Aphid transmission and a polypeptide are specified by a defined region of the cauliflower mosaic virus genome

Infection of young turnip leaves with an aphid-transmissible isolate, Cabb B-JI, of cauliflower mosaic virus (CaMV) causes synthesis of an Mr 18 000 polypeptide (p18) which co-purifies with virus inclusion bodies. This polypeptide is not detectable in leaves infected with either of two aphid non-transmissible isolates. Campbell and CM4-184. Construction in vitro, of hybrid genomes between Cabb B-JI and Campbell isolates demonstrates that aphid transmissibility and presence of p18 is dependent on the small genome fragment from the BstEII site to the XhoI site. A deletion made in this fragment within open reading frame (ORF) II causes loss of aphid transmissibility and also terminates production of p18. We conclude that aphid transmissibility and the presence of p18 are related to the expression of ORF II of the CaMV genome.     

Fine genetic mapping of the <Emphasis Type="Italic">TuNI</Emphasis> locus causing systemic veinal necrosis by turnip mosaic virus infection in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

In the pathosystem of turnip mosaic virus (TuMV) and Arabidopsis thaliana, two distinct symptoms (mosaic symptom and veinal necrosis) were observed that were dependent upon the combination of the TuMV isolate and the Arabidopsis ecotype. The Col-0 ecotype developed mosaic symptoms after infection with the TuMV isolate Azu while the Ler ecotype developed veinal necrosis after infection with the same TuMV isolate. The Ler phenotype is controlled by a single dominant gene TuNI (TuMV necrosis inducer) which is located on chromosome 1. The TuNI gene was precisely mapped to the ~105 kb interval between the two markers of mXF41 and mRF28 by using several types of DNA polymorphism markers. Within this region, which included largely duplicated sequences, a total of 19 putative genes were predicted and 15 of these were classified into five gene families. The genes belonging to the gene families At1g58480 and At1g58602 may function in response to infection by pathogens. The gene family At1g58480 encodes lipase-like proteins, which might be involved in the induction of defence responses that are mediated by salicylic acid. The gene family At1g58602 encodes the CC-NBS-LRR (CNL) proteins, which are known to function as one of the plant resistance (R) proteins against pathogens. In the present study, the possibility that TuNI might function as an R gene was discussed.Y.K. and T.I. contributed equally to this study     

Phyllosphere bacterial communities of trichome-bearing and trichomeless Arabidopsis thaliana leaves

This study aimed to investigate whether the presence of trichomes as conspicuous physical attributes of the leaf surface affects the microbial community composition on Arabidopsis thaliana leaves. The A. thaliana ecotype Col-0 and its trichomeless gl1 mutant were grown in growth cabinets under climate-controlled conditions. The gl1 mutant showed a similar wax composition as the Col-0 wild type with slightly reduced amounts of C₂₉, C₃₁ and C₃₃ alkanes by GC/MS and GC/FID analyses. 120 bacterial isolates representing 39 bacterial genera were obtained from A. thaliana Col-0 leaf surfaces. Phylogenetic analysis of nearly full-length 16S rRNA sequences from 29 selected isolates confirmed their affiliation to the Proteobacteria (Alpha-, Beta-, Gamma-), Actinobacteria, Bacteroidetes and Firmicutes. The bacterial diversity on A. thaliana ecotype Col-0 and its gl1 mutant, devoid of trichomes, were further compared by denaturing gradient gel electrophoresis (DGGE). Banding patterns and sequencing of representative DGGE bands revealed the presence of phylotypes related to Sphingomonas (Alphaproteobacteria), Methylophilus (Betaproteobacteria) and Dyadobacter (Bacteroidetes) which are common phyllosphere inhabitants. Furthermore, wildtype and trichomeless mutant plants were exposed to outdoor conditions for 4–5 weeks. The DGGE gels showed only minor differences between the two plant lines, thus suggesting that trichomes per se do not affect bacterial diversity on Arabidopsis leaves under the experimental conditions tested.     

