arabidopsis one does, although the rice genome is much larger than the Arabidopsis one. Eight groups of CDKs similar to those in Arabidopsis were identified in the rice genome through phylogenetic analysis, and the corresponding members in the different groups include E2F, CKI, Rb, CKS and Wee. The structures of the core cell regulators were relatively conserved between the rice and Arabidopsis genomes. Furthermore, the expression of the majority of the core cell cycle genes was spatially regulated, and the most closely related ones showed very similar patterns of expression, suggesting functional redundancy and conservation between the highly similar core cell cycle genes in rice and Arabidopsis. Following auxin or cytokinin treatment, the expression of the core cell cycle genes was either upregulated or downregulated, suggesting that auxin and/or cytokinin may directly regulate the expression of the core cell cycle genes. Our results provide basic information to understand the mechanism of cell cycle regulation and the functions of the rice cell cycle genes. Electronic supplementary material The online version of this article (doi: ) contains supplementary material, which is available to authorized users. Jing Guo and Jian Song have contributed equally.     

Genome-wide identification and expression profiling of ankyrin-repeat gene family in maize

Members of the ankyrin repeats (ANK) gene family encode ANK domain that are common in diverse organisms and play important roles in cell growth and development, such as cell-cell signal transduction and cell cycle regulation. Recently, genome-wide identification and evolutionary analyses of the ANK gene family have been carried out in Arabidopsis and rice. However, little is known regarding the ANK genes in the entire maize genome. In this study, we described the identification and structural characterization of 71 ANK genes in maize (ZmANK). Then, comprehensive bioinformatics analyses of ZmANK genes family were performed including phylogenetic, domain and motif analysis, chromosomal localization, intron/exon structural patterns, gene duplications and expression profiling. Domain composition analyses showed that ZmANK genes formed ten subfamilies. Five tandem duplications and 14 segmental duplications were identified in ZmANK genes. Furthermore, we took comparative analysis of the total ANK gene family in Arabidopsis, rice and maize, ZmANKs were more closely paired with OsANKs than with AtANKs. At last, expression profile analyses were performed. Forty-one members of ZmANK genes held EST sequences records. Semi-quantitative expression and microarray data analysis of these 41 ZmANK genes demonstrated that ZmANK genes exhibit a various expression pattern, suggesting that functional diversification of ZmANK genes family. The results will present significant insights to explore ANK genes expression and function in future studies in maize.     

Genome-wide identification and expression profiling of DNA methyltransferase gene family in maize

Key message

In this study, we identified eight DNA MTase genes in maize and the diversity of expression patterns of them was presented by EST mining, microarray and semi-quantitative expression profile analyses.

Abstract

DNA methylation plays a pivotal role in promoting genomic stability through diverse biological processes including regulation of gene expression during development and chromatin organization. Although this important biological process is mainly regulated by several conserved Cytosine-5 DNA methyltransferases encoded by a smaller multigene family in plants, investigation of the plant C5-MTase-encoding gene family will serve to elucidate the epigenetic mechanism diversity in plants. Recently, genome-wide identification and evolutionary analyses of the C5-MTase-encoding gene family have been characterized in multiple plant species including Arabidopsis, rice, carrot and wheat. However, little is known regarding the C5-MTase-encoding genes in the entire maize genome. Here, genome-wide identification and expression profile analyses of maize C5-MTase-encoding genes (ZmMETs) were performed from the latest version of the maize (B73) genome. Phylogenetic analysis indicated that the orthologs from the three species (maize, Arabidopsis and rice) were categorized into four classes. Chromosomal location of these genes revealed that they are unevenly distributed on 6 of all 10 chromosomes with three chromosomal/segmental duplication events, suggesting that gene duplication played a key role in expansion of the maize C5-MTase-encoding gene family. Furthermore, EST expression data mining, microarray data and semi-quantitative expression profile analyses detected in the leaves by two different abiotic stress treatments have demonstrated that these genes had temporal and spatial expression pattern and exhibited different expression levels in stress treatments, suggesting that functional diversification of ZmMET genes family. Overall, our study will serve to present signification insights to explore the plant C5-MTase-encoding gene expression and function and also be beneficial for future experimental research to further unravel the mechanisms of epigenetic regulation in plants.     

