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Phytoplasmas are pathogenic bacteria that reprogram plant host development for their own benefit. Previous studies have characterized a few different phytoplasma effector proteins that destabilize specific plant transcription factors. However, these are only a small fraction of the potential effectors used by phytoplasmas; therefore, the molecular mechanisms through which phytoplasmas modulate their hosts require further investigation. To obtain further insights into the phytoplasma infection mechanisms, we generated a protein-protein interaction network between a broad set of phytoplasma effectors and a large, unbiased collection of Arabidopsis thaliana transcription factors and transcriptional regulators. We found widespread, but specific, interactions between phytoplasma effectors and host transcription factors, especially those related to host developmental processes. In particular, many unrelated effectors target specific sets of TCP transcription factors, which regulate plant development and immunity. Comparison with other host-pathogen protein interaction networks shows that phytoplasma effectors have unusual targets, indicating that phytoplasmas have evolved a unique and unusual infection strategy. This study contributes a rich and solid data source that guides further investigations of the functions of individual effectors, as demonstrated for some herein. Moreover, the dataset provides insights into the underlying molecular mechanisms of phytoplasma infection., (© 2023 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

The Asteraceae is the largest angiosperm family with more than 25,000 species. Individual studies have shown that MADS-box and TCP transcription factors are regulators of the development and symmetry of flowers, contributing to their iconic flower-head (capitulum) and floret. However, a systematic study of MADS-box and TCP genes across the Asteraceae is lacking. We performed a comparative analysis of genome sequences of 33 angiosperm species including our de novo assembly of diploid sexual dandelion ( Taraxacum officinale ) and 11 other Asteraceae to investigate the lineage-specific evolution of MADS-box and TCP genes in the Asteraceae. We compared the phylogenomic results of MADS-box and TCP genes with their expression in T. officinale floral tissues at different developmental stages to demonstrate the regulation of genes with Asteraceae-specific attributes. Here, we show that MADS-box MIKC c and TCP-CYCLOIDEA ( CYC ) genes have expanded in the Asteraceae. The phylogenomic analysis identified AGAMOUS -like ( AG -like: SEEDSTICK [ STK ]-like), SEPALATA -like ( SEP 3-like), and TCP-PROLIFERATING CELL FACTOR ( PCF )-like copies with lineage-specific genomic contexts in the Asteraceae, Cichorioideae, or dandelion. Different expression patterns of some of these gene copies suggest functional divergence. We also confirm the presence and revisit the evolutionary history of previously named " Asteraceae-Specific MADS-box genes ( AS-MADS )." Specifically, we identify non-Asteraceae homologs, indicating a more ancient origin of this gene clade. Syntenic relationships support that AS-MADS is paralogous to FLOWERING LOCUS C ( FLC ) as demonstrated by the shared ancient duplication of FLC and SEP 3., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Xiong, Risse, Berke, Zhao, van de Geest, Oplaat, Busscher, Ferreira de Carvalho, van der Meer, Verhoeven, Schranz and Vijverberg.)

