Yunhai Li

BRITTLE CULM1, Which Encodes a COBRA-Like Protein, Affects the Mechanical Properties of Rice Plants

Plant mechanical strength is an important agronomic trait. To understand the molecular mechanism that controls the plant mechanical strength of crops, we characterized the classic rice mutant brittle culm1 (bc1) and isolated BC1 using a map-based cloning approach. BC1, which encodes a COBRA-like protein, is expressed mainly in developing sclerenchyma cells and in vascular bundles of rice. In these types of cells, mutations in BC1 cause not only a reduction in cell wall thickness and cellulose content but also an increase in lignin level, suggesting that BC1, a gene that controls the mechanical strength of monocots, plays an important role in the biosynthesis of the cell walls of mechanical tissues.

Control of final seed and organ size by the DA1 gene family in Arabidopsis thaliana

Although the size of an organism is a defining feature, little is known about the mechanisms that set the final size of organs and whole organisms. Here we describe Arabidopsis DA1, encoding a predicted ubiquitin receptor, which sets final seed and organ size by restricting the period of cell proliferation. The mutant protein encoded by the da1-1 allele has a negative activity toward DA1 and a DA1-related (DAR) protein, and overexpression of a da1-1 cDNA dramatically increases seed and organ size of wild-type plants, identifying this small gene family as important regulators of seed and organ size in plants.

SEPPA: a computational server for spatial epitope prediction of protein antigens

In recent years, a lot of efforts have been made in conformational epitope prediction as antigen proteins usually bind antibodies with an assembly of sequentially discontinuous and structurally compact surface residues. Currently, only a few methods for spatial epitope prediction are available with focus on single residue propensity scales or continual segments clustering. In the method of SEPPA, a concept of ‘unit patch of residue triangle’ was introduced to better describe the local spatial context in protein surface. Besides that, SEPPA incorporated clustering coefficient to describe the spatial compactness of surface residues. Validated by independent testing datasets, SEPPA gave an average AUC value over 0.742 and produced a successful pick-up rate of 96.64%. Comparing with peers, SEPPA shows significant improvement over other popular methods like CEP, DiscoTope and BEpro. In addition, the threshold scores for certain accuracy, sensitivity and specificity are provided online to give the confidence level of the spatial epitope identification. The web server can be accessed at http://lifecenter.sgst.cn/seppa/index.php. Batch query is supported.

The Ubiquitin Receptor DA1 Interacts with the E3 Ubiquitin Ligase DA2 to Regulate Seed and Organ Size in Arabidopsis

Although size control is a fundamental question in developmental biology, little is known about the mechanisms that determine seed and organ size in plants. Here, we show that two ubiquitin-related proteins, DA1 and DA2, interact genetically and physically with each other to regulate seed and organ size in Arabidopsis, suggesting that DA1 and DA2 are promising targets for crop improvement. Seed size in higher plants is determined by the coordinated growth of the embryo, endosperm, and maternal tissue. Several factors that act maternally to regulate seed size have been identified, such as AUXIN RESPONSE FACTOR2, APETALA2, KLUH, and DA1, but the genetic and molecular mechanisms of these factors in seed size control are almost totally unknown. We previously demonstrated that the ubiquitin receptor DA1 acts synergistically with the E3 ubiquitin ligase ENHANCER1 OF DA1 (EOD1)/BIG BROTHER to regulate the final size of seeds in Arabidopsis thaliana. Here, we describe another RING-type protein with E3 ubiquitin ligase activity, encoded by DA2, which regulates seed size by restricting cell proliferation in the maternal integuments of developing seeds. The da2-1 mutant forms large seeds, while overexpression of DA2 decreases seed size of wild-type plants. Overexpression of rice (Oryza sativa) GRAIN WIDTH AND WEIGHT2, a homolog of DA2, restricts seed growth in Arabidopsis. Genetic analyses show that DA2 functions synergistically with DA1 to regulate seed size, but does so independently of EOD1. Further results reveal that DA2 interacts physically with DA1 in vitro and in vivo. Therefore, our findings define the genetic and molecular mechanisms of three ubiquitin-related proteins DA1, DA2, and EOD1 in seed size control and indicate that they are promising targets for crop improvement.

