CLASS-II KNOX genes coordinate spatial and temporal patterns of the tomato ripening

Ripening is a complex developmental change of a mature organ, the fruit. In plants like a tomato, it involves softening, pigmentation, and biosynthesis of metabolites beneficial for the human diet. Examination of the transcriptional changes towards ripening suggests that redundant uncharacterized factors may be involved in the coordination of the ripening switch. Previous studies have demonstrated that Arabidopsis CLASS-II KNOX genes play a significant role in controlling the maturation of siliques and their transition to senescence. Here we examined the combined role of all four tomato CLASS-II KNOX genes in the maturation and ripening of fleshy fruits using an artificial microRNA targeting them simultaneously. As expected, the knockdown plants (35S::amiR-TKN-CL-II) exhibited leaves with increased complexity, reminiscent of the leaf phenotype of plants overexpressing CLASS- I KNOX, which antagonize CLASS-II KNOX gene functions. The fruits of 35S::amiR-TKN-CL-II plants were notably smaller than the control. While their internal gel/placenta tissue softened and accumulated the typical pigmentation, the pericarp color break took place ten days later than control, and eventually, it turned yellow instead of red. Additionally, the pericarp of 35S::amiR-TKN-CL-II fruits remained significantly firmer than control even after three weeks of shelf storage. Strikingly, the 35S::amiR-TKN-CL-II fruits showed early ethylene release and respiration peak, but these were correlated only with liquefaction and pigmentation of the internal tissues. Our findings suggest that CLASS-II KNOX genes are required to coordinate the spatial and temporal patterns of tomato fruit ripening.

In Vitro Plant Regeneration in Conifers: The Role of WOX and KNOX Gene Families

Conifers are a group of woody plants with an enormous economic and ecological importance. Breeding programs are necessary to select superior varieties for planting, but they have many limitations due to the biological characteristics of conifers. Somatic embryogenesis (SE) and de novo organogenesis (DNO) from in vitro cultured tissues are two ways of plant mass propagation that help to overcome this problem. Although both processes are difficult to achieve in conifers, they offer advantages like a great efficiency, the possibilities to cryopreserve the embryogenic lines, and the ability of multiplying adult trees (the main bottleneck in conifer cloning) through DNO. Moreover, SE and DNO represent appropriate experimental systems to study the molecular bases of developmental processes in conifers such as embryogenesis and shoot apical meristem (SAM) establishment. Some of the key genes regulating these processes belong to the WOX and KNOX homeobox gene families, whose function has been widely described in Arabidopsis thaliana. The sequences and roles of these genes in conifers are similar to those found in angiosperms, but some particularities exist, like the presence of WOXX, a gene that putatively participates in the establishment of SAM in somatic embryos and plantlets of Pinus pinaster.

A gene expression atlas for kiwifruit (Actinidia chinensis) and network analysis of transcription factors

Background Transcriptomic studies combined with a well annotated genome have laid the foundations for new understanding of molecular processes. Tools which visualise gene expression patterns have further added to these resources. The manual annotation of the Actinidia chinensis (kiwifruit) genome has resulted in a high quality set of 33,044 genes. Here we investigate gene expression patterns in diverse tissues, visualised in an Electronic Fluorescent Pictograph (eFP) browser, to study the relationship of transcription factor (TF) expression using network analysis. Results Sixty-one samples covering diverse tissues at different developmental time points were selected for RNA-seq analysis and an eFP browser was generated to visualise this dataset. 2839 TFs representing 57 different classes were identified and named. Network analysis of the TF expression patterns separated TFs into 14 different modules. Two modules consisting of 237 TFs were correlated with floral bud and flower development, a further two modules containing 160 TFs were associated with fruit development and maturation. A single module of 480 TFs was associated with ethylene-induced fruit ripening. Three “hub” genes correlated with flower and fruit development consisted of a HAF-like gene central to gynoecium development, an ERF and a DOF gene. Maturing and ripening hub genes included a KNOX gene that was associated with seed maturation, and a GRAS-like TF. Conclusions This study provides an insight into the complexity of the transcriptional control of flower and fruit development, as well as providing a new resource to the plant community. The Actinidia eFP browser is provided in an accessible format that allows researchers to download and work internally.

What can the phylogeny of class I KNOX genes and their expression patterns in land plants tell us about the evolution of shoot development?

