Understanding how pathogen access nutrients from plant are very important to study plant-pathogen interaction. According to previous research result from Chen et. al, SWEET gene family which encode sugar transport plays a crucial role in glucose delivery from plant to pathogen. Our long-term goal here is to determine the mechanism of Pseudomonas syringae pv tomato DC3000Arabidopsis interaction in SWEET sugar efflux pathway. Prior research published indicated that the infection of Pst . DC3000 on Arabidopsis is a type III secretion system-dependent manner. In this proposal, first, we plan to identify the type III effector AvrSWEET15 injected by Pst . DC3000 to trigger SWEET gene expression. Second, we will identify the transcription factor SWEEP (SWEET15-interacting Protein) directly binding to
SWEET gene promoter that supposed to act downstream of AvrSWEET15 effectortriggered nutrient transport signaling pathway. The proposal can provide a model system for pathogen-plant interaction and fill the gap that little is known about how pathogens obtain nutrients from living plant cells. The application of this knowledge would help solve crop yield loss caused by pathogen in agricultural field.
A majority number of plant pathogens cause disease and lead to dramatic loss in crops. At least 10% of global food production is lost by plant disease
(Strange, 2005). It is difficult to control pathogens because pathogens and plants coevolve all the time. Plants develop ways to limit pathogen access to nutrients and initiate defense response, whereas pathogens is smart enough to evolve approaches to fight against host immune response to obtain nutrients from plants. However, the mechanism behind the interaction of pathogen and host plant related to nutrients is not fully understood.
Fungal pathogens include two groups: necrotrophs and biotrophs. Necrotrophic pathogens can cause host plant programmed cell death and absorb nutrients from the dead plant cell to fuel their growth. In contrast, biotrophic pathogens can grow within living plant cells, which means they must obtain nutrients from cell alive. The bacterial pathogen P . syringae is often considered as biotroph, occasionally considered a necrotroph. Many strains, including P. syringae pv. tomato DC3000, infect plants by producing toxin that contribute to pathogenicity. P . syringae actively transport dozens of proteins (known as type III effector) into host cells through type
III secretion system (Glazebrook, 2005). It is reported that P.
syringae can access sugar from plant via type III effector, but how type III effector trigger the sugar efflux from plant to P.
syringae is still a question.
Chen et al. [1] found that besides inhibition of plant immunity, type III effectors and some TAL effectors can function specifically in diverting nutritional resources from

the host [1] . Xanthomonas oryzae pathovar oryzae (Xoo) accesses nutrients from rice by secreting transcription activator-like (TAL) effector to activate transcription of
OsSWEET11 , inducing sugar efflux to feed bacteria in the xylem and apoplasm. Also,
SWEET genes can be induced by Pseudomonas syringae DC3000 in Arabidopsis .
AtSWEET mRNAs is induced by the phytobacterium Psedomonas syringae pv. tomato
DC3000 (Li-Qing Chen, 2010). Among the seventeen members in the AtSWEET family, AtSWEET15 is highly induced, which may indicate its important role in the
Pst DC3000 infection. In addition, the Pst . DC3000 type III secretion system (T3SS) mutant (∆ hrcU ) cannot induce AtSWEET15 . Therefore, I mainly focus on studying the mechanism of AtSWEET15 induced by Pst DC3000 in a T3SS-dependent manner . SWEET15, also named SAG29, is a membrane-localized MtN3 protein, regulates senescing process and cell viability under high salinity (Pil Joon Seo, 2011).
AtSWEET15 serves as a molecular link that integrates environmental stress responses into the senescing process. Though it is known that a TAL effector binds to OsSWEET11 promoter to activate its transcription and promote mRNA level, the mechanism in Arabidopsis is not clearly determined by current research. There are still several questions to be solved. Does Pst DC3000 produce some type III effectors that have similar function as TAL effectors of Xoo to induce SWEET mRNA expression level directly? If not, is the increased SWEET mRNA expression level caused by host transcription factors, which is activated by type III effectors via signaling pathway? If it is, those transcription factors and the related signaling pathways are to be determined.
Nearly 60 Type III effectors are identified in Pseudomonas syringae pv DC3000
(Lindeberg, 2012). AvrPto blocks plant immune responses by binding receptor kinase FLS2, EFR in Arabidopsis and tomato LeFLS2 (Tingting Xiang, 2008).
AvrPtoB binds Bti9 and SILyk13 to suppress tomato immunity by inhibiting their kinase activity (Lirong Zeng, 2012). HopF2 interacts with Arabidopsis thaliana Map
Kinase Kinase5 (MKK5) to inhibit MPKs and PAMP-triggered immunity (Yujing
Wang, 2010). HopF2Pto targets Arabidopsis RIN4 protein to promote Pseudomonas syringae virulence (Mike Wilton, 2010). Besides pst DC3000 type III effectors interfere in plant immunity by binding to receptor kinase or other protein, type III effectors develop ways to negatively regulate host PAMP-triggered immunity (Sruti
DebRoy, 2004), effector-triggered immunity (Guo M, 2009), programmed cell death
(Jamir Y, 2004) or salicylic acid-mediated defense response (Sruti DebRoy, 2004).
However, research areas related to the pathogen access nutrients from host plants are very little. Therefore, this project can make up this gap and supplement the former discovery that pathogen employs SWEET family to transport sugar for their growth. Here, in our proposal, we use ArabidopsisPseudomonas syringae pv tomato
DC3000as the plant-pathogen model to explore the interaction between Type III effector and AtSWEET family.

