safae Hamdoun | Morgan State Unuversity (original) (raw)

Papers by safae Hamdoun

PLOS ONE, Dec 11, 2013

Plant-pathogen interactions involve sophisticated action and counteraction strategies from both p... more Plant-pathogen interactions involve sophisticated action and counteraction strategies from both parties. Plants can recognize pathogen derived molecules, such as conserved pathogen associated molecular patterns (PAMPs) and effector proteins, and subsequently activate PAMP-triggered immunity (PTI) and effector-triggered immunity (ETI), respectively. However, pathogens can evade such recognitions and suppress host immunity with effectors, causing effector-triggered susceptibility (ETS). The differences among PTI, ETS, and ETI have not been completely understood. Toward a better understanding of PTI, ETS, and ETI, we systematically examined various defenserelated phenotypes of Arabidopsis infected with different Pseudomonas syringae pv. maculicola ES4326 strains, using the virulence strain DG3 to induce ETS, the avirulence strain DG34 that expresses avrRpm1 (recognized by the resistance protein RPM1) to induce ETI, and HrcCthat lacks the type three secretion system to activate PTI. We found that plants infected with different strains displayed dynamic differences in the accumulation of the defense signaling molecule salicylic acid, expression of the defense marker gene PR1, cell death formation, and accumulation/localization of the reactive oxygen species, H 2 O 2. The differences between PTI, ETS, and ETI are dependent on the doses of the strains used. These data support the quantitative nature of PTI, ETS, and ETI and they also reveal qualitative differences between PTI, ETS, and ETI. Interestingly, we observed the induction of large cells in the infected leaves, most obviously with HrcCat later infection stages. The enlarged cells have increased DNA content, suggesting a possible activation of endoreplication. Consistent with strong induction of abnormal cell growth by HrcC-, we found that the PTI elicitor flg22 also activates abnormal cell growth, depending on a functional flg22-receptor FLS2. Thus, our study has revealed a comprehensive picture of dynamic changes of defense phenotypes and cell fate determination during Arabidopsis-P. syringae interactions, contributing to a better understanding of plant defense mechanisms.

<p>The fourth to sixth leaves of 30-day-old plants were infiltrated with <i>P. syring... more <p>The fourth to sixth leaves of 30-day-old plants were infiltrated with <i>P. syringae</i> strains at OD₆₀₀ 0.01, using 10 mM MgSO₄ treatment as a control. The infiltrated leaves were collected at the indicated times and were further cut into 1x2 mm sections. The sections were incubated with freshly prepared 5 mM CeCl₃ in 50 mM MOPS at pH 7.2 or MOPS without CeCl₃ for 1 h. The samples were then fixed and processed for TEM imaging. At least six different leaf samples for each treatment were fixed, and six sections were observed in each sample. (A) Cell morphology at 0 h. (B-C) H₂O₂ localization at 6 h (B) and 24 h (C) after DG34 inoculation. (D-F) H₂O₂ localization at 6 h (D), 18 h (E) and 24 h (F) after DG3 inoculation. (G-H) H₂O₂ localization at 18 h (G) and 48 h (H) after HrcC^- inoculation. Arrows indicate electron-dense cerium deposits. Asterisks indicate bacteria. Ch, chloroplast; CW, cell wall; M, mitochondrion; P, peroxisome.</p

<p>Leaves infiltrated with HrcC^- (OD₆₀₀ 0.1),... more <p>Leaves infiltrated with HrcC^- (OD₆₀₀ 0.1), flg22 (1 μM or 10 μM), or BTH (10 μM or 300 μM) were quantified for the formation of abnormal growths 4 days post treatment, using a dissecting microscope. (A) Flg22-induced cell enlargement is FLS2-dependent. (B) HrcC^- partially requires FLS2 to induce large cells. (C) BTH treatment does not induce abnormal growth. (D) HrcC^--induced abnormal growth requires SID2 and NPR1. For quantification of abnormal growths, at least 25 leaves from 12 plants were used for each BTH treatment or bacterial infection and 18 leaves from 7 plants were used for each flg22 treatment. Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Asterisks in (A), (B), and (D) indicate significant difference between treatments of the same genotype (P<0.05). </p

