Arabidopsis STAY-GREEN2 Is a Negative Regulator of Chlorophyll Degradation during Leaf Senescence

Chlorophyll （Chl） degradation causes leaf yellowing during senescence or under stress conditions. For Chl breakdown, STAY-GREEN1 （SGR1） interacts with Chl catabolic enzymes （CCEs） and light-harvesting complex II （LHCII） at the thylakoid membrane, possibly to allow metabolic channeling of potentially phototoxic Chl breakdown intermediates. Among these Chl catabolic components, SGR1 acts as a key regulator of leaf yellowing. In addition to SGR1 （At4g22920）, the Arabidopsis thaliana genome contains an additional homolog, SGR2 （At4g11910）, whose biological function remains elusive. Under senescence-inducing conditions, SGR2 expression is highly up-regulated, similarly to SGR1 expression. Here we show that SGR2 function counteracts SGR1 activity in leaf Chl degradation; SGR2-overexpressing plants stayed green and the sgr2-1 knockout mutant exhibited early leaf yellowing under age-, dark-, and stress-induced senescence conditions. Like SGR1, SGR2 interacted with LHCII but, in contrast to SGR1, SGR2 interactions with CCEs were very limited. Furthermore, SGR1 and SGR2 formed homo- or heterodimers, strongly suggesting a role for SGR2 in negatively regulat- ing Chl degradation by possibly interfering with the proposed CCE-recruiting function of SGR1. Our data indicate an antagonistic evolution of the functions of SGR1 and SGR2 in Arabidopsis to balance Chl catabolism in chloroplasts with the dismantling and remobilizing of other cellular components in senescing leaf cells.     

Molecular Evolution of VEF-Domain-Containing PcG Genes in Plants

Arabidopsis VERNALIZATION2 （VRN2）, EMBRYONIC FLOWER2 （EMF2）, and FERTILIZATION-INDEPENDENT SEED2 （FIS2） are involved in vernalization-mediated flowering, vegetative development, and seed development, respectively. Together with Arabidopsis VEF-L36, they share a VEF domain that is conserved in plants and animals. To investigate the evolution of VEF-domain-containing genes （VEF genes）, we analyzed sequences related to VEF genes across land plants. To date, 24 full-length sequences from 11 angiosperm families and 54 partial sequences from another nine families were identified. The majority of the full-length sequences identified share greatest sequence similarity with and possess the same major domain structure as Arabidopsis EMF2. EMF2-1ike sequences are not only widespread among angiosperms, but are also found in genomic sequences of gymnosperms, lycophyte, and moss. No FIS2- or VEF-L36-1ike sequences were recovered from plants other than Arabidopsis, including from rice and poplar for which whole genomes have been sequenced. Phylogenetic analysis of the full-length sequences showed a high degree of amino acid sequence conservation in EMF2 homologs of closely related taxa. VRN2 homologs are recovered as a clade nested within the larger EMF2 clade. FIS2 and VEF-L36 are recovered in the VRN2 clade. VRN2 clade may have evolved from an EMF2 duplication event that occurred in the rosids prior to the divergence of the eurosid I and eurosid II lineages. We propose that dynamic changes in genome evolution contribute to the generation of the family of VEF-domain-containing genes, Phylogenetic analysis of the VEF domain alone showed that VEF sequences continue to evolve following EM F2NRN2 divergence in accordance with species relationship. Existence of EMF2-1ike sequences in animals and across land plants suggests that a prototype form of EMF2 was present prior to the divergence of the plant and animal lineages. A proposed sequence of events, based on domain organization and occurrence of intermediate seque     

A Histone H3 Lysine-27 Methyltransferase Complex Represses Lateral Root Formation in Arabidopsis thaliana

Root branching or lateral root formation is crucial to maximize a root system acquiring nutrients and water from soil. A lateral root （LR） arises from asymmetric cell division of founder cells （FCs） in a pre-branch site of the primary root, and FC establishment is essential for lateral root formation. FCs are known to be specified from xylem pole pericycle cells, but the molecular genetic mechanisms underlying FC establishment are unclear. Here, we report that, in Arabidopsis thaliana, a PRC2 （for Polycomb repressive complex 2） histone H3 lysine-27 （H3K27） methyltransferase complex, functions to inhibit FC establishment during LR initiation. We found that functional loss of the PRC2 subunits EMF2 （for EMBRYONIC FLOWER 2） or CLF （for CURLY LEAF） leads to a great increase in the number of LRs formed in the primary root. The CLF H3K27 methyltransferase binds to chromatin of the auxin efflux carrier gene PIN FORMED 1 （PIN1）, deposits the repres- sive mark H3K27me3 to repress its expression, and functions to down-regulate auxin maxima in root tissues and inhibit FC establishment. Our findings collectively suggest that EMF2-CLF PRC2 acts to down-regulate root auxin maxima and show that this complex represses LR formation in Arabidopsis.     

