Down-regulation of <Emphasis Type="Italic">S</Emphasis>-adenosyl-<Emphasis Type="SmallCaps">l</Emphasis>-homocysteine hydrolase reveals a role of cytokinin in promoting transmethylation reactions

S-adenosyl-L: -homocysteine hydrolase (SAHH) is a key enzyme for maintenance of cellular transmethylation potential. Although a cytokinin-binding activity had been hypothesized for SAHH, the relation between cytokinin and transmethylation reactions has not been elucidated. Here we show that, of the two Arabidopsis thaliana SAHH genes, AtSAHH1 has a much higher expression level than AtSAHH2. A T-DNA insertion mutant of AtSAHH1 (sahh1-1) and the RNA interference (RNAi) plants (dsAtSAHH2) accumulated a higher level of cytokinins, exhibited phenotypic changes similar to those of cytokinin-overproducers, and their global DNA methylation status was reduced. On the other hand, cytokinins positively regulate the transmethylation pathway genes, including AtSAHH1, AtADK1 (for adenosine kinase), and this regulation involves the cytokinin activity. Furthermore, expression of three cytosine DNA methyltransferase genes examined was inducible by cytokinin treatment. Unlike adenine and adenosine which are SAHH inhibitors, the adenine-type cytokinins have no effect on SAHH activity at protein level. Changing of endogenous cytokinin levels by transgene expression resulted in alterations of DNA methylation status in the sahh1-1 background, suggesting that cytokinins promote DNA methylation, at least under transmethylation stringent conditions. These data demonstrate that the phytohormone cytokinin plays a role in promoting transmethylation reactions, including DNA methylation.     

Reactive oxygen species and root hairs in Arabidopsis root response to nitrogen, phosphorus and potassium deficiency

Plant root sensing and adaptation to changes in the nutrient status of soils is vital for long-term productivity and growth. Reactive oxygen species (ROS) have been shown to play a role in root response to potassium deprivation. To determine the role of ROS in plant response to nitrogen and phosphorus deficiency, studies were conducted using wild-type Arabidopsis and several root hair mutants. The expression of several nutrient-responsive genes was determined by Northern blot, and ROS were quantified and localized in roots. The monitored genes varied in intensity and timing of expression depending on which nutrient was deficient. In response to nutrient deprivation, ROS concentrations increased in specific regions of the Arabidopsis root. Changes in ROS localization in Arabidopsis and in a set of root hair mutants suggest that the root hair cells are important for response to nitrogen and potassium. In contrast, the response to phosphorus deprivation occurs in the cortex where an increase in ROS was measured. Based on these results, we put forward the hypothesis that root hair cells in Arabidopsis contain a sensing system for nitrogen and potassium deprivation.     

The GLABRA2 homeodomain protein directly regulates CESA5 and XTH17 gene expression in Arabidopsis roots

Arabidopsis root hair formation is determined by the patterning genes CAPRICE ( CPC ), GLABRA3 ( GL3 ), WEREWOLF ( WER ) and GLABRA2 ( GL2 ), but little is known about the later changes in cell wall material during root hair formation. A combined Fourier-transform infrared microspectroscopy–principal components analysis (FTIR-PCA) method was used to detect subtle differences in the cell wall material between wild-type and root hair mutants in Arabidopsis. Among several root hair mutants, only the gl2 mutation affected root cell wall polysaccharides. Five of the 10 genes encoding cellulose synthase ( CESA1 – 10 ) and 4 of 33 xyloglucan endotransglucosylase ( XTH1 – 33 ) genes in Arabidopsis are expressed in the root, but only CESA5 and XTH17 were affected by the gl2 mutation. The L1-box sequence located in the promoter region of these genes was recognized by the GL2 protein. These results indicate that GL2 directly regulates cell wall-related gene expression during root development.     

OsCSLD1, a cellulose synthase-like D1 gene, is required for root hair morphogenesis in rice

Root hairs are long tubular outgrowths that form on the surface of specialized epidermal cells. They are required for nutrient and water uptake and interact with the soil microflora. Here we show that the Oryza sativa cellulose synthase-like D1 (OsCSLD1) gene is required for root hair development, as rice (Oryza sativa) mutants that lack OsCSLD1 function develop abnormal root hairs. In these mutants, while hair development is initiated normally, the hairs elongate less than the wild-type hairs and they have kinks and swellings along their length. Because the csld1 mutants develop the same density and number of root hairs along their seminal root as the wild-type plants, we propose that OsCSLD1 function is required for hair elongation but not initiation. Both gene trap expression pattern and in situ hybridization analyses indicate that OsCSLD1 is expressed in only root hair cells. Furthermore, OsCSLD1 is the only member of the four rice CSLD genes that shows root-specific expression. Given that the Arabidopsis (Arabidopsis thaliana) gene KOJAK/AtCSLD3 is required for root hair elongation and is expressed in the root hair, it appears that OsCSLD1 may be the functional ortholog of KOJAK/AtCSLD3 and that these two genes represent the root hair-specific members of this family of proteins. Thus, at least part of the mechanism of root hair morphogenesis in Arabidopsis is conserved in rice.     

