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81.
Resistance (R) gene-mediated immunity provides plants with rapid and strain-specific protection against pathogen infection. Our recent study using the genetically tractable Arabidopsis and turnip crinkle virus (TCV) pathosystem revealed a novel component, named CRT1 (compromised for recognition of the TCV CP), that is involved in general R gene-mediated signaling, including that mediated by HRT, an R gene against TCV. The Arabidopsis CRT1 gene family contains six additional members, of which two share high homology to CRT1 (75 and 81% a.a. identity); either CRT1 or its closest homolog restore the cell death phenotype suppressed by crt1. Analysis of single knock-out mutants for CRT1 and its closest homologs suggest that each may have unique and redundant functions. Here, we provide insight into the screening conditions that enabled identification of a mutant gene despite the presence of functionally redundant family members. We also discuss a potential mechanism that may regulate the interaction between CRT1 and R proteins.Key words: resistance gene, ATPase, suppressor screening, Arabidopsis, turnip crinkle virusPlant resistance (R) proteins activate defense signaling pathways following detection of a specific pathogen-encoded effector, or perception that a host factor has been altered by a pathogen effector. The vast majority of R proteins contain nucleotide binding site (NBS) and leucine-rich repeat (LRR) domains. These R proteins can be further divided into two subgroups, TIR-NBS-LRR and CC-NBS-LRR, depending on whether the N terminus consists of a Toll-interleukin 1 receptor (TIR) or a coiled-coiled (CC) domain, respectively.1 Subsequent to pathogen perception, the signal(s) generated by various R proteins likely converge into a limited set of pathways, with CC-NBS-LRR proteins usually utilizing NDR1 and TIR-NBS-LRR proteins generally requiring EDS1.2 However, the molecular mechanism(s) through which R proteins recognize a pathogen(s) and initiate a defense signal(s) remains unclear.To gain insights into this elusive signaling process, several groups have performed genetic screens to isolate mutants whose R gene-mediated resistance responses are suppressed following either pathogen infection or expression of a transgene-encoded bacterial effector protein. Several proteins, including HSP90, SGT1 and RAR1, were shown to be required for resistance triggered by a variety of R proteins, suggesting their universal function in R protein-mediated resistance.3–7 However, while some R protein-mediated signaling pathways required both RAR1 and SGT1, others needed only one or neither protein. Thus, the requirement for RAR1 and SGT1 appears to be specific to each pathway.8 Further studies revealed that SGT1, RAR1 and HSP90 regulate the stability/accumulation of various R proteins,8–11 raising the possibility that they serve as (co)chaperones for assembling an active R protein complex.The Arabidopsis R protein HRT was previously shown to recognize the coat protein (CP) of turnip crinkle virus (TCV) and trigger necrotic lesion formation in the inoculated leaf, as well as local and systemic defense responses.12 To identify components of the HRT-mediated signaling pathway, a line containing HRT and an inducible CP transgene was constructed and screened for suppressors of CP-induced cell death.13 One mutant, named crt1 (compromised for recognition of the TCV CP), was identified; it contains a mutation in a GHKL (Gyrase, Hsp90, histidine kinase, MutL) ATPase.13 Interestingly, HSP90 also belongs to this recently recognized ATPase superfamily, although sequence homology between HSP90 and CRT1 is limited to the ATPase domain.14 Either wt CRT1 or its closest homolog, CRT1-h1 (81% a.a. identity to CRT1; 13 suggesting that CRT1 and CRT1-h1 are functionally redundant.
