The dynamics of HD-ZIP III - ZPR protein interactions play essential roles in embryogenesis, meristem function and organ development

Maintaining a stem cell population while developing lateral organs is a crucial aspect of plant development. Studies have shown that a family of micro proteins, the LITTLE ZIPPERS (ZPR), are involved in this process by repressing the activity of HD-ZIP III transcription factors. However, the unique role of each ZPR has not been thoroughly characterized. In this work, we use genetics, imaging, and biochemistry to create a detailed picture of ZPR family expression and their specific interactions with HD-ZIP IIIs. CRISPR/Cas9 was implemented to isolate single loss-of-function ZPR alleles as well as higher-order mutant combinations. A single mutation in ZPR1, ZPR3, and ZPR4 affects the development of the cotyledons during embryogenesis. Additionally, double mutant analyses indicates both unique roles for each ZPR protein as well as redundancy. Using ZPR tagged lines we show that while ZPR3 and ZPR4 have a similar pattern of accumulation as the HD-ZIP IIIs, ZPR1 and ZPR2 accumulation is more limited. Immuno-precipitations (IP) with tagged ZPR proteins are mainly enriched with the anticipated HD-ZIP III partners. Although ZPRs interact with all HD-ZIP IIIs, an apparent preference of heterodimer formation with REVOLUTA is observed. Our work highlights that the dynamics of ZPR protein accumulation together with the strength of ZPR-HD-ZIP III interactions provide an added layer of complexity to the regulation of HD-ZIP IIIs during plant development.

Genome-wide identification and analysis of the YABBY gene family in Moso Bamboo (Phyllostachys edulis (Carrière) J. Houz)

Background The YABBY gene family is a family of small zinc finger transcription factors associated with plant morphogenesis, growth, and development. In particular, it is closely related to the development of polarity in the lateral organs of plants. Despite being studied extensively in many plant species, there is little information on genome-wide characterization of this gene family in Moso bamboo. Methods In the present study, we identified 16 PeYABBY genes, which were unequally distributed on 11 chromosomes, through genome-wide analysis of high-quality genome sequences of M oso bamboo by bioinformatics tools and biotechnological tools. Gene expression under hormone stress conditions was verified by quantitative real-time PCR (qRT-PCR) experiments. Results Based on peptide sequences and similarity of exon-intron structures, we classified the PeYABBY genes into four subfamilies. Analysis of putative cis-acting elements in promoters of these genes revealed that PeYABBYs contained a large number of hormone-responsive and stress-responsive elements. Expression analysis showed that they were expressed at a high level in Moso bamboo panicles, rhizomes, and leaves. Expression patterns of putative PeYABBY genes in different organs and hormone-treated were analyzed using RNA-seq data, results showed that some PeYABBY genes were responsive to gibberellin (GA) and abscisic acid (ABA), indicating that they may play an important role in plant hormone responses. Gene Ontology (GO) analyses of YABBY proteins indicated that they may be involved in many developmental processes, particularly high level of enrichment seen in plant leaf development. In summary, our results provide a comprehensive genome-wide study of the YABBY gene family in bamboos, which could be useful for further detailed studies of the function and evolution of the YABBY genes, and to provide a fundamental basis for the study of YABBY in Gramineae for resistance to stress and hormonal stress.

Importance of cell division angle, position of cell proliferative area, and localization of AN3 in lateral organ morphology

Leaf meristem is a cell proliferative zone present in the lateral organ primordia, and it contributes to the expansion of lateral organ lamina. In this study, we investigated how the proliferative zone affects the final morphology of the lateral organs. We examined how cell proliferative zones differ in the primordia of polar-auxin transport inhibitor (PATI)-treated leaves and floral organs from normal foliage leaf primordia of Arabidopsis thaliana with focus on the spatial accumulation pattern of mRNA and protein of ANGUSTIFOLIA3 (AN3), a key element for leaf meristem positioning. As a result, we revealed that organ shape change by PATI treatment could not be attributed to changes in leaf-meristem positioning, size of the leaf meristem, or the expression pattern of AN3. Instead, it was attributed to altered cell division angles in the leaf meristem. In contrast, different shapes between sepals and petals compared with foliage leaves were observed to be correlated with both altered meristem position associated with altered AN3 expression patterns and different distributions of cell division angles. These results strongly indicate that lateral organ shapes are regulated via two aspects: position of meristem and cell division angles; the former is mainly governed by the AN3 expression pattern.