Genome-wide identification of the class III aminotransferase gene family in rice and expression analysis under abiotic stress

Aminotransferases are pyridoxal 5′-phosphate-dependent enzymes that play crucial roles in plant growth, development, and responses to abiotic stress. The class III aminotransferase family (ATIII family) is an important subfamily. However, no characterization of rice ATIII genes has been previously reported. Using available rice genome sequence information, we identified 12 japonica and 13 indica ATIII genes that were randomly localized on chromosomes 2, 3, 4, 5, 7, 8, and 11. Information provided by the Plant Genome Duplication Database revealed that four japonica and four indica ATIII genes are the results of segmental duplications, and two japonica and six indica genes resulted from tandem duplications. A phylogenetic analysis of the ATIII genes in japonica, indica and Arabidopsis enabled the classification of the genes into six different groups, and the characteristics were established before the monocot-dicot and japonica–indica split. An analysis of the Ka/Ks, divergence time and average indel length suggested the diverse selection styles of the duplicated gene pairs. Gene structure and motif analyses revealed that the ATIII gene family has experienced extensive divergence. Real-time PCR was performed to examine the expression pattern of the japonica ATIII genes in response to various abiotic stresses including drought, salt, and cold. The results suggested that most of the genes were differentially up- or down-regulated in rice seedlings in response to at least one stress factor, which indicates the key role of the rice ATIII gene family in responding to abiotic stresses. These results provide a basis for elucidating the roles of the ATIII genes and their further functional analysis under abiotic stresses.     

番茄WOX转录因子家族的鉴定及其进化、表达分析

WUSCHEL相关的同源异型盒(WUSCHEL-related homeobox,WOX)是一类植物特异的转录因子家族,具有调控植物干细胞分裂分化动态平衡等重要功能。本研究利用番茄(Solanum lycopersicum)基因组数据,通过建立隐马尔科夫模型并进行检索,鉴定了番茄10个WOX转录因子家族成员。多序列比对发现,番茄WOX转录因子家族成员具有高度保守的同源异型结构域;以拟南芥WOX转录因子家族成员序列为参照,通过邻接法、极大似然法、贝叶斯法重建了系统发育树,三者呈现出类似的拓扑结构,番茄和拟南芥WOX转录因子家族共25个成员被分为3个进化支(Clade)和9个亚家族(Subgroup);利用MEME和GSDS对WOX转录因子家族成员的蛋白保守结构域和基因结构进行了分析,同一亚家族内的WOX转录因子家族成员的保守结构域的种类、组织形式以及基因结构具有高度的一致性;利用Perl和Orthomcl对家族成员的染色体定位和同源性关系进行分析,结果表明串联重复的SlWOX3a和SlWOX3b可能来源于一次复制事件;利用番茄转录组数据和qRT-PCR进行表达分析,结果显示家族成员在不同组织中的表达存在差异,暗示了WOX家族的不同成员在功能上可能具有多样性。本研究对番茄WOX转录因子家族成员进行GO(Gene Ontology)注释和比较分析,结果表明该家族成员作为转录因子,可能在组织器官发育、细胞间通讯等过程中发挥作用。     

Genome-wide expression analysis of HSP70 family genes in rice and identification of a cytosolic HSP70 gene highly induced under heat stress

The heat shock protein 70 (HSP70) gene family plays a key role in protecting plant cells or tissues from thermal or oxidative stress. Although many studies have elucidated the molecular functions of individual family members, genome-wide analysis of this family is still limited, especially for crop species. Our objective was to integrate various meta-profiling data into the context of a phylogenetic tree, which would enable us to perform fine evaluation of functional dominancy or redundancy within this family. Our data indicated that a loss-of-function mutant of a rice cytosolic HSP70 gene (OsctHSP70-1) did not show a clear defective phenotype in response to high temperature because of the existence of another gene family member that was closely clustered with OsctHSP70-1 and had similar expression patterns. Moreover, the second gene showed much stronger anatomical expression. We indirectly analyzed the function of OsctHSP70-1 by studying GUS activity under the control of the endogenous promoter. We also designed a probable interaction network mediated by OsctHSP70-1 and used co-expression analysis among its components to refine the network, suggesting more probable model to explain the function of OsctHSP70-1.