We hypothesize that a type III effector protein of
Pst DC3000 binds AtSWEET promoter directly to induce transcription or indirectly by activating downstream components in SWEET signaling pathway. However, based on the functions of nearly 60 type III effectors identified in Pst . DC3000, none of them has the function like TAL effector of Xoo . Therefore, this proposal would mainly focus on the latter: Pst . DC3000 effector injected by type III secretion system indirectly induces AtSWEET transcription by activating downstream factors of
SWEET signaling pathway.
However, the effectors related to sugar-efflux signaling pathway is still need to identify since no such effectors have been reported. Our first step is to identify Pst
DC3000 effectors by screening strain mutants. After that, we plan to find other host transcription factors involving in the activation of AtSWEET15 gene expression upon
Pst DC3000 infection.
My main objective here is to identify this type III effector of Pst. DC3000 (named
AvrSweet15) contributing to the increasing expression level of AtSWEET15 and the mechanism about whether it directly binds to AtSWEET15 promoter region or indirectly.
Specific objective 1: Identify Pst . DC3000 effector that increase AtSWEET15 mRNA level. Working hypothesis: An effector secreted by Pst . DC3000 T3SS would indirectly induce the transcription level AtSWEET15 gene.
Specific objective 2: Determine the host transcription factors that directly interact with SWEET15 gene to induce its transcription. Working hypothesis: Some transcription factors targeting AtSWEET promoter would activate its transcription.
Pst
AtSWEET15
General Hypothesis: a type III effector injected by Pst . DC3000 indirectly activates
AtSWEET15 transcription.
Working hypothesis:
1) A type III Pst . DC3000 effector (name AvrSWEET15) will be identified by Pst .
DC3000 mutants screens.
2) AvrSWEET15 does not bind to AtSWEET15 promoter by gel mobility shift assay
(GMSA).
3) Arabidopsis accumulates more glucose when infected with Pst . DC3000 mutant
ΔavrSWEET15 compared with wild-type Pst . DC3000. The number of Pst . DC3000 is larger in overexpression SWEET15 line than sweet15 mutant.

Preliminary results: Based on the preliminary results from Chen et.al (Li-Qing Chen,
2010), AtSWEET mRNAs are induced highly by the bacterium Pseudomonas syringae
Figure 1
Induction of
AtSWEET mRNAs by either the bacterium
Pseudomonas syringae pv. tomato DC3000 (2 ×108 c.f.u. ml−1, 8 h after inoculation, measured by qPCR, normalized by MgCl2 buffer treatment) pv. tomato DC3000 infection measured by qPCR in Arabidopsis leaves. As shown in
Figure 1, AtSWEET15 mRNA increased highest among those AtSWEET mRNAs. That is also the main reason for us to pick AtSWEET15 as the main gene to identify Pst .
DC3000 effector.
Inoculation Arabidopsis leaves with Pst . DC3000 type III secretion mutant ( hrcU ) that cannot inject type III effector into the host, AtSWEET15 mRNA expression level decreased dramatically 6h after inoculation compared with wild type Pst . DC3000
(Figure 2). This indicates that SWEET mRNA abundance is modulated in a type-IIIdependent manner. Based on these preliminary data (Li-Qing Chen, 2010), therefore, we focus on type III effectors of Pst . DC3000 that induce AtSWEET15 mRNA expression. Seen from the pseudomonas website (Lindeberg, 2012), almost all of the sixty effectors are proteins secreted by type III secretion system.

