Genome-Wide Identification and Expression Profiling Analysis of WOX Family Protein-Encoded Genes in Triticeae Species

The WOX family is a group of plant-specific transcription factors which regulate plant growth and development, cell division and differentiation. From the available genome sequence databases of nine Triticeae species, 199 putative WOX genes were identified. Most of the identified WOX genes were distributed on the chromosomes of homeologous groups 1 to 5 and originated via the orthologous evolution approach. Parts of WOX genes in Triticum aestivum were confirmed by the specific PCR markers using a set of Triticum. durum-T. aestivum genome D substitution lines. All of these identified WOX proteins could be grouped into three clades, similar to those in rice and Arabidopsis. WOX family members were conserved among these Triticeae plants; all of them contained the HOX DNA-binding homeodomain, and WUS clade members contained the characteristic WUS-box motif, while only WUS and WOX9 contained the EAR motif. The RNA-seq and qPCR analysis revealed that the TaWOX genes had tissue-specific expression feature. From the expression patterns of TaWOX genes during immature embryo callus production, TaWOX9 is likely closely related with the regulation of regeneration process in T. aestivum. The findings in this study could provide a basis for evolution and functional investigation and practical application of the WOX family genes in Triticeae species.

A novel ERF transcription factor, CmERF12, negatively influences chrysanthemum embryo development

Embryo abortion often occurs during distant hybridization events. Apetala 2/ethylene-responsive factor (AP2/ERF) proteins are key transcription factor (TF) regulators of plant development and stress resistance, but their roles in hybrid embryo development are poorly understood. We isolated a novel AP2/ERF TF, CmERF12, from chrysanthemum and showed that it adversely affects embryo development during distant hybridization. Transcriptome and real-time quantitative PCR data demonstrated that CmERF12 is expressed at significantly higher levels in aborted ovaries compared with normal ovaries. CmERF12 localizes to the cell nucleus and contains a conserved EAR motif that mediates its transcription repressor function in yeast and plant cells. We generated an amiR-CmERF12 transgenic Chrysanthemum morifolium (C.m.) var. ‘Yuhualuoying’ and conducted distant hybridization with the wild-type tetraploid, Chrysanthemum nankingense (C.n.), revealing that CmERF12 knockdown significantly promoted embryo development and increased the seed setting rates during hybridization. The expression of various embryo development-related genes was up-regulated in developing ovaries from the ♀amiR-CmERF12-C.m. × ♂C.n. cross. Furthermore, CmERF12 directly interacted with CmSUF4 and significantly reduced its ability to activate its target gene CmEC1. Overall, we invented an original method to overcome plant distant hybridization barriers and unraveled the mechanism by which CmERF12 negatively affects chrysanthemum embryo development.

The EAR Motif in the Arabidopsis MADS Transcription Factor AGAMOUS-Like 15 Is Not Necessary to Promote Somatic Embryogenesis

AGAMOUS-like 15 (AGL15) is a member of the MADS domain family of transcription factors (TFs) that can directly induce and repress target gene expression, and for which promotion of somatic embryogenesis (SE) is positively correlated with accumulation. An ethylene-responsive element binding factor-associated amphiphilic repression (EAR) motif of form LxLxL within the carboxyl-terminal domain of AGL15 was shown to be involved in repression of gene expression. Here, we examine whether AGL15′s ability to repress gene expression is needed to promote SE. While a form of AGL15 where the LxLxL is changed to AxAxA can still promote SE, another form with a strong transcriptional activator at the carboxy-terminal end, does not promote SE and, in fact, is detrimental to SE development. Select target genes were examined for response to the different forms of AGL15.

An updated census of the maize TIFY family

The TIFY gene family is a plant-specific gene family encoding a group of proteins characterized by its namesake, the conservative TIFY domain and members can be organized into four subfamilies: ZML, TIFY, PPD and JAZ (Jasmonate ZIM-domain protein) by presence of additional conserved domains. The TIFY gene family is intensively explored in several model and agriculturally important crop species and here, yet the composition of the TIFY family of maize has remained unresolved. This study increases the number of maize TIFY family members known by 40%, bringing the total to 47 including 38 JAZ, 5 TIFY, and 4 ZML genes. The majority of the newly identified genes were belonging to the JAZ subfamily, six of which had aberrant TIFY domains, suggesting loss JAZ-JAZ or JAZ-NINJA interactions. Six JAZ genes were found to have truncated Jas domain or an altered degron motif, suggesting resistance to classical JAZ degradation. In addition, seven membranes were found to have an LxLxL-type EAR motif which allows them to recruit TPL/TPP co-repressors directly without association to NINJA. Expression analysis revealed that ZmJAZ14 was specifically expressed in the seeds and ZmJAZ19 and 22 in the anthers, while the majority of other ZmJAZs were generally highly expressed across diverse tissue types. Additionally, ZmJAZ genes were highly responsive to wounding and JA treatment. This study provides a comprehensive update of the maize TIFY/JAZ gene family paving the way for functional, physiological, and ecological analysis.