New and interesting palpigrades (Arachnida, Palpigradi) of the genera Koeneniodes Silvestri, 1913 and Prokoenenia Börner, 1901 from Asia

Two new species of palpigrades are described: a soil-dwelling species of the genus Koeneniodes Silvestri, 1913 from a broadleaf forest in Tibet and an extraordinary cave-dwelling species from Jinhua cave in China belonging to Prokoenenia Börner, 1901. Koeneniodes tibetanus sp. n. is related to Koeneniodes spiniger from Thailand. The two species share the presence of four thick and spiniform setae on the second lobe of the female genitalia; they differ in the number of thick setae on opisthosomal sternite IV, the number of cheliceral teeth, the coxal setal formula, and the morphology of the spiniform setae. Prokoenenia sarcodactylica sp. n. is based on an immature female from Jinhua Cave, Beijing. The presence of 18 finger-shaped blades in the lateral organs—unique among palpigrades –, the large body size (2150 μm) and the extremely long basitarsus IV (205 μm) indicate that the new species is the first undoubtedly cave–adapted Prokoenenia. This is also the first record of the genus Prokoenenia from China.

Arabidopsis ABIG1 Functions in Laminar Growth and Polarity Formation through Regulation by REVOLUTA and KANADI

Leaf laminar growth and adaxial-abaxial boundary formation are fundamental outcomes of plant development. Boundary and laminar growth coordinate the further patterning and growth of the leaf, directing the differentiation of cell types within the top and bottom domains and promoting initiation of lateral organs along their adaxial/abaxial axis. Leaf adaxial-abaxial polarity specification and laminar out-growth are regulated by two transcription factors, REVOLUTA (REV) and KANADI (KAN). ABA INSENSITIVE TO GROWTH 1 (ABIG1) is a HOMEODOMAIN-LEUCINE ZIPPER (HD-ZIP) Class II transcription factor and is a direct target of the adaxial-abaxial regulators REV and KAN. To investigate the role of ABIG1 in the leaf development and establishment of polarity, we examined the phenotypes of both gain-of-function and loss-of-function mutants. Through genetic interaction analysis with REV and KAN mutants, we have determined that ABIG1 plays a role in leaf laminar-growth as well as in adaxial-abaxial polarity establishment. Genetic and physical interaction assays showed that ABIG1 interacts with the transcriptional corepressor TOPLESS (TPL). This study provides new evidence that another HD-ZIP II gene, ABIG1, facilitates growth through the corepressor TPL.

LsaYAB7, A Homologous Gene of FILAMENTOUS FLOWER, Participating In The Regulation of Adaxial-Abaxial Polarity of Leaves In Lettuce

Background: The plant-specific YABBY transcription factor plays important roles in the development of lateral organs, the establishment of adaxial-abaxial polarity and abiotic stress response. However, the function of YABBY gene family in lettuce (Lactuca sativa), an important leaf vegetable is still unclear yet. Results: In this study, we analyzed the chromosomal distribution, gene structure and cis-transcriptional elements of the YABBY gene family in lettuce. Phylogenetic analysis of YABBY genes in lettuce, Arabidopsis, maize and rice classified them into five groups. In addition, the expression profile of YABBY genes in lettuce indicated that they may play distinct functions in different tissues and developmental processes. The subcellular localization analysis and transactivation assay showed that LsaYAB7 probably functions as a transcription factor in the nucleus. Furthermore, the ectopic expression of LsaYAB7 exhibited abaxially curled leaves, not only in Arabidopsis, but also in lettuce. Conclusion: Altogether, LsaYAB7 is a key functional gene in determining the adaxial-abaxial polarity of lettuce leaves. Our study laid foundation for the molecular research of functional genes in lettuce.