A new,pathogen-inducible gene of Arabidopsis is expressed in an ecotype-specific manner

In this study we describe a novel gene, which was isolated in an attempt to search for specific plant resistance genes of Arabidopsis against isolates of the phytopathogenic bacterium Xanthomonas campestris pv. campestris. The gene was cloned by differential screening of a genomic library of the Xcc 750-resistant ecotype Col-0, using cDNA populations derived from ecotype Col-0 and the Xcc 750-susceptible ecotype Oy-0. The isolated gene, CXc750, is differentially expressed in ecotypes of Arabidopsis thaliana. In addition, although highly expressed in uninfected plants, gene expression increases in response to pathogen attack. CXc750 potentially codes for a small, basic protein of about 10 kDa. The predicted protein product contains a potential signal leader peptide at the amino-terminal end but no ER retention sequence and no further transmembrane domain. This indicates that the gene product is transported to other compartments or out of the cell.The possible function of CXc750 as a member of the plant defense response system is discussed.     

<Emphasis Type="Italic">Arabidopsis TTR1</Emphasis> causes LRR-dependent lethal systemic necrosis,rather than systemic acquired resistance,to Tobacco ringspot virus

Most Arabidopsis ecotypes display tolerance to the Tobacco ringspot virus (TRSV), but a subset of Arabidopsis ecotypes, including Estland (Est), develop lethal systemic necrosis (LSN), which differs from the localized hypersensitive responses (HRs) or systemic acquired resistance (SAR) characteristic of incompatible reactions. Neither viral replication nor the systemic movement of TRSV was restricted in tolerant or sensitive Arabidopsis ecotypes; therefore, the LSN phenotype shown in the sensitive ecotypes might not be due to viral accumulation. In the present study, we identified the Est TTR1 gene (tolerance to Tobacco ringspot virus 1) encoding a TIR-NBS-LRR protein that controls the ecotype-dependent tolerant/sensitive phenotypes by a map-based cloning method. The tolerant Col-0 ecotype Arabidopsis transformed with the sensitive Est TTR1 allele developed an LSN phenotype upon TRSV infection, suggesting that the Est TTR1 allele is dominant over the tolerant ttr1 allele of Col-0. Multiple sequence alignments of 10 tolerant ecotypes from those of eight sensitive ecotypes showed that 10 LRR amino acid polymorphisms were consistently distributed across the TTR1/ttr1 alleles. Site-directed mutagenesis of these amino acids in the LRR region revealed that two sites, L956S and K1124Q, completely abolished the LSN phenotype. VIGS study revealed that TTR1 is dependent on SGT1, rather than EDS1. The LSN phenotype by TTR1 was shown to be transferred to Nicotiana benthamiana, demonstrating functional conservation of TTR1 across plant families, which are involved in SGT-dependent defense responses, rather than EDS1-dependent signaling pathways.     

Isolation and characterization of faithful and altered clones of the genomes of cauliflower mosaic virus isolates Cabb B-JI, CM4-184, and Bari I

Full-length genomes of cauliflower mosaic virus (CaMV) isolates Cabb B-JI, CM4-184, and Bari I have been cloned in the SalGI site of plasmid pAT 153. The cloned DNAs were characterized by restriction mapping and infectivity assays. All the sites present in the virion DNAs were found in the cloned DNAs. Comparison of restriction maps with those of DNA from two other isolates which have been recently completely sequenced revealed a close relationship among the different isolates. Some of the clones appear to be faithful copies of the viral genomes and these viral inserts are infectious when inoculated into turnip plants. Various clones with deletions in the CaMV DNA have been isolated and characterized. Some of them may correspond to deletions naturally occurring in a subpopulation of the virus whereas others occurred during cloning. None of the deleted fragments are infectious when inoculated into plants. Strikingly, all the deletions overlap one or two of the specific single-stranded breaks characteristic of caulimoviruses, suggesting that sequences surrounding the breaks are not dispensable.     

Environmental regulation of stomatal response in the <Emphasis Type="Italic">Arabidopsis</Emphasis> Cvi-0 ecotype

The Arabidopsis Cape Verde Islands (Cvi-0) ecotype is known to differ from other ecotypes with respect to environmental stress responses. We analyzed the stomatal behavior of Cvi-0 plants, in response to environmental signals. We investigated the responses of stomatal conductance and aperture to high [CO₂] in the Cvi-0 and Col-0 ecotypes. Cvi-0 showed constitutively higher stomatal conductance and more stomatal opening than Col-0. Cvi-0 stomata opened in response to light, but the response was slow. Under low humidity, stomatal opening was increased in Cvi-0 compared to Col-0. We then assessed whether low humidity affects endogenous ABA levels in Cvi-0. In response to low humidity, Cvi-0 had much higher ABA levels than Col-0. However, epidermal peels experiments showed that Cvi-0 stomata were insensitive to ABA. Measurements of organic and inorganic ions in Cvi-0 guard cell protoplasts indicated an over-accumulation of osmoregulatory anions (malate and Cl⁻). This irregular anion homeostasis in the guard cells may explain the constitutive stomatal opening phenotypes of the Cvi-0 ecotype, which lacks high [CO₂]-induced and low humidity-induced stomatal closure.     