<p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>P. syr... more <p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>P. syringae</i> strains and observed for leaf morphology. (A) Pictures of leaf cross sections. Infected leaves were collected at 4 dpi and fixed for embedding with LR White resin. One-micron sections were cut and stained with 1% toluidine blue O for photographing. Leaves infected with DG3 (0.01) were mostly dead at 4 dpi and thus no data is available. Arrows indicate enlarged cells. The size bar represents 200 μm and applies to all images. Each growth (or a protrusion) has multiple enlarged cells. (B) Quantitation of abnormal growths. The number of abnormal growths, appearing to be transparent protrusions on the treated leaves, was counted at 4 dpi with a dissecting microscope. At least 25 leaves from each treatment were used in the counting. Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Different letters indicate significant difference among the samples (P<0.05). </p

<p><i>P. syringae</i>- infected leaves were embedded in paraplast and cut into ... more <p><i>P. syringae</i>- infected leaves were embedded in paraplast and cut into 15 μm sections for DAPI staining. Images of leaf cross-sections stained with DAPI to show nuclei were captured with a constant exposure time, using a Nikon DS cooled camera attached to a compound microscope. The images are from the following treatments: (A) 10 mM MgSO4, (B) DG34 0.01, (C) DG34 0.001, (D) HrcC^- 0.1, and (E) HrcC^- 0.01. (F) Typical nuclei of guard cells from a mock-treated leaf. Guard cells from leaves infected with DG34 0.01, DG34 0.001, HrcC^- 0.1, or HrcC^- 0.01 show visually similar nuclei (data not shown). (G) A typical nucleus of a normal mesophyll cell from a mock-treated leaf. Normal sized mesophyll cells from leaves infected with DG34 0.01, DG34 0.001, HrcC^- 0.1, or HrcC^- 0.01 show visually similar nuclei (data not shown). (H) A typical nucleus of an enlarged mesophyll cell induced by HrcC^- 0.01. Enlarged cells from leaves infected with DG34 0.01, DG34 0.001, or HrcC^- 0.1 also show large nuclei (data not shown). Arrows indicate the large nuclei in (A) to (E). The size bar in (A) represents 100 μm and applies to panels (A) to (E) while the size bar in (F) represents 5 μm and applies to panels (F) to (H). (J) Relative nuclear DNA content. The average fluorescence of nuclei of guard cells from mock-treated leaves (1) was set as 2C and was used to quantify relative nuclear DNA content of normal mesophyll cells from mock-treated leaves (2) and the enlarged mesophyll cells induced by HrcC^- (0.01) (3). At least 60 nuclei were used for each data point. Nuclear DNA contents of guard cells and normal mesophyll cells from HrcC^- (0.01)-infected leaves are similar to those of their corresponding cells from mock-treated leaves (data not shown). Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Different letters indicate significant difference among the samples (P<0.05).</p

<p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>Pseudo... more <p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>Pseudomonas syringae. </i><i>pv. </i><i>maculicola</i> ES4326 strains, DG3 at OD₆₀₀ 0.01 or 0.001, DG34 at OD₆₀₀ 0.01 or 0.001, or HrcC^- at OD₆₀₀ 0.1 or 0.01. The infected leaves were collected at the indicated times for SA and RNA analysis. (A) SA quantitation by HPLC analysis. Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Different letters indicate significant difference among the samples at the same time point (P<0.05). (B) Northern blotting for PR1 expression. Image of rRNA was used for a loading control. </p

Plants are constantly challenged by pathogens. Studies from several plant-pathosystems indicate t... more Plants are constantly challenged by pathogens. Studies from several plant-pathosystems indicate that manipulation of host cell fate might be a common strategy that pathogens use to succeed in their attacks. Genetic perturbation of the key defense signaling mediated by salicylic acid (SA) is also known to lead to altered cell fate in some Arabidopsis mutants. One of such mutants is accelerated cell death 6 (acd6-1), which demonstrates aberrant cell death coupled with cell growth, constitutive defense, and extreme dwarf phenotypes. However, the mechanism by which cell fate is determined during pathogen infection and in some defense mutants has not been well understood. I hypothesize that precise regulation of cell fate is intimately linked to plant disease resistance. I tested this hypothesis during my PhD research. I have found that: 1) There are dynamic, quantitative, and qualitative differences in the accumulation of the defense signaling molecule salicylic acid, expression of the ...