Open in a separate windowGiven the presence of a functionally redundant homolog sharing 81% a.a. identity to CRT1, it is surprising that the crt1 mutant was identified. Because a previous study using the dexamethasone inducible system reported severe growth arrest and induction of defense-related genes when any transgene was highly expressed,15 we started with a transgenic line expressing CP at a level that was low (particularly in comparison to those attained during TCV infection), yet was sufficient to trigger cell death in non-mutant plants. The low level of CP expression in our transgenic line may have inadvertently provided screening conditions under which a rather modest compromise in R protein-mediated signaling could be detected, such as a mutation in a gene with functionally redundant family members. The crt1 and other crt mutants indeed showed cell death when CP was highly expressed via TCV infection. Thus, it is likely that crt1 would have escaped the suppressor screen if expression of the CP transgene had been higher. Another anti-viral R protein of Arabidopsis, RCY1, was utilized for a similar suppressor screen except that the effector protein was provided via viral infection.16 This screen identified mutations only in RCY1, consistent with our hypothesis that weak activation of the defense signaling pathway facilitated detection of a mutation in a gene that is part of a functionally redundant family.Since HRT-mediated resistance to TCV was impaired in crt1 and was further compromised by silencing closely related CRT1 family members,13 the functional copy number of CRT1 family members appears to be important for resistance. This result, combined with the semi-dominant nature of the crt1 mutation led us to test whether the mutant phenotype is due to haploid insufficiency. Analysis of single T-DNA knockout mutants for CRT1 or CRT1-h1 revealed that resistance to Pseudomonas syringae was not compromised, although it was suppressed in a double knockout mutant (unpublished). These results suggest that loss of a single copy of CRT1 is not sufficient to compromise TCV resistance signaling, thereby arguing that the crt1 phenotype is due to a dosage effect of disabled CRT1 family members. An alternative, although mutually not exclusive, possibility is that crt1 suppresses TCV resistance via a negative gain of function. Ectopic expression of some truncated CRT1 variants suppressed cell death triggered by the constitutively activated R protein ssi4.13 Thus, crt1 might suppress resistance signaling by competing with wild type CRT1 for an interacting partner, likely an R protein. Such a scenario could explain why CRT1 dosage affects TCV resistance.An intriguing possibility raised in a preview to our paper is that CRT1 may activate/prime a cytosolic R protein, which is then localized to the nucleus.17 Several lines of evidence suggest that nuclear localization of some R proteins is required for their function.18–20 Thus, CRT1 could be an important player that transits R proteins from one subcellular location to another, although it remains to be demonstrated whether HRT and the other R proteins shown to interact with CRT1 change subcellular location during resistance signaling. Another important question is what triggers CRT1 to activate/prime a client R protein. Western analysis has revealed that CRT1 is present as two distinct isoforms; the larger isoform presumably is created by an unknown post-translational modification.13 Interestingly, the larger CRT1 isoform interacts poorly with the NBS domain of HRT,13 suggesting that this putative modification is a crucial signal to release a client R protein. Thus, characterization of this post-translational modification may provide crucial insight into an R protein-mediate signaling pathway(s) that has been enigmatic for over a decade. 相似文献
Table 1
Amino-acid sequence identity between CRTI family members in ArabidopsisOpen in a separate window |
82.
Tao Tan Jun Wu Chenyang Si Shaoxing Dai Youyue Zhang Nianqin Sun E Zhang Honglian Shao Wei Si Pengpeng Yang Hong Wang Zhenzhen Chen Ran Zhu Yu Kang Reyna Hernandez-Benitez Llanos Martinez Martinez Estrella Nuñez Delicado W. Travis Berggren Juan Carlos Izpisua Belmonte 《Cell》2021,184(8):2020-2032.e14
83.
Carbon accumulation and distribution in Pinus massoniana and Cunninghamia lanceolata mixed forest ecosystem in Daqingshan, Guangxi, China 总被引:1,自引:0,他引:1 下载免费PDF全文
Carbon accumulation and distribution were studied at three sampling plots in a 13-year-old mixed planatation of Pinus massoniana and Cunninghamia lanceolata in Daqingshan, Guangxi, China. The results showed that carbon content varied with tissues and tree species, but the total carbon content of Pinus massoniana was higher than that of Cunninghamia lanceolata. The average tissue carbon contents of Pinus massoniana were: wood (58.6%) > root (56.3%) > branch (51.2%) > bark (49.8%) > leaf (46.8%), while those of Cunninghamia lanceolata were: bark (52.2%) > leaf (51.8%) > wood (50.2%) > root (47.5%) > branch (46.7%). The carbon contents of the soil (at a depth of 60cm) ranged from 1.45% to 1.84% with an average of 1.70%. Carbon contents were higher in the surface soil (0–20cm) than in the deep layer (below 20cm). The average carbon contents were the highest for trees (51.1%), followed by litter (48.3%), shrubs (44.1%), and herbs (33.0%). The biomass of the trees in the three plots ranged from 85.35 t hm-2 to 101.35 t hm-2 with an average of 93.83 t hm-2, in which 75.7%–82.6% was Pinus massoniana. The biomass of the understory was 2.10–3.95 t hm-2 with an average of 2.72 t hm-2, while the standing stock of ground litter was 5.49–7.91 t hm-2 with an average of 6.75 t hm-2. The carbon storage in the mixed plantation reached the maximum in the soil layer (69.02%), followed by vegetation (29.03%), and standing litter (1.82%). The carbon storage in the tree layer occupied 23.90% of the total ecosystem and 97.7% of the vegetation layer. Pinus massoniana accounted for 65.39% of the total carbon storage in the tree layer. Tissue carbon storage was directly related to the corresponding amount of biomass. Trunks had the highest carbon storage, accounting for 53.23% of the trees in Pinus massoniana and 55.57% in Cunninghamia lanceolata, respectively. Roots accounted for about 19.22% of the total tree carbon. The annual net productivity of the mixed plantation was 11.46 t hm-2a-1, and that of sequestered carbon was 5.96 t hm-2a-1, which was equivalent to fixing CO2 of 21.88 t hm-2a-1. The plantation was found to be an important sink of atmospheric CO2. 相似文献
84.