Control of final organ size by Mediator complex subunit 25 in Arabidopsis thaliana

Control of organ size by cell proliferation and cell expansion is a fundamental developmental process, but the mechanisms that establish the final size of organs and whole organisms remain elusive in plants and animals. We have previously demonstrated that DA1, which encodes a predicted ubiquitin receptor, controls the final size of seeds and organs by restricting cell proliferation in Arabidopsis. Through a genetic screen for mutations that enhance the floral organ size of da1-1, we have identified an enhancer of da1-1 (eod8-1). The eod8-1 mutation was identified, using a map-based cloning approach, in Mediator complex subunit 25 (MED25; also known as PFT1), which is involved in the transcriptional regulation of gene expression. Loss-of-function mutants in MED25 form large organs, with larger and slightly increased numbers of cells as a result of an increased period of cell proliferation and cell expansion, whereas plants overexpressing MED25 have small organs owing to decreases in both cell number and cell size. Our genetic and physiological data suggest that MED25 acts to limit cell and organ growth independently of MED25-mediated phytochrome signaling and the jasmonate pathway. Genetic analyses show that MED25 functions redundantly with DA1 to control organ growth by restricting cell proliferation. Collectively, our findings show that MED25 plays a crucial role in setting final organ size, suggesting that it constitutes an important point of regulation in plant organ size control within the transcriptional machinery.

Maternal control of seed size by EOD3/CYP78A6 in Arabidopsis thaliana

Seed size in higher plants is coordinately determined by the growth of the embryo, endosperm and maternal tissue, but relatively little is known about the genetic and molecular mechanisms that set final seed size. We have previously demonstrated that Arabidopsis DA1 acts maternally to control seed size, with the da1-1 mutant producing larger seeds than the wild type. Through an activation tagging screen for modifiers of da1-1, we have identified an enhancer of da1-1 (eod3-1D) in seed size. EOD3 encodes the Arabidopsis cytochrome P450/CYP78A6 and is expressed in most plant organs. Overexpression of EOD3 dramatically increases the seed size of wild-type plants, whereas eod3-ko loss-of-function mutants form small seeds. The disruption of CYP78A9, the most closely related family member, synergistically enhances the seed size phenotype of eod3-ko mutants, indicating that EOD3 functions redundantly with CYP78A9 to affect seed growth. Reciprocal cross experiments show that EOD3 acts maternally to promote seed growth. eod3-ko cyp78a9-ko double mutants have smaller cells in the maternal integuments of developing seeds, whereas eod3-1D forms more and larger cells in the integuments. Genetic analyses suggest that EOD3 functions independently of maternal factors DA1 and TTG2 to influence seed growth. Collectively, our findings identify EOD3 as a factor of seed size control, and give insight into how plants control their seed size.

Regulation of OsGRF4 by OsmiR396 controls grain size and yield in rice

An increase in grain yield is crucial for modern agriculture1. Grain size is one of the key components of grain yield in rice and is regulated by quantitative trait loci (QTLs)2,3. Exploring new QTLs for grain size will help breeders develop elite rice varieties with higher yields3,4. Here, we report a new semi-dominant QTL for grain size and weight (GS2) in rice, which encodes the transcription factor OsGRF4 (GROWTH-REGULATING FACTOR 4) and is regulated by OsmiR396. We demonstrate that a 2 bp substitution mutation in GS2 perturbs OsmiR396-directed regulation of GS2, resulting in large and heavy grains and increased grain yield. Further results reveal that GS2 interacts with the transcription coactivitors OsGIF1/2/3, and overexpression of OsGIF1 increases grain size and weight. Thus, our findings define the regulatory mechanism of GS2, OsGIFs and OsmiR396 in grain size and weight control, suggesting this pathway could be used to increase yields in crops.