Molecular Plant Pathology, 2017

Erwinia amylovora is the causal agent of the fire blight disease in some plants of the Rosaceae f... more Erwinia amylovora is the causal agent of the fire blight disease in some plants of the Rosaceae family. The non-host plant Arabidopsis serves as a powerful system for the dissection of mechanisms of resistance to E. amylovora. Although not yet known to mount gene-for-gene resistance to E. amylovora, we found that Arabidopsis activated strong defence signalling mediated by salicylic acid (SA), with kinetics and amplitude similar to that induced by the recognition of the bacterial effector avrRpm1 by the resistance protein RPM1. Genetic analysis further revealed that SA signalling, but not signalling mediated by ethylene (ET) and jasmonic acid (JA), is required for E. amylovora resistance. Erwinia amylovora induces massive callose deposition on infected leaves, which is independent of SA, ET and JA signalling and is necessary for E. amylovora resistance in Arabidopsis. We also observed tumour-like growths on E. amylovora-infected Arabidopsis leaves, which contain enlarged mesophyll cells with increased DNA content and are probably a result of endoreplication. The formation of such growths is largely independent of SA signalling and some E. amylovora effectors. Together, our data reveal signalling requirements for E. amylovora-induced disease resistance, callose deposition and cell fate change in the non-host plant Arabidopsis. Knowledge from this study could facilitate a better understanding of the mechanisms of host defence against E. amylovora and eventually improve host resistance to the pathogen.

Plant physiology, Jan 11, 2015

Precise cell cycle control is critical for plant development and responses to pathogen invasion. ... more Precise cell cycle control is critical for plant development and responses to pathogen invasion. Two homologous cyclin-dependent kinase inhibitor (CKI) genes, SIAMESE (SIM) and SIM-RELATED 1 (SMR1), were recently shown to regulate Arabidopsis defense. However whether these two genes play differential roles in cell cycle and defense control is unknown. In this report, we showed that while acting synergistically to promote endoreplication, SIM and SMR1 play different roles in affecting the ploidy of trichome and leaf cells. In addition, we found that the smr1-1 mutant but not sim-1 was more susceptible to a virulent Pseudomonas syringae strain and this susceptibility could be rescued by activating salicylic acid (SA)-mediated defense. Consistent with these results, smr1-1 partially suppressed dwarfism, high SA levels, and cell death phenotypes in acd6-1, a mutant used to gauge the change of defense levels. Thus SMR1 functions partly through SA in defense control. The differential role...

Plant Physiology, 2011

The salicylic acid (SA) regulatory gene HOPW1-1-INTERACTING3 (WIN3) was previously shown to confe... more The salicylic acid (SA) regulatory gene HOPW1-1-INTERACTING3 (WIN3) was previously shown to confer resistance to the biotrophic pathogen Pseudomonas syringae. Here, we report that WIN3 controls broad-spectrum disease resistance to the necrotrophic pathogen Botrytis cinerea and contributes to basal defense induced by flg22, a 22-amino acid peptide derived from the conserved region of bacterial flagellin proteins. Genetic analysis indicates that WIN3 acts additively with several known SA regulators, including PHYTOALEXIN DEFICIENT4, NONEXPRESSOR OF PR GENES1 (NPR1), and SA INDUCTION-DEFICIENT2, in regulating SA accumulation, cell death, and/or disease resistance in the Arabidopsis (Arabidopsis thaliana) mutant acd6-1. Interestingly, expression of WIN3 is also dependent on these SA regulators and can be activated by cell death, suggesting that WIN3-mediated signaling is interconnected with those derived from other SA regulators and cell death. Surprisingly, we found that WIN3 and NPR1 ...