KNOX genes encode transcription factors (TFs), several of which act non-cell-autonomously. KNOX genes evolved in algae, and two classes, class I KNOX and class II KNOX genes, were already present in charophytes. In tracheophytes, class I KNOX genes are expressed in shoot apical meristems (SAMs) and thought to inhibit cell differentiation, whereas class II KNOX genes are expressed in mature organs regulating differentiation. In this review, we summarize the data available on gene families and expression patterns of class I and class II KNOX genes in embryophytes. The expression patterns of class I KNOX genes should be seen in the context of SAM structure and of leaf primordium development where the inhibition of cell differentiation needs to be lifted. Although the SAMs of angiosperms and gnetophytes almost always belong to the duplex type, several other types are distributed in gymnosperms, ferns, lycopods and bryophytes. KNOX gene families remained small (maximally five genes) in the representatives of bryophytes, lycopods and ferns examined thus far; however, they expanded to some extent in gymnosperms and, independently and much more strongly, in angiosperms. The growing sophistication of mechanisms to repress and re-induce class KNOX I expression played a major role in the evolution of leaf shape.

An Actinidia chinensis (kiwifruit) eFP browser and network analysis of transcription factors

BackgroundTranscriptomic studies combined with a well annotated genome have laid the foundations for new understanding of molecular processes. Tools which visualise gene expression patterns have further added to these resources. The manual annotation of the Actinidia chinensis (kiwifruit) genome has resulted in a high quality set of 33,044 genes. Here we investigate gene expression patterns in diverse tissues, visualised in an Electronic Fluorescent Pictograph (eFP) browser, to study the relationship of transcription factor (TF) expression using network analysis. ResultsSixty-one samples covering diverse tissues at different developmental time points were selected for RNAseq analysis and an eFP browser was generated to visualise this dataset. 2,839 TFs representing 57 different classes were identified and named. Network analysis of the TF expression patterns separated TFs into 14 different modules. Two modules consisting of 237 TFs were correlated with floral bud and flower development, a further two modules containing 160 TFs were associated with fruit development and maturation. A single module of 480 TFs was associated with ethylene-induced fruit ripening. Three “hub” genes correlated with flower and fruit development consisted of a HAF-like gene central to gynoecium development, an ERF and a DOF gene. Maturing and ripening hub genes included a KNOX gene that was associated with seed maturation, and a GRAS-like TF.ConclusionsThis study provides an insight into the complexity of the transcriptional control of flower and fruit development, as well as providing a new resource to the plant community. The eFP browser is provided in an accessible format that allows researchers to download and work internally.

Functional divergence and adaptive selection of KNOX gene family in plants

KNOTTED-like homeodomain (KNOX) genes are transcriptional regulators that play an important role in morphogenesis. In the present study, a comparative analysis was performed to investigate the molecular evolution of the characteristics of the KNOX gene family in 10 different plant species. We identified 129 KNOX gene family members, which were categorized into two subfamilies based on multiple sequence alignment and phylogenetic tree reconstruction. Several segmental duplication pairs were found, indicating that different species share a common expansion model. Functional divergence analysis identified the 15 and 52 amino acid sites with significant changes in evolutionary rates and amino acid physicochemical properties as functional divergence sites. Additional selection analysis showed that 14 amino acid sites underwent positive selection during evolution, and two groups of co-evolutionary amino acid sites were identified by Coevolution Analysis using Protein Sequences software. These sites could play critical roles in the molecular evolution of the KNOX gene family in these species. In addition, the expression profiles of KNOX duplicated genes demonstrated functional divergence. Taken together, these results provide novel insights into the structural and functional evolution of the KNOX gene family.

Overexpression of a Novel LcKNOX Transcription Factor from Liriodendron chinense Induces Lobed Leaves in Arabidopsis thaliana

Liriodendron chinense is a common ornamental tree that has attractive leaves, which is a valuable trait for use in landscape architecture. In this work, we aimed to identify the potential genes that control and regulate the development of L. chinense leaf lobes. Based on the transcriptome data for the leaf developmental stages we previously generated, two candidate genes were identified in this study. KNOTTED-LIKE HOMEOBOX(KNOX), encoding homeobox family proteins, play a large role in leaf lobe and leaf complexity regulation. Here, two full length KNOX genes from L. chinense were amplified and named LcKNOX1 and LcKNOX6 according to their sequence similarities with the respective Arabidopsis thaliana KNOX family genes. Overexpression vectors were constructed and subsequently transformed into wild type (WT) A. thaliana. Additionally, LcKNOX6 was expressed in tobacco leaves to examine its subcellular localization, and the 35S::LcKNOX6 transgenic A. thaliana leaf cells were imaged with the use of SEM. The expression of several genes that participate in KNOX gene regulation were validated by quantitative real-time PCR. The results show that LcKNOX1 produces almost the same phenotype as that found in WT A. thaliana. Notably, the LcKNOX6-1 lines presented deep leaf lobes that were similar to L. chinense leaf lobes. Two 35S::LcKNOX6 lines induced an abnormal growth phenotype whose seeds were abortive. In short, these results indicate that the LcKNOX6 gene might affect leaf development in A. thaliana and provide insights into the regulation of L. chinense leaf shaping.