Apomixis, the clonal formation of seeds, is a rare yet widely distributed trait in flowering plants. We have isolated the PARTHENOGENESIS (PAR) gene from apomictic dandelion that triggers embryo development in unfertilized egg cells. PAR encodes a K2-2 zinc finger, EAR-domain protein. Unlike the recessive sexual alleles, the dominant PAR allele is expressed in egg cells and has a miniature inverted-repeat transposable element (MITE) transposon insertion in the promoter. The MITE-containing promoter can invoke a homologous gene from sexual lettuce to complement dandelion LOSS OF PARTHENOGENESIS mutants. A similar MITE is also present in the promoter of the PAR gene in apomictic forms of hawkweed, suggesting a case of parallel evolution. Heterologous expression of dandelion PAR in lettuce egg cells induced haploid embryo-like structures in the absence of fertilization. Sexual PAR alleles are expressed in pollen, suggesting that the gene product releases a block on embryogenesis after fertilization in sexual species while in apomictic species PAR expression triggers embryogenesis in the absence of fertilization., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Certain obligate parasites induce complex and substantial phenotypic changes in their hosts in ways that favor their transmission to other trophic levels. However, the mechanisms underlying these changes remain largely unknown. Here we demonstrate how SAP05 protein effectors from insect-vectored plant pathogenic phytoplasmas take control of several plant developmental processes. These effectors simultaneously prolong the host lifespan and induce witches' broom-like proliferations of leaf and sterile shoots, organs colonized by phytoplasmas and vectors. SAP05 acts by mediating the concurrent degradation of SPL and GATA developmental regulators via a process that relies on hijacking the plant ubiquitin receptor RPN10 independent of substrate ubiquitination. RPN10 is highly conserved among eukaryotes, but SAP05 does not bind insect vector RPN10. A two-amino-acid substitution within plant RPN10 generates a functional variant that is resistant to SAP05 activities. Therefore, one effector protein enables obligate parasitic phytoplasmas to induce a plethora of developmental phenotypes in their hosts., Competing Interests: Declaration of interests Three patent applications related to the work described here have been submitted by the John Innes Centre., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Long non-coding RNAs (lncRNAs) have emerged as new class of regulatory molecules in animals where they regulate gene expression at transcriptional and post-transcriptional level. Recent studies also identified lncRNAs in plant genomes, revealing a new level of transcriptional complexity in plants. Thousands of lncRNAs have been predicted in the Arabidopsis thaliana genome, but only a few have been studied in depth., Results: Here we report the identification of Arabidopsis lncRNAs that are expressed during the vegetative stage of development in either the shoot apical meristem or in leaves. We found that hundreds of lncRNAs are expressed in these tissues, of which 50 show differential expression upon an increase in ambient temperature. One of these lncRNAs, FLINC, is down-regulated at higher ambient temperature and affects ambient temperature-mediated flowering in Arabidopsis., Conclusion: A number of ambient temperature responsive lncRNAs were identified with potential roles in the regulation of temperature-dependent developmental changes, such as the transition from the vegetative to the reproductive (flowering) phase. The challenge for the future is to characterize the biological function and molecular mode of action of the large number of ambient temperature-regulated lncRNAs that have been identified in this study.

Various environmental signals integrate into a network of floral regulatory genes leading to the final decision on when to flower. Although a wealth of qualitative knowledge is available on how flowering time genes regulate each other, only a few studies incorporated this knowledge into predictive models. Such models are invaluable as they enable to investigate how various types of inputs are combined to give a quantitative readout. To investigate the effect of gene expression disturbances on flowering time, we developed a dynamic model for the regulation of flowering time in Arabidopsis thaliana. Model parameters were estimated based on expression time-courses for relevant genes, and a consistent set of flowering times for plants of various genetic backgrounds. Validation was performed by predicting changes in expression level in mutant backgrounds and comparing these predictions with independent expression data, and by comparison of predicted and experimental flowering times for several double mutants. Remarkably, the model predicts that a disturbance in a particular gene has not necessarily the largest impact on directly connected genes. For example, the model predicts that SUPPRESSOR OF OVEREXPRESSION OF CONSTANS (SOC1) mutation has a larger impact on APETALA1 (AP1), which is not directly regulated by SOC1, compared to its effect on LEAFY (LFY) which is under direct control of SOC1. This was confirmed by expression data. Another model prediction involves the importance of cooperativity in the regulation of APETALA1 (AP1) by LFY, a prediction supported by experimental evidence. Concluding, our model for flowering time gene regulation enables to address how different quantitative inputs are combined into one quantitative output, flowering time.