PLOS ONE, Dec 11, 2013

Plant-pathogen interactions involve sophisticated action and counteraction strategies from both p... more Plant-pathogen interactions involve sophisticated action and counteraction strategies from both parties. Plants can recognize pathogen derived molecules, such as conserved pathogen associated molecular patterns (PAMPs) and effector proteins, and subsequently activate PAMP-triggered immunity (PTI) and effector-triggered immunity (ETI), respectively. However, pathogens can evade such recognitions and suppress host immunity with effectors, causing effector-triggered susceptibility (ETS). The differences among PTI, ETS, and ETI have not been completely understood. Toward a better understanding of PTI, ETS, and ETI, we systematically examined various defenserelated phenotypes of Arabidopsis infected with different Pseudomonas syringae pv. maculicola ES4326 strains, using the virulence strain DG3 to induce ETS, the avirulence strain DG34 that expresses avrRpm1 (recognized by the resistance protein RPM1) to induce ETI, and HrcCthat lacks the type three secretion system to activate PTI. We found that plants infected with different strains displayed dynamic differences in the accumulation of the defense signaling molecule salicylic acid, expression of the defense marker gene PR1, cell death formation, and accumulation/localization of the reactive oxygen species, H 2 O 2. The differences between PTI, ETS, and ETI are dependent on the doses of the strains used. These data support the quantitative nature of PTI, ETS, and ETI and they also reveal qualitative differences between PTI, ETS, and ETI. Interestingly, we observed the induction of large cells in the infected leaves, most obviously with HrcCat later infection stages. The enlarged cells have increased DNA content, suggesting a possible activation of endoreplication. Consistent with strong induction of abnormal cell growth by HrcC-, we found that the PTI elicitor flg22 also activates abnormal cell growth, depending on a functional flg22-receptor FLS2. Thus, our study has revealed a comprehensive picture of dynamic changes of defense phenotypes and cell fate determination during Arabidopsis-P. syringae interactions, contributing to a better understanding of plant defense mechanisms.

<p>The fourth to sixth leaves of 30-day-old plants were infiltrated with <i>P. syring... more <p>The fourth to sixth leaves of 30-day-old plants were infiltrated with <i>P. syringae</i> strains at OD₆₀₀ 0.01, using 10 mM MgSO₄ treatment as a control. The infiltrated leaves were collected at the indicated times and were further cut into 1x2 mm sections. The sections were incubated with freshly prepared 5 mM CeCl₃ in 50 mM MOPS at pH 7.2 or MOPS without CeCl₃ for 1 h. The samples were then fixed and processed for TEM imaging. At least six different leaf samples for each treatment were fixed, and six sections were observed in each sample. (A) Cell morphology at 0 h. (B-C) H₂O₂ localization at 6 h (B) and 24 h (C) after DG34 inoculation. (D-F) H₂O₂ localization at 6 h (D), 18 h (E) and 24 h (F) after DG3 inoculation. (G-H) H₂O₂ localization at 18 h (G) and 48 h (H) after HrcC^- inoculation. Arrows indicate electron-dense cerium deposits. Asterisks indicate bacteria. Ch, chloroplast; CW, cell wall; M, mitochondrion; P, peroxisome.</p

<p>Leaves infiltrated with HrcC^- (OD₆₀₀ 0.1),... more <p>Leaves infiltrated with HrcC^- (OD₆₀₀ 0.1), flg22 (1 μM or 10 μM), or BTH (10 μM or 300 μM) were quantified for the formation of abnormal growths 4 days post treatment, using a dissecting microscope. (A) Flg22-induced cell enlargement is FLS2-dependent. (B) HrcC^- partially requires FLS2 to induce large cells. (C) BTH treatment does not induce abnormal growth. (D) HrcC^--induced abnormal growth requires SID2 and NPR1. For quantification of abnormal growths, at least 25 leaves from 12 plants were used for each BTH treatment or bacterial infection and 18 leaves from 7 plants were used for each flg22 treatment. Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Asterisks in (A), (B), and (D) indicate significant difference between treatments of the same genotype (P<0.05). </p