Further insights into the isoenzyme composition and activity of glutamate dehydrogenase in Arabidopsis thaliana

Following the discovery that in Arabidopsis, a third isoenzyme of NADH-dependent glutamate dehydrogenase (GDH) is expressed in the mitochondria of the root companion cells, we have re-examined the GDH isoenzyme composition. By analyzing the NADH-GDH isoenzyme composition of single, double and triple mutants deficient in the expression of the three genes encoding the enzyme, we have found that the α, β and γ polypeptides that comprise the enzyme can be assembled into a complex combination of heterohexamers in roots. Moreover, we observed that when one or two of the three root isoenzymes were missing from the mutants, the remaining isoenzymes compensated for this deficiency. The significance of such complexity is discussed in relation to the metabolic and signaling function of the NADH-GDH enzyme. Although it has been shown that a fourth gene encoding a NADPH-dependent enzyme is present in Arabidopsis, we were not able to detect corresponding enzyme activity, even in the triple mutant totally lacking NADH-GDH activity.     

Somatic Embryogenesis Receptor Kinases Control Root Development Mainly via Brassinosteroid-Independent Actions in Arabidopsis thaliana

Brassinosteroids(BRs),a group of plant steroidal hormones,play critical roles in many aspects of plant growth and development.Previous studies showed that BRI1-mediated BR signaling regulates cell division and differentiation during Arabidopsis root development via interplaying with auxin and other phytohormones.Arabidopsis somatic embryogenesis receptor-like kinases(SERKs),as co-receptors of BRI1,were found to play a fundamental role in an early activation step of BR signaling pathway.Here we report a novel function of SERKs in regulating Arabidopsis root development.Genetic analyses indicated that SERKs control root growth mainly via a BR-independent pathway.Although BR signaling pathway is completely disrupted in the serk1 bak1 bkk1 triple mutant,the root growth of the triple mutant is much severely damaged than the BR deficiency or signaling null mutants.More detailed analyses indicated that the triple mutant exhibited drastically reduced expression of a number of genes critical to polar auxin transport,cell cycle,endodermis development and root meristem differentiation,which were not observed in null BR biosynthesis mutant cpd and null BR signaling mutant bri1-701.     

Both vegetative and reproductive actin isovariants complement the stunted root hair phenotype of the Arabidopsis act2-1 mutation

The ACT2 gene, encoding one of eight actin isovariants in Arabidopsis, is the most strongly expressed actin gene in vegetative tissues. A search was conducted for physical defects in act2-1 mutant plants to account for their reduced fitness compared with wild type in population studies. The act2-1 insertion fully disrupted expression of ACT2 RNA and significantly lowered the level of total actin protein in vegetative organs. The root hairs of the act2-1 mutants were 10% to 70% the length of wild-type root hairs, and they bulged severely at the base. The length of the mutant root hairs and degree of bulging at the base were affected by adjusting the osmolarity and gelling agent of the growth medium. The act2-1 mutant phenotypes were fully rescued by an ACT2 genomic transgene. When the act2-1 mutation was combined with another vegetative actin mutation, act7-1, the resulting double mutant exhibited extensive synergistic phenotypes ranging from developmental lethality to severe dwarfism. Transgenic overexpression of the ACT7 vegetative isovariant and ectopic expression of the ACT1 reproductive actin isovariant also rescued the root hair elongation defects of the act2-1 mutant. These results suggest normal ACT2 gene regulation is essential to proper root hair elongation and that even minor differences may cause root defects. However, differences in the actin protein isovariant are not significant to root hair elongation, in sharp contrast to recent reports on the functional nonequivalency of plant actin isovariants. Impairment of root hair functions such as nutrient mining, water uptake, and physical anchoring are the likely cause of the reduced fitness seen for act2-1 mutants in multigenerational studies.     