Byeong-Teck Kang Dong-Pyo Jang Su-Hyun Gu Jong-Hwan Lee Dong-In Jung Chae-Young Lim Ha-Jung Kim Young-Bo Kim Hyung-Joong Kim Eung-Je Woo Zang-Hee Cho Hee-Myung Park 《Comparative medicine》2009,59(5):459-464
The purpose of this study was to evaluate the diagnostic value of magnetic resonance imaging (MRI) and assess the correlation between the volume of the ischemic lesion and neurobehavioral status during the subacute stage of ischemic stroke. Ischemic stroke was induced in 6 healthy laboratory beagles through permanent occlusion of the middle cerebral artery (MCAO). T2-weighted and fluid-attenuated inversion recovery (FLAIR) imaging, diffusion-weighted imaging (DWI), measurement of the apparent diffusion coefficient (ADC) ratio, and neurobehavioral evaluation were performed 3 times serially by using a 1.5-T MR system: before and 3 and 10 d after MCAO. Ischemic lesions demonstrated T2 hyperintensity, FLAIR hyperintensity, and DWI hyperintensity. The ADC ratio was decreased initially but then was increased at 10 d after MCAO. Ischemic lesion volumes on T2-weighted and FLAIR imaging were not significantly different from those on DWI. The lesion volume and neurobehavioral score showed strong correlation. Our results suggest that conventional MRI may be a reliable diagnostic tool during the subacute stage of canine ischemic stroke.Abbreviations: ADC, apparent diffusion coefficient; DWI, diffusion-weighted imaging; FLAIR, fluid-attenuated inversion recovery; MCAO, middle cerebral artery occlusion; MRI, magnetic resonance imaging; PWI, perfusion-weighted imagingIn human medicine, stroke is a leading cause of adult mortality and neurologic disability worldwide.1 Strokes previously were thought to be uncommon in small animals, but the true prevalence is unknown.4 These events are now recognized more frequently in dogs because of increased use of magnetic resonance imaging (MRI).5,14,17Because the infusion of thrombolytic agents, such as urokinase or tissue plasminogen activator, within 3 to 6 h of the onset symptoms is effective in restoring blood flow and improving stroke outcome in humans,19 the detection of early ischemic changes is now thought to be necessary to improve patient outcome. Computed tomography and conventional MRI are not sufficiently sensitive to predict the presence and extent of ischemic damage during the acute stage after a stroke.12,20 Therefore several MRI sequences, such as fluid-attenuated inversion recovery (FLAIR), diffusion-weighted imaging (DWI), perfusion-weighted imaging (PWI), and MR angiography, have been developed for early diagnosis and subsequent follow-up of ischemic stroke.3 High-field magnetic strengths (at least 1.5 T) are necessary to perform these sequences.In contrast to the situation in humans, ischemic stroke in many dogs is diagnosed during the subacute stage—24 h to 6 wk after the vascular insult—due to the time lag between the onset of clinical signs to referral and to the lack of standard diagnostic protocols for ischemic stroke in dogs. In most reports of strokes in dogs, the median interval between the onset of neurologic dysfunction and performance of an MRI was more than 2 d.5,14,17 Whereas DWI has marked sensitivity to very early ischemic changes in the brain, T2-weighted and FLAIR images gradually become more hyperintense later (that is, during the first 24 h after the insult).3 Therefore, hyperintensity on T2-weighted and FLAIR images is believed to be representative of mature lesions.15 In light of these findings, we hypothesized that conventional MR sequences, such as T2-weighted and FLAIR imaging as well as DWI would be used for the diagnosis of the subacute stage of ischemic stroke in dogs.The purpose of this study was to evaluate the diagnostic value of MRI and assess the correlation between the volume of ischemic lesions and neurobehavioral status during the subacute stage of ischemic stroke in dogs. We therefore investigated the lesion volume of T2-weighted and FLAIR images compared with that on DWI images. Furthermore, we assessed the relationship between the apparent diffusion coefficient (ADC) of the ischemic lesions and the neurobehavioral status of the dogs. 相似文献
85.
86.