<p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>P. syr... more <p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>P. syringae</i> strains and observed for leaf morphology. (A) Pictures of leaf cross sections. Infected leaves were collected at 4 dpi and fixed for embedding with LR White resin. One-micron sections were cut and stained with 1% toluidine blue O for photographing. Leaves infected with DG3 (0.01) were mostly dead at 4 dpi and thus no data is available. Arrows indicate enlarged cells. The size bar represents 200 μm and applies to all images. Each growth (or a protrusion) has multiple enlarged cells. (B) Quantitation of abnormal growths. The number of abnormal growths, appearing to be transparent protrusions on the treated leaves, was counted at 4 dpi with a dissecting microscope. At least 25 leaves from each treatment were used in the counting. Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Different letters indicate significant difference among the samples (P<0.05). </p

<p><i>P. syringae</i>- infected leaves were embedded in paraplast and cut into ... more <p><i>P. syringae</i>- infected leaves were embedded in paraplast and cut into 15 μm sections for DAPI staining. Images of leaf cross-sections stained with DAPI to show nuclei were captured with a constant exposure time, using a Nikon DS cooled camera attached to a compound microscope. The images are from the following treatments: (A) 10 mM MgSO4, (B) DG34 0.01, (C) DG34 0.001, (D) HrcC^- 0.1, and (E) HrcC^- 0.01. (F) Typical nuclei of guard cells from a mock-treated leaf. Guard cells from leaves infected with DG34 0.01, DG34 0.001, HrcC^- 0.1, or HrcC^- 0.01 show visually similar nuclei (data not shown). (G) A typical nucleus of a normal mesophyll cell from a mock-treated leaf. Normal sized mesophyll cells from leaves infected with DG34 0.01, DG34 0.001, HrcC^- 0.1, or HrcC^- 0.01 show visually similar nuclei (data not shown). (H) A typical nucleus of an enlarged mesophyll cell induced by HrcC^- 0.01. Enlarged cells from leaves infected with DG34 0.01, DG34 0.001, or HrcC^- 0.1 also show large nuclei (data not shown). Arrows indicate the large nuclei in (A) to (E). The size bar in (A) represents 100 μm and applies to panels (A) to (E) while the size bar in (F) represents 5 μm and applies to panels (F) to (H). (J) Relative nuclear DNA content. The average fluorescence of nuclei of guard cells from mock-treated leaves (1) was set as 2C and was used to quantify relative nuclear DNA content of normal mesophyll cells from mock-treated leaves (2) and the enlarged mesophyll cells induced by HrcC^- (0.01) (3). At least 60 nuclei were used for each data point. Nuclear DNA contents of guard cells and normal mesophyll cells from HrcC^- (0.01)-infected leaves are similar to those of their corresponding cells from mock-treated leaves (data not shown). Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Different letters indicate significant difference among the samples (P<0.05).</p

<p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>Pseudo... more <p>The fourth to sixth leaves of 30-day-old Col-0 plants were infected with <i>Pseudomonas syringae. </i><i>pv. </i><i>maculicola</i> ES4326 strains, DG3 at OD₆₀₀ 0.01 or 0.001, DG34 at OD₆₀₀ 0.01 or 0.001, or HrcC^- at OD₆₀₀ 0.1 or 0.01. The infected leaves were collected at the indicated times for SA and RNA analysis. (A) SA quantitation by HPLC analysis. Statistical analysis was performed with one-way ANOVA Fisher’s PLSD tests (StatView 5.0.1). Different letters indicate significant difference among the samples at the same time point (P<0.05). (B) Northern blotting for PR1 expression. Image of rRNA was used for a loading control. </p

Plants are constantly challenged by pathogens. Studies from several plant-pathosystems indicate t... more Plants are constantly challenged by pathogens. Studies from several plant-pathosystems indicate that manipulation of host cell fate might be a common strategy that pathogens use to succeed in their attacks. Genetic perturbation of the key defense signaling mediated by salicylic acid (SA) is also known to lead to altered cell fate in some Arabidopsis mutants. One of such mutants is accelerated cell death 6 (acd6-1), which demonstrates aberrant cell death coupled with cell growth, constitutive defense, and extreme dwarf phenotypes. However, the mechanism by which cell fate is determined during pathogen infection and in some defense mutants has not been well understood. I hypothesize that precise regulation of cell fate is intimately linked to plant disease resistance. I tested this hypothesis during my PhD research. I have found that: 1) There are dynamic, quantitative, and qualitative differences in the accumulation of the defense signaling molecule salicylic acid, expression of the ...