Figure 2
I nduction of AtSWEET4,
AtSWEET5 and AtSWEET15 by P. syringae DC3000 depends on a functional type III secretion system
(T3S). Samples were collected at
6, 12 and 24 h after infiltration with 2 × 108 c.f.u. ml−1 of
DC3000 or DC3000 hrcU, a T3S mutant.
We plan to identify this type III effector by screening all the isogenic strains of Pst .
DC3000 T1 published online and investigate the change of AtSWEET15 mRNA expression level infected by these strains.
I. Obtain isogenic strains of Pst . DC3000 effector mutants and identify candidate effector(s)
Nearly 60 effectors published on Pst.DC3000 webpage, we will screen the effectors to find those can induce AtSWEET15 mRNA expression level. Firstly, we would contact owners that possess already known Pst . DC3000 strains. For those cannot obtain successfully, we would generate the isogenic strains Pst . DC3000 T1 carrying a plasmid with or without the aimed effector by our lab. We can learn from (Yao Luo,
2009), they have such experience when constructing Pst . DC3000 effector mutants with and without avrPto.
After we obtain the candidate Pst . DC3000 isogenic strains. We will inoculate wild type Arabidopsis with those isogenic strains and wild type Pst . DC3000 as control. 8 hours after inoculation, we will analyze AtSWEET15 transcript level in Arabidopsis leaves by qPCR.
Expected results: The strain(s) without the candidate effector(s) would result in significantly decreased AtSWEET15 mRNA expression level compared with WT, because it cannot induce AtSWEET15 transcription without the effector(s). Whereas the rest strains have similar AtSWEET15 mRNA expression level as the control Pst .
DC3000, since they contain the candidate effectors that would induce AtSWEET15

transcription. If identify more than one effector, we would focus on the one that has the most obvious effect and name it AvrSWEET15.
II. Determine the mechanism of AvrSWEET15 in activation of AtSWEET15 mRNA expression
Our objective here is to find AvrSWEET15 that can activate AtSWEET15 transcription by directly binds to AtSWEET15 promoter or indirectly.
Hypothesis: There is no direct interaction between AvrSWEET15 and AtSWEET15 promoter.
To test this hypothesis, we will use gel mobility shift assay (GMSA). Set both negative and positive controls. We plan to prepare DNA of AtSWEET15 promoter probe first. After binding reaction, do electrophoresis analysis.
Expected results: we would expect no bands are detected, which means that
AvrSWEET15 does not bind to AtSWEET15 promoter region. It will address our hypothesis that AvrSWEET15 interacts with AtSWEET15 is indirect, leading to the hypothesis that there maybe other transcription factors induced by AvrSWEET15 involving in AtSWEET15 transcription.
III. Characterize the glucose accumulation of Arabidopsis when inoculated by
Pst . DC3000 and growth of Pst . DC3000 strain
We first generate
1) Plant material: SWEET15 Arabidopsis knockout line and overexpression line;
2) Pathogen material: Pst . DC3000 strains: Pst . DC3000 avrSWEET15, wild-type Pst .
DC3000 strains
Then we 1) inoculate wide type Arabidopsis with Pst . DC3000 avrSWEET15 and wild-type Pst . DC3000 strains at 5X10 4 CFU/ml (Jorge L. Badel, 2006)to measure glucose accumulation and observe the growth of plant to determine the bacterial virulence; 2) inoculate SWEET15 , WT Col-0, and OX SWEET15 Arabidopsis with wild-type Pst . DC3000 to do growth assay of Pst . DC3000. 4 Days after infiltration with bacteria, we would record the disease symptoms of plant, records including two aspects: whole plant, individual leaves. At the same time, we will quantify bacterial populations in leaves of sweet15 , WT Col-0, and overexpression SWEET15
Arabidopsis transgenic plants after 0 and 4 days following inoculation by DC3000.
Expected results: 1) Inoculate WT Col-0 with Pst . DC3000 avrSWEET15, wild-type
Pst . DC3000 see host plant’s phenotype. WT Col-0 grows much better under Pst .
DC3000 avrSWEET15 than Pst . DC3000 in which the leaves may turn yellow; WT
Col-0 accumulates more glucose when infected with Pst . DC3000 mutant avrSWEET15 than wild-type Pst . DC3000.
2) Infiltrate sweet15 , WT, and OX SWEET15 Arabidopsis by Pst . DC3000, do quantitative bacterial populations. No significant difference between these plants 0 dai (day after inoculation). Whereas 4 dai , compared with Pst . DC3000 growth