Tomato is one of the most cultivated vegetables in the world and an important ingredient of the human diet. Tomato breeders and growers face a continuous challenge of combining high quantity (production volume) with high quality (appearance, taste and perception for the consumers, processing quality for the processing industry). To improve the quality of tomato, it is important to understand the regulation of fruit development and of fruit cellular structure, which is in part determined by the sizes and numbers of cells within a tissue. The role of the cell cycle therein is poorly understood. Plant cyclin-dependent kinases (CDKs) are homologues of yeast cdc2, an important cell cycle regulator conserved throughout all eukaryotes. CDKA1 is constitutively expressed during the cell cycle and has dual functions in S- and M-phase progression. We have produced transgenic tomato plants with increased expression of CDKA1 under the control of the fruit-specific TPRP promoter, which despite a reduced number of seeds and diminished amount of jelly, developed fruits with weight and shape comparable to that of wild-type fruits. However, the phenotypic changes with regard to the pericarp thickness and placenta area were remarkable. Fruits of tomato plants with the highest expression of CDKA1 had larger septa and columella (placenta), compared with wild-type fruits. Our data demonstrate the possibility of manipulating the ratio between cell division and expansion by changing the expression of a key cell cycle regulator and probably its activity with substantial effects on structural traits of the harvested fruit., (© 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.)

Background: TCP proteins are plant-specific transcription factors, which are known to have a wide range of functions in different plant species such as in leaf development, flower symmetry, shoot branching, and senescence. Only a small number of TCP genes has been characterised from tomato (Solanum lycopersicum). Here we report several functional features of the members of the entire family present in the tomato genome., Results: We have identified 30 Solanum lycopersicum SlTCP genes, most of which have not been described before. Phylogenetic analysis clearly distinguishes two homology classes of the SlTCP transcription factor family - class I and class II. Class II differentiates in two subclasses, the CIN-TCP subclass and the CYC/TB1 subclass, involved in leaf development and axillary shoots formation, respectively. The expression patterns of all members were determined by quantitative PCR. Several SlTCP genes, like SlTCP12, SlTCP15 and SlTCP18 are preferentially expressed in the tomato fruit, suggesting a role during fruit development or ripening. These genes are regulated by RIN (RIPENING INHIBITOR), CNR (COLORLESS NON-RIPENING) and SlAP2a (APETALA2a) proteins, which are transcription factors with key roles in ripening. With a yeast one-hybrid assay we demonstrated that RIN binds the promoter fragments of SlTCP12, SlTCP15 and SlTCP18, and that CNR binds the SlTCP18 promoter. This data strongly suggests that these class I SlTCP proteins are involved in ripening. Furthermore, we demonstrate that SlTCPs bind the promoter fragments of members of their own family, indicating that they regulate each other. Additional yeast one-hybrid studies performed with Arabidopsis transcription factors revealed binding of the promoter fragments by proteins involved in the ethylene signal transduction pathway, contributing to the idea that these SlTCP genes are involved in the ripening process. Yeast two-hybrid data shows that SlTCP proteins can form homo and heterodimers, suggesting that they act together in order to form functional protein complexes and together regulate developmental processes in tomato., Conclusions: The comprehensive analysis we performed, like phylogenetic analysis, expression studies, identification of the upstream regulators and the dimerization specificity of the tomato TCP transcription factor family provides the basis for functional studies to reveal the role of this family in tomato development.

Growth of tomato fruits is determined by cell division and cell expansion, which are tightly controlled by factors that drive the core cell cycle. The cyclin-dependent kinases (CDKs) and their interacting partners, the cyclins, play a key role in the progression of the cell cycle. In this study the role of CDKA1, CDKB1, and CDKB2 in fruit development was characterized by fruit-specific overexpression and down-regulation. CDKA1 is expressed in the pericarp throughout development, but is strongly up-regulated in the outer pericarp cell layers at the end of the growth period, when CDKB gene expression has ceased. Overexpression of the CDKB genes at later stages of development and the down-regulation of CDKA1 result in a very similar fruit phenotype, showing a reduction in the number of cell layers in the pericarp and alterations in the desiccation of the fruits. Expression studies revealed that CDKA1 is down-regulated by the expression of CDKB1/2 in CDKB1 and CDKB2 overexpression mutants, suggesting opposite roles for these types of CDK proteins in tomato pericarp development.