Molecular Plant Pathology, 2017

Erwinia amylovora is the causal agent of the fire blight disease in some plants of the Rosaceae f... more Erwinia amylovora is the causal agent of the fire blight disease in some plants of the Rosaceae family. The non-host plant Arabidopsis serves as a powerful system for the dissection of mechanisms of resistance to E. amylovora. Although not yet known to mount gene-for-gene resistance to E. amylovora, we found that Arabidopsis activated strong defence signalling mediated by salicylic acid (SA), with kinetics and amplitude similar to that induced by the recognition of the bacterial effector avrRpm1 by the resistance protein RPM1. Genetic analysis further revealed that SA signalling, but not signalling mediated by ethylene (ET) and jasmonic acid (JA), is required for E. amylovora resistance. Erwinia amylovora induces massive callose deposition on infected leaves, which is independent of SA, ET and JA signalling and is necessary for E. amylovora resistance in Arabidopsis. We also observed tumour-like growths on E. amylovora-infected Arabidopsis leaves, which contain enlarged mesophyll cells with increased DNA content and are probably a result of endoreplication. The formation of such growths is largely independent of SA signalling and some E. amylovora effectors. Together, our data reveal signalling requirements for E. amylovora-induced disease resistance, callose deposition and cell fate change in the non-host plant Arabidopsis. Knowledge from this study could facilitate a better understanding of the mechanisms of host defence against E. amylovora and eventually improve host resistance to the pathogen.

Plant physiology, Jan 11, 2015

Precise cell cycle control is critical for plant development and responses to pathogen invasion. ... more Precise cell cycle control is critical for plant development and responses to pathogen invasion. Two homologous cyclin-dependent kinase inhibitor (CKI) genes, SIAMESE (SIM) and SIM-RELATED 1 (SMR1), were recently shown to regulate Arabidopsis defense. However whether these two genes play differential roles in cell cycle and defense control is unknown. In this report, we showed that while acting synergistically to promote endoreplication, SIM and SMR1 play different roles in affecting the ploidy of trichome and leaf cells. In addition, we found that the smr1-1 mutant but not sim-1 was more susceptible to a virulent Pseudomonas syringae strain and this susceptibility could be rescued by activating salicylic acid (SA)-mediated defense. Consistent with these results, smr1-1 partially suppressed dwarfism, high SA levels, and cell death phenotypes in acd6-1, a mutant used to gauge the change of defense levels. Thus SMR1 functions partly through SA in defense control. The differential role...

Plant Physiology, 2011

The salicylic acid (SA) regulatory gene HOPW1-1-INTERACTING3 (WIN3) was previously shown to confe... more The salicylic acid (SA) regulatory gene HOPW1-1-INTERACTING3 (WIN3) was previously shown to confer resistance to the biotrophic pathogen Pseudomonas syringae. Here, we report that WIN3 controls broad-spectrum disease resistance to the necrotrophic pathogen Botrytis cinerea and contributes to basal defense induced by flg22, a 22-amino acid peptide derived from the conserved region of bacterial flagellin proteins. Genetic analysis indicates that WIN3 acts additively with several known SA regulators, including PHYTOALEXIN DEFICIENT4, NONEXPRESSOR OF PR GENES1 (NPR1), and SA INDUCTION-DEFICIENT2, in regulating SA accumulation, cell death, and/or disease resistance in the Arabidopsis (Arabidopsis thaliana) mutant acd6-1. Interestingly, expression of WIN3 is also dependent on these SA regulators and can be activated by cell death, suggesting that WIN3-mediated signaling is interconnected with those derived from other SA regulators and cell death. Surprisingly, we found that WIN3 and NPR1 ...