under Col-0, the number of Pst . DC3000 would significantly increase under OX
SWEET15 showing enhanced growth and significantly reduce under sweet15 .
Problems and limitations: It is laborious and time-consuming to identify the expected effector by screening all of Pst . DC3000 strains. In addition, it is not so easy to obtain all of the known Pst. DC3000 strains. If cannot acquire from other owners, we would construct the strain mutants by our lab. It is possible that some effectors function simultaneously to induce AtSWEET15 transcription, but here we only focus on the one that has the most obvious impact on AtSWEET15 transcription. Secondly, if the phenotype of sweet15 mutant is not obvious, which may mean that other
SWEET family members may function redundantly, then we will generate sweet
RNA interference plant to silence this gene family. Thirdly, since our hypothesis is based on the functions of type III effectors of Pst . DC3000 discovered by now, we come to suspect there is also no such effector function like TAL effector of Xoo .
However, if our actual result were contrast with the expected results, we would come to conclude that a new function of type III Pst . DC3000 effector is identified, which can also target DNA promoter as TAL effector. It would be definitely an encouraging result because TAL effectors are attracting more and more attentions.
Scientists exploit the modularity of TAL effector-DNA recognition for DNA targeting to achieve control over the genetic material in vivo (Adam J. Bogdanove1, 2011).
Therefore, if Pst . DC3000 has this type effector, we expect it would be applied in the real world.
Timetable:
Working Objectives Year
Collect all the identified Pst . DC3000 mutants
Screen the collected mutants to identify the interested effector
Construct sweet , OX SWEET transgenic plants
Confirm the function of AvrSWEET15
Hypothesis: in this objective, based on the main hypothesis that AvrSWEET15 induce AtSWEET15 mRNA expression indirectly, we hypothesize that a host transcription factor results in the activation of AtSWEET15 transcription by binding to AtSWEET15 promoter.
Working hypothesis:
1) A host transcription factor (named SWEEP) directly binds to AtSWEET15 promoter will be identified by yeast-one-hybrid screens.
2) SWEEP transcription factor is in the same signaling pathway with AvrSWEET15 contributing to the sugar delivery from host to bacterial.
1 st – 2 nd
2 nd – 3 rd
1 st – 2 nd
3 rd

Preliminary results: Xoo PXO99 A infects rice by injecting type III effector PthXo1 that is a transcriptional activator-like (TAL) effector (Yang B, 2006). Chen et. al indicated that PthXo1 directly interacts with OsSWEET11 promoter by chromatin immune precipitation (Figure 3) (Li-Qing Chen, 2010). By this way, PthXo1 activates transcription of OsSWEET11 , presumably to induce sugar efflux to feed bacterial in the xylem and apoplasm (Figure 4) (Li-Qing Chen, 2010).
Figure 3
Enrichment of 5’-upstream
OsSWEET11 promoter DNA upon infection of rice with PXO99A containing Flag-tagged PthXo1-2F
Figure 4
The pathogenic Xoo strain
PXO99A injects TAL effector
PthXo1 via the type III secretion system into rice cells.
PthXo1 directly induces
OsSWEET11 leading to sugar efflux. Bacteria take up glucose and multiply.
However, no such type III effectors have ever been reported in Pst . DC3000. It is the main reason for us not to emphasis our research on identifying the effector that directly binds to AtSWEET15 promoter. Instead, we suppose AvrSWEET15 indirectly induce the transcription of AtSWEET15. Therefore, besides identifying
AvrSWEET15, our aim is to identify the most directly transcription factor (TF) that binds AtSWEET15 gene, even though there are maybe a lot of factors linking this type III effector and TF. Here, we will use biochemical approach to identify this TF and confirm its interaction with AtSWEET15 gene in vivo.