Michael Bauer Lifeng Kang Yiling Qiu Jinhui Wu Michelle Peng Howard H. Chen Gulden Camci-Unal Ahmad F. Bayomy David E. Sosnovik Ali Khademhosseini Ronglih Liao 《PloS one》2012,7(11)
Background
A major hurdle in the use of exogenous stems cells for therapeutic regeneration of injured myocardium remains the poor survival of implanted cells. To date, the delivery of stem cells into myocardium has largely focused on implantation of cell suspensions.Methodology and Principal Findings
We hypothesize that delivering progenitor cells in an aggregate form would serve to mimic the endogenous state with proper cell-cell contact, and may aid the survival of implanted cells. Microwell methodologies allow for the culture of homogenous 3D cell aggregates, thereby allowing cell-cell contact. In this study, we find that the culture of cardiac progenitor cells in a 3D cell aggregate augments cell survival and protects against cellular toxins and stressors, including hydrogen peroxide and anoxia/reoxygenation induced cell death. Moreover, using a murine model of cardiac ischemia-reperfusion injury, we find that delivery of cardiac progenitor cells in the form of 3D aggregates improved in vivo survival of implanted cells.Conclusion
Collectively, our data support the notion that growth in 3D cellular systems and maintenance of cell-cell contact improves exogenous cell survival following delivery into myocardium. These approaches may serve as a strategy to improve cardiovascular cell-based therapies. 相似文献87.
88.
Characterisation and transferability of apple SSRs to two European pear F<Subscript>1</Subscript> populations 总被引:1,自引:0,他引:1
Pierantoni L Cho KH Shin IS Chiodini R Tartarini S Dondini L Kang SJ Sansavini S 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2004,109(7):1519-1524
European pear (Pyrus communis L.) is among the important fruit species for which only few genetic studies have been carried out. Available evidence indicates that simple sequence repeats (SSR) are very useful as molecular markers because they are codominant, highly polymorphic, abundant and reproducible. The present paper reports more than 100 apple SSR markers in two populations of European pear; a total of 41 SSR markers were then positioned on a genetic linkage map of the cross Passe Crassane × Harrow Sweet and 31 in the map Abbè Fétel × Max Red Bartlett. Syntenic relationships between pear and apple maps have been considered for the chromosomes carrying two or more SSR markers. The alignment among the two maps supports the colinearity of the two genomes with respect both to identification and to orientation of the linkage groups. 相似文献
89.
Moon HJ Tiwari MK Singh R Kang YC Lee JK 《Applied and environmental microbiology》2012,78(9):3079-3086
Ribitol dehydrogenase from Zymomonas mobilis (ZmRDH) catalyzes the conversion of ribitol to d-ribulose and concomitantly reduces NAD(P)(+) to NAD(P)H. A systematic approach involving an initial sequence alignment-based residue screening, followed by a homology model-based screening and site-directed mutagenesis of the screened residues, was used to study the molecular determinants of the cofactor specificity of ZmRDH. A homologous conserved amino acid, Ser156, in the substrate-binding pocket of the wild-type ZmRDH was identified as an important residue affecting the cofactor specificity of ZmRDH. Further insights into the function of the Ser156 residue were obtained by substituting it with other hydrophobic nonpolar or polar amino acids. Substituting Ser156 with the negatively charged amino acids (Asp and Glu) altered the cofactor specificity of ZmRDH toward NAD(+) (S156D, [k(cat)/K(m)(,NAD)]/[k(cat)/K(m)(,NADP)] = 10.9, where K(m)(,NAD) is the K(m) for NAD(+) and K(m)(,NADP) is the K(m) for NADP(+)). In contrast, the mutants containing positively charged amino acids (His, Lys, or Arg) at position 156 showed a higher efficiency with NADP(+) as the cofactor (S156H, [k(cat)/K(m)(,NAD)]/[k(cat)/K(m)(,NADP)] = 0.11). These data, in addition to those of molecular dynamics and isothermal titration calorimetry studies, suggest that the cofactor specificity of ZmRDH can be modulated by manipulating the amino acid residue at position 156. 相似文献
90.
Inhibition of CLIC4 enhances autophagy and triggers mitochondrial and ER stress-induced apoptosis in human glioma U251 cells under starvation 总被引:1,自引:0,他引:1
CLIC4/mtCLIC, a chloride intracellular channel protein, localizes to mitochondria, endoplasmic reticulum (ER), nucleus and cytoplasm, and participates in the apoptotic response to stress. Apoptosis and autophagy, the main types of the programmed cell death, seem interconnected under certain stress conditions. However, the role of CLIC4 in autophagy regulation has yet to be determined. In this study, we demonstrate upregulation and nuclear translocation of the CLIC4 protein following starvation in U251 cells. CLIC4 siRNA transfection enhanced autophagy with increased LC3-II protein and puncta accumulation in U251 cells under starvation conditions. In that condition, the interaction of the 14-3-3 epsilon isoform with CLIC4 was abolished and resulted in Beclin 1 overactivation, which further activated autophagy. Moreover, inhibiting the expression of CLIC4 triggered both mitochondrial apoptosis involved in Bax/Bcl-2 and cytochrome c release under starvation and endoplasmic reticulum stress-induced apoptosis with CHOP and caspase-4 upregulation. These results demonstrate that CLIC4 nuclear translocation is an integral part of the cellular response to starvation. Inhibiting the expression of CLIC4 enhances autophagy and contributes to mitochondrial and ER stress-induced apoptosis under starvation. 相似文献