Inappropriate annotation of a key defence marker in<Emphasis Type="Italic"> Arabidopsis</Emphasis>: will the real<Emphasis Type="Italic"> PR-1</Emphasis> please stand up?

PR-1 has been extensively used as a marker for salicylic acid (SA)-mediated defence and systemic and local acquired resistance. The Arabidopsis Genome Project annotates At2g19990 as PR-1. This gene is also identified as PR-1 in two full genome Arabidopsis microarrays, and TAIR cites approximately 60 articles to describe its patterns of expression. However, most of these citations are incorrect; the probes used were not At2g19990, but a homologous gene At2g14610, which is annotated as PR-1-like. Because of the potential for confusion, we analyzed the expression of both genes in Arabidopsis thaliana (L.) Heynh. At2g14610 (PR-1-like) showed the archetypal patterns of SA-responsive expression: mRNA levels increased following SA-treatment, inoculation with an avirulent (but not a virulent) strain of Pseudomonas syringae, and in wild-type (but not NahG) Arabidopsis infected with cauliflower mosaic virus (CaMV). In cpr5 mutants it was expressed constitutively. In contrast, expression of At2g19990 (annotated as PR-1) was detectable in neither SA-treated Col-0 nor in cpr5. Infection by virulent and avirulent isolates of P. syringae up-regulated expression, but to a similar level, and infection by CaMV induced a modest increase in expression in both the wild type and NahG. At2g19990, although pathogen responsive, does not show the SA-dependent patterns of expression expected from a member of the PR-1 regulon, and its annotation as PR-1 is inappropriate. The annotations should identify At2g14610 as the authentic PR-1.     

AvrAC(Xcc8004), a type III effector with a leucine-rich repeat domain from Xanthomonas campestris pathovar campestris confers avirulence in vascular tissues of Arabidopsis thaliana ecotype Col-0

Xanthomonas campestris pathovar campestris causes black rot, a vascular disease on cruciferous plants, including Arabidopsis thaliana. The gene XC1553 from X. campestris pv. campestris strain 8004 encodes a protein containing leucine-rich repeats (LRRs) and appears to be restricted to strains of X. campestris pv. campestris. LRRs are found in a number of type III-secreted effectors in plant and animal pathogens. These prompted us to investigate the role of the XC1553 gene in the interaction between X. campestris pv. campestris and A. thaliana. Translocation assays using the hypersensitive-reaction-inducing domain of X. campestris pv. campestris AvrBs1 as a reporter revealed that XC1553 is a type III effector. Infiltration of Arabidopsis leaf mesophyll with bacterial suspensions showed no differences between the wild-type strain and an XC1553 gene mutant; both strains induced disease symptoms on Kashmir and Col-0 ecotypes. However, a clear difference was observed when bacteria were introduced into the vascular system by piercing the central vein of leaves. In this case, the wild-type strain 8004 caused disease on the Kashmir ecotype, but not on ecotype Col-0; the XC1553 gene mutant became virulent on the Col-0 ecotype and still induced disease on the Kashmir ecotype. Altogether, these data show that the XC1553 gene, which was renamed avrAC_Xcc8004, functions as an avirulence gene whose product seems to be recognized in vascular tissues.     

A compatible interaction of Alternaria brassicicola with Arabidopsis thaliana ecotype DiG: evidence for a specific transcriptional signature

Background  

The interaction of Arabidopsis with Alternaria brassicicola provides a model for disease caused by necrotrophs, but a drawback has been the lack of a compatible pathosystem. Infection of most ecotypes, including the widely-studied line Col-0, with this pathogen generally leads to a lesion that does not expand beyond the inoculated area. This study examines an ecotype, Dijon G (DiG), which is considered sensitive to A. brassicicola.     