The plant‐specific G protein γ subunit AGG3 influences organ size and shape in Arabidopsis thaliana

• Control of organ size and shape by cell proliferation and cell expansion is a fundamental developmental process, but the mechanisms that set the size and shape of determinate organs are largely unknown in plants. • Molecular, genetic, cytological and biochemical approaches were used to characterize the roles of the Arabidopsis thaliana G protein γ subunit (AGG3) gene in organ growth. • Here, we describe A. thaliana AGG3, which promotes petal growth by increasing the period of cell proliferation. Both the N-terminal region and the C-terminal domains of AGG3 are necessary for the function of AGG3. By contrast, analysis of a series of AGG3 derivatives with deletions in specific domains showed that the deletion of any of these domains cannot completely abolish the function of AGG3. The GFP-AGG3 fusion protein is localized to the plasma membrane. The predicted transmembrane domain plays an important role in the plasma membrane localization of AGG3. Genetic analyses revealed that AGG3 action requires a functional G protein α subunit (GPA1) and G protein β subunit (AGB1). • Our findings demonstrate that AGG3, GPA1 and AGB1 act in the same genetic pathway to influence organ size and shape in A. thaliana.

Arabidopsis NAP and PIR Regulate Actin-Based Cell Morphogenesis and Multiple Developmental Processes

The actin cytoskeleton mediates cellular processes through the dynamic regulation of the time, location, and extent of actin polymerization. Actin polymerization is controlled by several types of evolutionarily conserved proteins, including those comprising the ARP2/3 complex. In animal cells ARP2/3 activity is regulated by WAVE complexes that contain WAVE/SCAR proteins, PIR121, Nap125, and other proteins. The activity of the WAVE complex is regulated by Rho-GTPase-mediated signaling that leads to ARP2/3 activation by WAVE/SCAR proteins. We describe in this report Arabidopsis (Arabidopsis thaliana) genes encoding Nap and PIR proteins. Light-grown Atnap-1 and Atpir-1 mutant plants displayed altered leaf, inflorescence, silique, and seed set phenotypes. Dark-grown Atnap-1 and Atpir-1 seedlings also exhibited longer roots, enhanced skotomorphogenesis and Glc responses, and shorter thicker hypocotyls than those of wild type, showing that AtNAP and AtPIR participate in a variety of growth and developmental processes. Mutations in AtNAP and AtPIR caused cell morphology defects in cotyledon pavement cells and trichomes seen in mutants in ARP2/3 subunits and in plants expressing constitutively active Rop2 GTPase. The patterns and levels of actin polymerization observed in Atnap-1 and Atpir-1 mutant trichome cells and epidermal pavement cell morphology is consistent with Arabidopsis NAP and PIR proteins forming a WAVE complex that activates ARP2/3 activity. The multiple growth and developmental phenotypes of Atnap and Atpir mutants reveals these proteins are also required for a wider variety of cellular functions in addition to regulating trichome cell growth.

Signaling from an Altered Cell Wall to the Nucleus Mediates Sugar-Responsive Growth and Development in Arabidopsis thaliana

Sugars such as glucose function as signal molecules that regulate gene expression, growth, and development in plants, animals, and yeast. To understand the molecular mechanisms of sugar responses, we isolated and characterized an Arabidopsis thaliana mutant, high sugar response8 (hsr8), which enhances sugar-responsive growth and gene expression. Light-grown hsr8 plants exhibited increased starch and anthocyanin and reduced chlorophyll content in response to glucose treatment. Dark-grown hsr8 seedlings showed glucose-hypersensitive hypocotyl elongation and development. The HSR8 gene, isolated using map-based cloning, was allelic to the MURUS4 (MUR4) gene involved in arabinose synthesis. Dark-grown mur1 and mur3 seedlings also exhibited similar sugar responses to hsr8/mur4. The sugar-hypersensitive phenotypes of hsr8/mur4, mur1, and mur3 were rescued by boric acid, suggesting that alterations in the cell wall cause hypersensitive sugar-responsive phenotypes. Genetic analysis showed that sugar-hypersensitive responses in hsr8 mutants were suppressed by pleiotropic regulatory locus1 (prl1), indicating that nucleus-localized PRL1 is required for enhanced sugar responses in hsr8 mutant plants. Microarray analysis revealed that the expression of many cell wall–related and sugar-responsive genes was altered in mur4-1, and the expression of a significant proportion of these genes was restored to wild-type levels in the mur4-1 prl1 double mutant. These findings reveal a pathway that signals changes in the cell wall through PRL1 to altered gene expression and sugar-responsive metabolic, growth, and developmental changes.