Floral organs are specified by the combinatorial action of MADS-domain transcription factors, yet the mechanisms by which MADS-domain proteins activate or repress the expression of their target genes and the nature of their cofactors are still largely unknown. Here, we show using affinity purification and mass spectrometry that five major floral homeotic MADS-domain proteins (AP1, AP3, PI, AG, and SEP3) interact in floral tissues as proposed in the "floral quartet" model. In vitro studies confirmed a flexible composition of MADS-domain protein complexes depending on relative protein concentrations and DNA sequence. In situ bimolecular fluorescent complementation assays demonstrate that MADS-domain proteins interact during meristematic stages of flower development. By applying a targeted proteomics approach we were able to establish a MADS-domain protein interactome that strongly supports a mechanistic link between MADS-domain proteins and chromatin remodeling factors. Furthermore, members of other transcription factor families were identified as interaction partners of floral MADS-domain proteins suggesting various specific combinatorial modes of action.

MADS-domain transcription factors are essential for proper flower and seed development in angiosperms and their role in determination of floral organ identity can be described by the 'ABC model' of flower development. Recently, close relatives of the B-type genes were identified by phylogenetic studies, which are referred to as B(sister) (B(s)) genes. Here, we report the isolation and characterization of a MADS-box B(s) member from petunia, designated FBP24. An fbp24 knock-down line appeared to closely resemble the Arabidopsis B(s) mutant abs and a detailed and comparative analysis led to the conclusion that both FBP24 and ABS are necessary to determine the identity of the endothelial layer within the ovule. Protein interaction studies revealed the formation of higher-order complexes between B(s)-C-E and B(s)-D-E type MADS-box proteins, suggesting involvement of these specific complexes in determination of endothelium identity. However, although there are many similarities between the two genes and their products and functions, interestingly FBP24 cannot replace ABS in Arabidopsis. The results presented here demonstrate the importance of the comparative analysis of key regulatory genes in various model systems to fully understand all aspects of plant development.

Interactions between proteins are essential for their functioning and the biological processes they control. The elucidation of interaction maps based on yeast studies is a first step toward the understanding of molecular networks and provides a framework of proteins that possess the capacity and specificity to interact. Here, we present a comprehensive plant protein-protein interactome map of nearly all members of the Arabidopsis thaliana MADS box transcription factor family. A matrix-based yeast two-hybrid screen of >100 members of this family revealed a collection of specific heterodimers and a few homodimers. Clustering of proteins with similar interaction patterns pinpoints proteins involved in the same developmental program and provides valuable information about the participation of uncharacterized proteins in these programs. Furthermore, a model is proposed that integrates the floral induction and floral organ formation networks based on the interactions between the proteins involved. Heterodimers between flower induction and floral organ identity proteins were observed, which point to (auto)regulatory mechanisms that prevent the activity of flower induction proteins in the flower.

Over the last decade, the yeast two-hybrid system has become the tool to use for the identification of protein-protein interactions and recently, even complete interactomes were elucidated by this method. Nevertheless, it is an artificial system that is sensitive to errors resulting in the identification of false-positive and false-negative interactions. In this study, plant MADS box transcription factor interactions identified by yeast two-hybrid systems where studied in living plant cells by a technique based on fluorescence resonance energy transfer (FRET). Petunia MADS box proteins were fused to either cyan fluorescent protein or yellow fluorescent protein and transiently expressed in protoplasts followed by FRET-spectral imaging microscopy and FRET-fluorescence lifetime imaging microscopy to detect FRET and hence protein-protein interactions. All petunia MADS box heterodimers identified in yeast were confirmed in protoplasts. However, in contrast to the yeast two-hybrid results, homodimerization was demonstrated in plant cells for three petunia MADS box proteins. Heterodimers were identified between the ovule-specific MADS box protein FLORAL BINDING PROTEIN 11 and members of the petunia FLORAL BINDING PROTEIN 2 subfamily, which are also expressed in ovules, suggesting that these dimers play a role in ovule development. Furthermore, the role of dimerization in translocation of MADS box protein dimers to the nucleus is demonstrated, and the nuclear localization signal of MADS box proteins has been mapped to the N-terminal region of the MADS domain by means of mutant analyses.