Identification of EIL and ERF Genes Related to Fruit Ripening in Peach

Peach (Prunus persica) is a climacteric fruit with a relatively short shelf life due to its fast ripening or softening process. Here, we report the association of gene families encoding ethylene insensitive-3 like (EIL) and ethylene response factor (ERF) with fruit ripening in peach. In total, 3 PpEILs and 12 PpERFs were highly expressed in fruit, with the majority showing a peak of expression at different stages. All three EILs could activate ethylene biosynthesis genes PpACS1 and PpACO1. One out of the 12 PpERFs, termed PpERF.E2, is a homolog of ripening-associated ERFs in tomato, with a consistently high expression throughout fruit development and an ability to activate PpACS1 and PpACO1. Additionally, four subgroup F PpERFs harboring the EAR repressive motif were able to repress the PpACO1 promoter but could also activate the PpACS1 promoter. Promoter deletion assay revealed that PpEILs and PpERFs could participate in transcriptional regulation of PpACS1 through either direct or indirect interaction with various cis-elements. Taken together, these results suggested that all three PpEILs and PpERF.E2 are candidates involved in ethylene biosynthesis, and EAR motif-containing PpERFs may function as activator or repressor of ethylene biosynthesis genes in peach. Our study provides an insight into the roles of EILs and ERFs in the fruit ripening process.

ZmCLA4 regulates leaf angle through multiple plant hormone-mediated signal pathways in maize

Leaf angle in cereals is an important agronomic trait contributing to plant architecture and grain yield by determining the plant compactness. Although ZmCLA4 was identified to shape plant architecture by affecting leaf angle, the detailed regulatory mechanism of ZmCLA4 in maize remains unclear. ZmCLA4 was identified as a transcriptional repressor using the Gal4-LexA/UAS system and transactivation analysis in yeast. The DNA affinity purification (DAP)-seq assay showed that ZmCLA4 not only acts as a repressor containing the EAR motif (CACCGGAC), but was also found to have two new motifs, CCGARGS and CDTCNTC. On analyzing the ZmCLA4-bound targeted genes, we found that ZmCLA4, as a cross node of multiple plant hormone-mediated pathways, directly bound to ARF22 and IAA26 to regulate auxin transport and mediated brassinosteroid signaling by directly binding to BZR3 and 14-3-3. ZmCLA4 bound two WRKY genes involved with abscisic acid, two genes (CYP75B1, CYP93D1) involved with jasmonic acid, B3 involved in the response to ethylene, and thereby negatively regulated leaf angle formation. We built a new regulatory network for the ZmCLA4 gene controlling leaf angle in maize, which contributed to the understanding of ZmCLA4’s regulatory mechanism and will improve grain yields by facilitating optimization of plant architecture.

Fungal effector Jsi1 hijacks plant JA/ET signaling through Topless

Ustilago maydis (U. maydis) is the causal agent of maize smut disease. During the colonization process, the fungus secretes effector proteins which suppress immune responses and redirect the host-metabolism in favor of the pathogen. Here we describe a novel strategy by which U. maydis induces plant jasmonate/ethylene (JA/ET) hormone signaling and thereby biotrophic susceptibility. The U. maydis effector Jasmonate/Ethylene signaling inducer 1 (Jsi1) possesses an ethylene-responsive element binding factor-associated amphiphilic repression (EAR) motif, DLNxxP, which interacts with the second WD40 domain of the conserved plant co-repressor family Topless/Topless related (TPL/TPR). Jsi1 interaction with TPL/TPRs leads to derepression of the ethylene response factor (ERF) branch of the JA/ET signaling pathway, supporting biotrophic susceptibility. Jsi1 likely activates the ERF branch by interfering with the binding of endogenous DLNxxP motif-containing ERF transcription factors to TPL/TPR proteins. The identification of effector proteins possessing a DLNxxP motif in different fungal species with biotrophic and hemibiotrophic lifestyles together with the validation of the interaction between such effectors from Magnaporthe oryzae (M. oryzae), Sporisorium scitamineum (S. scitamineum), and S. reilianum with TPL proteins indicates the convergent evolution of effectors for modulating the TPL/TPR co-repressor hub.