The Response of Lettuce Cultivars against Two Lettuce mosaic virus isolates,One Able to Systemically Infect the Resistant Cultivar Salinas 88

The response of seven lettuce cultivars to two geographically different Lettuce mosaic virus (LMV) isolates (LMV‐A, LMV‐T) was statistically evaluated based on infection rate, virus accumulation and symptom severity in different time trials. LMV‐A is characterized by the ability to systemically infect cv. Salinas 88 (mo1²‐carrying resistant cultivar), and inducing mild mosaic symptoms. Among lettuce cultivars, Varamin (a native cultivar) similar to cv. Salinas showed the most susceptibility to both LMV isolates, whereas another native cultivar, Varesh, was tolerant to the virus with minimal viral accumulation and symptom scores, significantly different from other cultivars at P < 0.05. LMV‐A systemically infects all susceptible lettuce cultivars more rapidly and at a higher rate than LMV‐T. This isolate accumulated in lettuce cultivars at a significantly higher level, determined by semiquantitative ELISA and induced more severe symptoms than LMV‐T isolate at 21 dpi. This is the first evidence for a LMV isolate with ability to systemically infect mo1²‐carrying resistant cultivar of lettuce from Iran. In this study, accumulation level of LMV showed statistically meaningful positive correlation with symptom severity on lettuce plants. Based on the results, three evaluated parameters differed considerably by lettuce cultivar and virus isolate.     

A Pleiotropic Phenotype is Associated with Altered Endogenous Hormone Balance in the Developmentally Stunted Mutant (<Emphasis Type="Italic">dsm1</Emphasis>)

A developmentally stunted mutant (dsm1) of Arabidopsis, isolated from an EMS mutant screen, had a pleiotropic phenotype, including repressed germination, retarded growth, delayed flowering, and impaired fertility. Additionally, dsm1 had a lifespan of approximately 160 days, which was more than twice the lifespan of the wild type (Col-0). Fine morphological and anatomical characters, such as the shoot apical meristem, root apical meristem, seed shape, and seed surface, were obviously altered in dsm1. We found that both abscisic acid and zeatin riboside levels were significantly greater in dsm1 than in Col-0 at all stages of development, while the levels of indole-3-acetic acid and gibberellins varied by age. The expressions of some abscisic acid-related genes were higher in dsm1 than in Col-0. These data indicate that DSM1 may play a general role in plant growth and development.     

Functional analysis of the C-terminal region of the vacuolar cadmium-transporting rice OsHMA3

Rice OsHMA3 is a vacuolar cadmium (Cd) transporter belonging to the P_1B-ATPase family and has a long (273aa) C-terminal region. We analyzed the function of the region related to Cd using the transgenic Arabidopsis Col-0 ecotype, which is sensitive to Cd. The OsHMA3 variant containing a truncated (58aa) C-terminal region did not confer Cd tolerance, whereas an OsHMA3 variant containing a longer truncated (105aa) C-terminal region conferred Cd tolerance to transgenic Arabidopsis. We conclude that the C-terminal region, particularly the region containing the first 105aa, has an important role in OsHMA3 activity.     

Plant growth-promoting rhizobacteria systemically protect Arabidopsis thaliana against Cucumber mosaic virus by a salicylic acid and NPR1-independent and jasmonic acid-dependent signaling pathway

Arabidopsis thaliana ecotype Columbia plants (Col-0) treated with plant growth-promoting rhizobacteria (PGPR) Serattia marcescens strain 90-166 and Bacillus pumilus strain SE34 had significantly reduced symptom severity by Cucumber mosaic virus (CMV). In some cases, CMV accumulation was also significantly reduced in systemically infected leaves. The signal transduction pathway(s) associated with induced resistance against CMV by strain 90-166 was determined using mutant strains and transgenic and mutant Arabidopsis lines. NahG plants treated with strains 90-166 and SE34 had reduced symptom severity indicating that the resistance did not require salicylic acid (SA). Strain 90-166 naturally produces SA under iron-limited conditions. Col-0 and NahG plants treated with the SA-deficient mutant, 90-166-1441, had significantly reduced CMV symptom severity with reduced virus accumulation in Col-0 plants. Another PGPR mutant, 90-166-2882, caused reduced disease severity in Col-0 and NahG plants. In a time course study, strain 90-166 reduced virus accumulation at 7 but not at 14 and 21 days post-inoculation (dpi) on the non-inoculated leaves of Col-0 plants. NahG and npr1-1 plants treated with strain 90-166 had reduced amounts of virus at 7 and 14 dpi but not at 21 dpi. In contrast, no decrease in CMV accumulation occurred in strain 90-166-treated fad3-2 fad7-2 fad8 plants. These data indicate that the protection of Arabidopsis against CMV by strain 90-166 follows a signaling pathway for virus protection that is independent of SA and NPR1, but dependent on jasmonic acid.     