A Role for Auxin in Triggering Lamina Outgrowth of Unifacial Leaves

A common morphological feature of typical angiosperms is the patterning of lateral organs along primary axes of asymmetry—a proximodistal, a mediolateral, and an adaxial–abaxial axis. Angiosperm leaves usually have distinct adaxial–abaxial identity, which is required for the development of a flat shape. By contrast, many unifacial leaves, consisting of only the abaxial side, show a flattened morphology. This implicates a unique mechanism that allows leaf flattening independent of adaxial–abaxial identity. In this study, we report a role for auxin in outgrowth of unifacial leaves. In two closely related unifacial-leaved species of Juncaceae, Juncus prismatocarpus with flattened leaves, and J. wallichianus with transversally radialized leaves, the auxin-responsive gene GLYCOSIDE HYDROLASE3 (GH3) displayed spatially different expression patterns within leaf primordia. Treatment of J. prismatocarpus seedlings with exogenous auxin or auxin transport inhibitors, which disturb endogenous auxin distribution, eliminated leaf flatness, resulting in a transversally radialized morphology. These treatments did not affect the radialized morphology of leaves of J. wallichianus. Moreover, elimination of leaf flatness by these treatments accompanied dysregulated expression of genetic factors needed to specify the leaf central-marginal polarity in J. prismatocarpus. The findings imply that lamina outgrowth of unifacial leaves relies on proper placement of auxin, which might induce initial leaf flattening and subsequently act to specify leaf polarity, promoting further flattening growth of leaves.

Gene duplication at the Fascicled ear1 locus controls the fate of inflorescence meristem cells in maize

Plant meristems are self-renewing groups of pluripotent stem cells that produce lateral organs in a stereotypical pattern. Of interest is how the radially symmetrical meristem produces laminar lateral organs. Both the male and female inflorescence meristems of the dominant Fascicled ear (Fas1) mutant fail to grow as a single point and instead show deep branching. Positional cloning of two independent Fas1 alleles identified an ∼160 kb region containing two floral genes, the MADS-box gene, zmm8, and the YABBY gene, drooping leaf2 (drl2). Both genes are duplicated within the Fas1 locus and spatiotemporally misexpressed in the mutant inflorescence meristems. Increased zmm8 expression alone does not affect inflorescence development; however, combined misexpression of zmm8, drl2, and their syntenic paralogs zmm14 and drl1, perturbs meristem organization. We hypothesize that misexpression of the floral genes in the inflorescence and their potential interaction cause ectopic activation of a laminar program, thereby disrupting signaling necessary for maintenance of radially symmetrical inflorescence meristems. Consistent with this hypothesis, RNA sequencing and in situ analysis reveal altered expression patterns of genes that define distinct zones of the meristem and developing leaf. Our findings highlight the importance of strict spatiotemporal patterns of expression for both zmm8 and drl2 and provide an example of phenotypes arising from tandem gene duplications.

Overexpression of a Pak Choi Gene, BcAS2, Causes Leaf Curvature in Arabidopsis thaliana

The LBD (Lateral Organ Boundaries Domain) family are a new group of plant-specific genes, which encode a class of transcription factors containing conserved Lateral Organization Boundary (LOB) domains, and play an important role in regulating the adaxial–abaxial polarity of plant leaves. In Arabidopsis thaliana, ASYMMETRIC LEAVES 2 (AS2) has a typical LOB domain and is involved in determining the adaxial cell fate. In this study, we isolated the BcAS2 gene from the pak choi cultivar “NHCC001”, and analyzed its expression pattern. The results showed that the BcAS2 encoded a protein made up of 202 amino acid residues which were located in the nucleus and cytomembrane. The Yeast two-hybrid system (Y2H) assay indicated that BcAS2 interacts with BcAS1-1 and BcAS1-2 (the homologous genes of AS1 gene in pak choi). In the transgenic Arabidopsis thaliana that overexpressed BcAS2 gene, it presented an abnormal phenotype with a curly shape. Taken together, our findings not only validate the function of BcAS2 in leaf development in Arabidopsis thaliana, but also contribute in unravelling the molecular regulatory mechanism of BcAS2, which fulfills a special role by forming complexes with BcAS1-1/2 in the establishment of the adaxial–abaxial polarity of the lateral organs in pak choi.