GNOM-LIKE 2, Encoding an Adenosine Diphosphate-Ribosylation Factor-Guanine Nucleotide Exchange Factor Protein Homologous to GNOM and GNL1, is Essential for Pollen Germination in Arabidopsis

In flowering plants, male gametes are delivered to female gametophytes by pollen tubes. Although it is important for sexual plant reproduction, little is known about the genetic mechanism that controls pollen germination and pollen tube growth. Here we report the identification and characterization of two novel mutants, gnom-like 2-1 （gnl2-1） and gn12-2 in Arabidopsis thaliana, in which the pollen grains failed to germinate in vitro and in vivo. GNL2 encodes a protein homologous to the adenosine diphosphate-ribosylation factor-guanine nucleotide exchange factors, GNOM and GNL1 that are involved in endosomal recycling and endoplasmic reticulum-Golgi vesicular trafficking. It was prolifically expressed in pollen grains and pollen tubes. The results of the present study suggest that GNL2 plays an important role in pollen germination.     

Protective effects of DIDS against ethanol-induced gastric mucosal injury in rats

The compound 4,4＇-diisothiocyanostilbene-2,2＇-disulfonic acid （DIDS） is an efficient anion exchanger inhibitor that can block the activities of anion exchanger 2 （AE2）, which plays an indispensable role in gastric acid secretion. DIDS also has potent anti-oxidative and antiapoptosis activities. This study aimed to investigate the effect of DIDS on ethanol-induced mucosal damage in rats and to evaluate the underlying mechanisms that mediate the action of the compound. The rats received 1 ml of absolute ethanol or saline orally. DIDS [50 mg/kg intravenous （i.v.）] was given 5 min before ethanol administration. Gastric lesions were evaluated macroscopically, microscopically, and electron microscopically at 60 min after ethanol challenge. Gastric myeloperoxidase （MPO） activity, malonyldialdehyde （MDA） level, prostaglandin E2 （PGE2） synthesis, and cyclooxygenase-2 （COX-2） expression were assessed. For the evaluation of the effect of DIDS on gastric acid secretion, histamine-stimulatory gastric acid secretion was examined with or without pretreatment of DIDS （50 mg/kg; i.v.）. Ethanol-induced gastric lesions were characterized by increasing gastric MDA level, MPO activity, and COX-2 expression, and decreasing PGE2 synthesis. It was found that DIDS significantly reduced the extent of gastric mucosal damage and reversed tissue MDA level and MPO activity. DIDS further enhanced the expression of COX-2 and reversed the decrease of PGE2. Our results suggested that DIDS is beneficial in rat model of gastric injury through mechanisms that involve inhibiting inflammatory cell infiltration and lipid peroxidation and up-regulating the COX-2/PGE2 pathway.     

Hydrogen peroxide induces the activation of the phospholipase C-γ1 survival pathway in PC12 cells： protective role in apoptosis

It has been reported that phospholipase C-γ1 （PLC-γ1） plays an important protective role in hydrogen peroxide （H2O2）-induced pheochromocytoma （PC） 12 cells death. However, most studies have used high doses of H2O2 and the downstream targets of PLC-γ1 activation remain to be identified. The present study was designed to examine the roles of PLC-γ1 signaling pathway in the apoptosis of PC12 cells induced by low dose of H2O2, as well as the downstream factors involved in this pathway. Low-dose treatment of H2O2 resulted in PLC-γ1 tyrosine phosphorylation in a time-dependent manner and H2O2 killed the PC12 cells by inducing necrosis. In contrast, pretreatment of PCI2 cells with U73122, a specific inhibitor of PLC, markedly increased the percentage of dead cells. The mode of cell death was converted to apoptosis as determined by Hoechst/PI nuclear staining and fluorescence microscopy. Western blot analysis demonstrated that the expression of Bcl-2 protein and the activation of pro-caspase-3 were not significantly affected by low dose of H2O2 alone. However, after pretreatment with U73122, Bcl-2 protein expression was dramatically decreased and the activation of pro-caspase-3 was significantly increased. We concluded that PLC-γ1 plays an important protective role in H2O2-induced PC12 cells death. Bcl-2 and caspase-3 probably participate in the signaling pathway as downstream factors.     

CD44 clustering is involved in monocyte differentiation

Differentiation of monocytes into macrophages is an import ant process under physiological and pathological conditions, but the underlying mechanism of monocyte differentiation is not completely clear. Some adhesion molecules have been reported to play an important role in cell differentiation. CD44 is an important adhesion molecule that mediates cell cell and cellmatrix interaction, and participates in a wide variety of cellular functions. As CD44 has been reported to show different activated states between monocytes and macrophages, we propose that CD44 may be involved in monocyte differentiation. In this study, we explored the role of CD44 in monocyte differentiation and further studied the mechanisms that were involved in. THP1 cells （human monocyfic leukemia cell line） were induced with phorbol 12myristate 13acetate （PMA） to establish the model of monocyte differentiation in vitro. It was found that CD44 expression and binding capacity to hyaluronic acid were increased significantly, and the distribution of CD44 was con verted into clusters during differentiation. The PMAinduced CD44 clustering and CD44 high expression were suppressed by blocking CD44, which resulted in the inhibition of CD14 expression. PMAinduced phosphorylation of ERK1/2 signal was also suppressed by blocking CD44. Our results suggested that CD44 was involved in monocyte differentiation. The mechanisms of monocyte differentiation following CD44 acti vation may include CD44 high expression and clustering which in turn lead to phosphorylation of ERK1/2.     