Satellite RNA-mediated resistance to turnip crinkle virus in Arabidopsis involves a reduction in virus movement.

Satellite RNAs (sat-RNAs) are parasites of viruses that can mediate resistance to the helper virus. We previously showed that a sat-RNA (sat-RNA C) of turnip crinkle virus (TCV), which normally intensifies symptoms of TCV, is able to attenuate symptoms when TCV contains the coat protein (CP) of cardamine chlorotic fleck virus (TCV-CPCCFV). We have now determined that sat-RNA C also attenuates symptoms of TCV containing an alteration in the initiating AUG of the CP open reading frame (TCV-CPm). TCV-CPm, which is able to move systemically in both the TCV-susceptible ecotype Columbia (Col-0) and the TCV-resistant ecotype Dijon (Di-0), produced a reduced level of CP and no detectable virions in infected plants. Sat-RNA C reduced the accumulation of TCV-CPm by < 25% in protoplasts while reducing the level of TCV-CPm by 90 to 100% in uninoculated leaves of Col-0 and Di-0. Our results suggest that in the presence of a reduced level of a possibly altered CP, sat-RNA C reduces virus long-distance movement in a manner that is independent of the salicylic acid-dependent defense pathway.     

A tobamovirus infecting capsicum in Australia

A tobamovirus infection of Capsicum annuum is recorded for the first time in New South Wales, Australia. The causal virus is described and shown to differ from tobacco (TMV) and tomato (ToMV) mosaic viruses in its host reactions and serology. Seventeen capsicum cultivars were tested for reaction to the Australian isolate and ranked according to symptom severity. Field and glasshouse observations indicated that the virus causes a decrease in height and weight of plants, fruit malformation and leaf mosaic symptoms. It is proposed that the capsicum tobamovirus strains are sufficiently distinctive from TMV and ToMV to be grouped together and designated a separate virus: capsicum mosaic virus (CaMV).     

Selective allele loss and interference between cauliflower mosaic virus DNAs

Summary Some cauliflower mosaic virus (CaMV) alleles are selectively lost during growth of the virus in mixedly infected turnip plants. Viral DNA from plants co-inoculated with DNA of the cabbage S isolate and infectious cabbage S DNA with an extra EcoRI restriciion site lacked the extra site. The EcoRI allele was also lost in most plants co-inoculated with a non-infectious mutant of cabbage S DNA while little selective allele loss was observed with two other non-infectious mutant DNAs. Plants co-inoculated with DNAs of closely-related isolates (CM4-184 and W) contained both parental viral DNAs and some DNAs with characteristics of both parents. Interference, scored as a reduced frequency of infection or a delay in symptom appearance relative to plants inoculated with wild-type DNA, occurred when plants were inoculated with wild-type and mutant DNAs covalently attached to one another in partial dimer plasmid DNAs. Similarities in the conditions leading to selective allele loss and those leading to interference suggest that both may have been due to active gene conversion between CaMV DNA molecules.     

HMA3 is a key factor for differences in Cd- and Zn-related phenotype between <Emphasis Type="Italic">Arabidopsis</Emphasis> Ws and Col-0 ecotypes

Columbia-0 (Col-0) appears to be less tolerant to cadmium (Cd) than the Wassilewskija (Ws) ecotype that exhibits the full Heavy Metal ATPase3 (HMA3) coding sequence. However, the physiological and molecular mechanisms of HMA3 encoded by point mutation genes in Col-0 remain unknown. In this study, we investigate whether the different metal-related phenotype observed in Col-0 (with HMA3 mutation) when compared to that of Ws (functional HMA3) is a result only of the HMA3 mutation. This investigation was carried out with a further study using plant materials as follows: Ws and Col-0 ecotypes, two HMA3 (Ws) overexpressing lines in Col-0, hma3 knock-out line in Ws. The results indicate that the Col-0 and hma3 mutant in Ws were less tolerant to Cd and Zn because HMA3 has lost the function of sequestration of Cd and Zn into the root vacuoles, thereby readily translocating Cd and Zn to the aerial parts. In addition, the root-to-shoot metal translocation rates of the Ws- and HMA3-overexpressing lines were lower than those of the Col-0 and hma3 mutants. These results indicate that HMA3 is important for the Cd and Zn detoxification in Arabidopsis.