I. Identification of potential TF interacting with AtSWEET15 by Yeast One-
Hybrid screens and GMSA
By Yeast one-hybrid screening, we will firstly identify the potential TF using our interested regionAtSWEET15 promoter as the bait sequence. After we get yeast clones, we will focus on potential TF we are mostly interested even though maybe there are several candidate proteins and name it SWEET15-interacting Protein
(SWEEP).
To confirm the direct interaction between SWEEP protein and AtSWEET15 gene promoter in vivo, we will conduct gel mobility shift assay. Here, we will use the probe as mentioned in the objective 1-II.
Expected results: In yeast one-hybrid screening, we will identify several promising transcription factor proteins and for this proposal we would only focus on one protein SWEEP. In GMSA, we would expect there are DNA bands with the probe compared with that have specific competitor which SWEET protein binds to instead of binding to AtSWEET15 gene.
II. Determine whether SWEEP function in the same signaling pathway with
AvrSWEET15 by genetic approach
After we identify SWEEP transcription factor, we want to explore the function of
SWEEP in the sugar efflux-signaling pathway. We have known that Pst . DC3000 injects AvrSWEET15 type III effector into plant cell. Then we would ask, does
AvrSWEET15 release signal to SWEEP via other unknown factors, then SWEEP transcription factor activating AtSWEET15 transcription to help the transport of sugar from plant to bacterial?
To validate our hypothesis, we will generate a T-DNA knockout mutant sweep , sweep complementary line, SWEEP overexpression line driven by CaMV 35S promoter. We first make constructs of SWEEP gene with a GFP-tag driven by CaMV 35S promoter.
Then we will introduce 35S::SWEEP-GFP into Arabidopsis plants by floral-dip methods to produce overexpression line. In addition, we would order SWEEP T-DNA knock out seeds from Arabidopsis Biological Resource Center ( www.arabidopsis.org
) and identify homozygous mutant T-DNA insertion in promoter by PCR genotyping with specific primers. In addition, we will generate a complementary line for sweep mutant with 35S::SWEET-GFP to see whether it can restore the phenotype of sweet mutant. After we get these transgenic plants, we will inoculate sweep , complementary plants of sweep , WT, overexpression SWEEP plant with infiltration of Pst . DC3000 carrying the plasmid with AvrSWEET15, without AvrSWEET15 as the control. Next, we will measure the bacterial growth in leaves over a period of 6 days and observe the phenotype of these plants upon Pst . DC3000 infection.
Expected results:
Inoculation of Pst . DC3000: over a period of 6 days (Robert B.Abramovitch, 2003), the growth curve of bacterial population would increase gradually within a plant.
For the different transgenic plants, we would expect that bacterial population of Pst .
DC3000 with AvrSWEET15 is the largest in SWEEP overexpression line, the smallest