Potato Responds to Salt Stress by Increased Activity of Antioxidant Enzymes

To understand the response of potato to salt stress, antioxidant enzyme activities and ion content were analyzed for a sensitive and a tolerant cultivar. Nodal cuttings of the tolerant cultivar, Kennebec, and the sensitive cultivar, Concord, were exposed to media without or with 30, 60, 90 or 120 mmol/L NaCI for 4 weeks. On exposure to NaCI, the length and fresh and dry weight of both shoots and roots of Concord showed greater decrease than those of Kennebec. The decrease in shoot growth was more severe than that of the root for both cultivars. The K^＋ content of shoots and roots of both cultivars was reduced in a dose-dependent manner by exposure to NaCl; the Na^＋ content increased. Activities of ascorbate peroxidase, catalase and glutathione reductase were increased in NaCl-exposed shoots of Kennebec; the corresponding activities in NaCI-exposed shoots of Concord were decreased. Roots of both cultivars showed similar changes in the activities of these enzymes on exposure to NaCI. These studies established that enzyme activities in Concord shoots are inversely related to the NaCI concentration, whereas those in Kennebec do not show a dose dependency, which is also the case for the roots of both cultivars. Our findings suggest that an increase in activity of antioxidant enzymes, such as ascorbate peroxidase, catalase and glutathione reductase, can contribute to salt tolerance in Kennebec, a salt resistant cultivar of potato.     

Conserved Functions of Arabidopsis and Rice CC-Type Glutaredoxins in Flower Development and Pathogen Response

Glutaredoxins （GRXs） are ubiquitous oxidoreductases that play a crucial role in response to oxidative stress by reducing disulfides in various organisms. In planta, three different GRX classes have been identified according to their active site motifs. CPYC and CGFS classes are found in all organisms, whereas the CC-type class is specific for higher land plants. Recently, two Arabidopsis CC-type GRXs, ROXY1 and ROXY2, were shown to exert crucial functions in petal and anther initiation and differentiation. To analyze the function of CC-type GRXs in the distantly related monocots, we isolated and characterized OsROXY1 and OsROXY2-two rice homologs of ROXY1. Both genes are expressed in vegetative and reproductive stages. Although rice flower morphology is distinct from eudicots, OsROXY1/2 floral expression patterns are similar to their Arabidopsis counterparts ROXY1/2. Complementation experiments demonstrate that OsROXY1 and OsROXY2 can fully rescue the roxyl floral mutant phenotype. Overexpression of OsROXY1, OsROXY2, and ROXY1 in Arabidopsis causes similar vegetative and reproductive plant developmental defects. ROXY1 and its rice homologs thus exert a conserved function during eudicot and monocot flower development. Strikingly, overexpression of these CC-type GRXs also leads to an increased accumulation of hydrogen peroxide levels and hyper-susceptibility to infection from the necrotrophic pathogen Botrytis cinerea, revealing the importance of balanced redox processes in flower organ develop- ment and pathogen defence.     

Arabidopsis Profilin Isoforms,PRF1 and PRF2 Show Distinctive Binding Activities and Subcellular Distributions

Profilin is an actin-binding protein that shows complex effects on the dynamics of the actin cytoskeleton. There are five profilin isoforms in Arabidopsis thaliana L. However, it is still an open question whether these isoforms are functionally different. In the present study, two profilin isoforms from Arabidopsis, PRF1 and PRF2 were fused with green fuorescent protein （GFP） tag and expressed in Escherichia coil and A. thaliana in order to compare their biochemical properties in vitro and their cellular distributions in vivo. Biochemical analysis revealed that fusion proteins of GFP-PRF1 and GFP-PRF2 can bind to poly-L-proline and G-actin showing remarkable differences. GFP-PRF1 has much higher affinities for both poly-L-proline and G-actin compared with GFP-PRF2. Observations of living cells in stable transgenic A. thaliana lines revealed that 35S：：GFP-PRF1 formed a filamentous network, while 35S：：GFP-PRF2 formed polygonal meshes. Results from the treatment with latrunculin A and a subsequent recovery experiment indicated that filamentous alignment of GFP-PRF1 was likely associated with actin filaments. However, GFP-PRF2 localized to polygonal meshes resembling the endoplasmic reticulum. Our results provide evidence that Arabidopsis profllin isoforms PRF1 and PRF2 have different biochemical affinities for poly-L-proline and G-actin, and show distinctive Iocalizations in living cells. These data suggest that PRF1 and PRF2 are functionally different isoforms.