Dissecting the role of CHITINASE-LIKE1 in nitrate-dependent changes in root architecture

The root phenotype of an Arabidopsis (Arabidopsis thaliana) mutant of CHITINASE-LIKE1 (CTL1), called arm (for anion-related root morphology), was previously shown to be conditional on growth on high nitrate, chloride, or sucrose. Mutants grown under restrictive conditions displayed inhibition of primary root growth, radial swelling, proliferation of lateral roots, and increased root hair density. We found here that the spatial pattern of CTL1 expression was mainly in the root and root tips during seedling development and that the protein localized to the cell wall. Fourier-transform infrared microspectroscopy of mutant root tissues indicated differences in spectra assigned to linkages in cellulose and pectin. Indeed, root cell wall polymer composition analysis revealed that the arm mutant contained less crystalline cellulose and reduced methylesterification of pectins. We also explored the implication of growth regulators on the phenotype of the mutant response to the nitrate supply. Exogenous abscisic acid application inhibited more drastically primary root growth in the arm mutant but failed to repress lateral branching compared with the wild type. Cytokinin levels were higher in the arm root, but there were no changes in mitotic activity, suggesting that cytokinin is not directly involved in the mutant phenotype. Ethylene production was higher in arm but inversely proportional to the nitrate concentration in the medium. Interestingly, eto2 and eto3 ethylene overproduction mutants mimicked some of the conditional root characteristics of the arm mutant on high nitrate. Our data suggest that ethylene may be involved in the arm mutant phenotype, albeit indirectly, rather than functioning as a primary signal.     

S-adenosylhomocysteine Hydrolase Participates in DNA Methylation Inheritance

DNA (cytosine-5) methyltransferase 1 (DNMT1) is essential for mammalian development and maintenance of DNA methylation following DNA replication in cells. The DNA methylation process generates S-adenosyl-l-homocysteine, a strong inhibitor of DNMT1. Here we report that S-adenosylhomocysteine hydrolase (SAHH/AHCY), the only mammalian enzyme capable of hydrolyzing S-adenosyl-l-homocysteine binds to DNMT1 during DNA replication. SAHH enhances DNMT1 activity in vitro, and its overexpression in mammalian cells led to hypermethylation of the genome, whereas its inhibition by adenosine periodate or siRNA-mediated knockdown resulted in hypomethylation of the genome. Hypermethylation was consistent in both gene bodies and repetitive DNA elements leading to aberrant gene regulation. Cells overexpressing SAHH specifically up-regulated metabolic pathway genes and down-regulated PPAR and MAPK signaling pathways genes. Therefore, we suggest that alteration of SAHH level affects global DNA methylation levels and gene expression.     

ATMYB123和ATKOR1基因参与拟南芥根发育的调控

从拟南芥T-DNA插入突变体库中筛选到2个根发育相关基因ATMYB123和ATKOR1表达缺失的突变体atmyb123和atkor1,通过杂交构建这两个基因表达缺失的双突变体atmyb123/atkor1,以明确这两个基因在根发育中的作用。结果显示:(1)ATMYB123表达缺失突变体atmyb123植株地上部分发育减缓,种皮颜色变黄,而ATKOR1表达缺失突变体atkor1植株在这两方面与其野生型没有明显差异;两基因缺失均显著影响了拟南芥根的发育,根生长受到了严重抑制。(2)双突变体atmyb123/atkor1在植株形态和种皮颜色方面表现出单突变体AT-MYB123的特点,而其根长却介于两单突变体的中间。(3)进一步研究发现,培养基pH改变、NaCl处理、外源GA施用均没有改变突变体根生长趋势,显示这3种因素与两基因缺失突变引起的根发育抑制无关。研究表明,AT-MYB123和ATKOR1基因参与拟南芥根的发育调控,转录因子ATMYB123可能作为主调控因子参与ATKOR1对拟南芥根发育的调控。     

Morphological,genetic and molecular characteristics of barley root hair mutants

Root hairs are tubular outgrowths of specialized epidermal cells called trichoblasts. They affect anchoring plants in soil, the uptake of water and nutrients and are the sites of the interaction between plants and microorganisms. Nineteen root hair mutants of barley representing different stages of root hair development were subjected to detailed morphological and genetic analyses. Each mutant was monogenic and recessive. An allelism test revealed that nine loci were responsible for the mutated root hair phenotypes in the collection and 1–4 mutated allelic forms were identified at each locus. Genetic relationships between the genes responsible for different stages of root hair formation were established. The linkage groups of four loci rhl1, rhp1, rhi1 and rhs1, which had previously been mapped on chromosomes 7H, 1H, 6H and 5H, respectively, were enriched with new markers that flank the genes at a distance of 0.16 cM to 4.6 cM. The chromosomal position of three new genes – two that are responsible for the development of short root hairs (rhs2 and rhs3) and the gene that controls an irregular root hair pattern (rhi2) – were mapped on chromosomes 6H, 2H and 1H, respectively. A comparative analysis of the agrobotanical parameters between some mutants and their respective parental lines showed that mutations in genes responsible for root hair development had no effect on the agrobotanical performance of plants that were grown under controlled conditions. The presented mutant collection is a valuable tool for further identification of genes controlling root hair development in barley.