in sweep mutant compared with WT; while for those in sweep mutant, it is similar with the control that inoculated with only MgCl
2
since we hypothesize that sweep mutant is insensitive to the infection of AvrSWEET15. For sweep complementary mutant, the number of bacterial is more than that in sweep , which has similar result with that in WT, because SWEEP restores the insensitivity of sweep to AvrSWEET15.
For Pst . DC3000 without AvrSWEET15, we expect that the populations in the three plants should be of no significant difference.
If we get those expected results, we would come to the conclusion that SWEEP transcription factor is indeed in the same pathway with AvrSWEET15 involving sugar delivery from host to bacterial.
Problems and limitations:
First, from Yeast one-Hybrid screens, we may obtain more than one candidate TFs, but in our research we only focus on one TF that contribute most to the SWEET15 transcription activation. Therefore, in the future work, we would explore the function and mechanism of other TFs (maybe as cofactors of SWEEP TF) to give a broad network of transcriptional regulation in the sugar-efflux signaling pathway.
Timetable:
Working Objectives
Identify TF SWEEP
Construct sweep , sweep /SWEEP, OX SWEEP transgenic plants
Characterize the function of SWEEP
Year
1 st
2 nd – 3 rd
3 rd
The newly identified sugar transporters, SWEETs, show some evidence to the long unsolved question: how pathogens obtain nutrients from plant? According to previous research, SWEET mRNA expression level increased upon pathogen infection in Arabidopsis and rice. The mechanism for rice is clearly explained. A transcription-activation-like effector injected by Xoo binds to
SWEET promoter to activate SWEET transcription, whereas there is no such explanation for Arabidopsis because no similar effector of Pst . DC3000 with Xoo.
Therefore, our proposal here would fill the gap about the mechanism how AtSWEET transcription is triggered by Pst. DC3000 infection. Additionally, our work can be applied into agricultural field and act as a guideline for those want to control pathogens growth by cutting their way to nutrients.
Our lab has rich experience in conducting research by genetic and biochemical approach. The approaches list here: cross technique, generate mutant or RNAi plant or overexpression mutant, Yeast one hybrid, gel mobility shift assay, transient coexpression assay. Besides, our lab includes a range of equipment that is needed in this research: greenhouse chamber, refrigerator, PCR machine, 37 and 30 incubator, and shaker. The facilities in university are very close to our lab. The DNA facility in Molecular Biology Building provides support in areas like DNA sequencing and synthesis. The Microscopy and NanoImaging Facility (MNIF) in Bessey Hall

provides a variety of instrumentation in light and fluorescence microscopy. In general, our research would be carried out well with the above experience and support.
In this proposal, we focus on exploring the mechanism about how Pst . DC3000 manipulate sugar transport from Arabidopsis to favor pathogen growth by inducing AtSWEET15 mRNA expression via transcriptional regulation. In here, our aims are to identify the type III effector AvrSWEET15 and transcription factor SWEEP in this sugar-efflux signaling pathway. If we obtain the expected results, we would come to conclude that Pst . DC3000 induces SWEET15 transcription by injecting type III effector AvrSWEET15 that can transduce signals to transcription factor SWEEP to activate AtSWEET15 transcription by binding to the promoter, thus facilitating sugar delivery from plant to bacterial.
However, predicted from our results, factors that downstream of AvrSWEET15 and upstream of SWEEP is unknown. In our future work, we are interested in characterize those components that involved in sugar-efflux signaling pathway. It would help to form a more detailed work model about how type III effector promotes AtSWEET15 mRNA expression. In addition, we are interested in the translational regulation after
AtSWEET15 mRNA is synthesized and the role of AtSWEET15 protein in sugar transport.
1) Identify components between type III effector AvrSWEET15 and transcription factor SWEEP in sugar-efflux signaling pathway.
2) Translational regulation after SWEET15 mRNA expression and the function of
AtSWEET15 protein in sugar transport.
SWEET sugar-efflux pathway model

Objective
Specific objective 1: identify Pst . DC3000 type III effector AvrSWEET15
Collect all the identified Pst . DC3000 mutants
Screen the collected mutants to identify the interested effector
Construct sweet , OX SWEET transgenic plants
Confirm the function of AvrSWEET15
Specific objective 2: identify transcription factor SWEEP in Arabidopsis
Identify TF SWEEP
Construct sweep , sweep /SWEEP, OX SWEEP transgenic plants
Characterize the function of SWEEP
1.
Adam J. Bogdanove1, D. F. (2011). TAL Effectors: Customizable Proteins for
DNA Targeting. Science , 1843-1846.
2.
Glazebrook, J. (2005). CONTRASTING MECHANISMS OF DEFENSE AGAINST
BIOTROPHIC AND NECROTROPHIC PATHOGENS . Annu. Rev. Phytopathol ,
205–27.
3.
Guo M, T. F. (2009). The Majority of the Type III Effector Inventory of
Pseudomonas syringae pv. tomato DC3000 Can Suppress Plant Immunity.
Mol Plant Microbe Interact , 1069-1080.
4.
Jamir Y, G. M.-O. (2004). Identification of Pseudomonas syringae type III effectors that can suppress programmed cell death in plants and yeast. Plant
Journal , 554-65.
5.
Jorge L. Badel, R. S.-S. (2006). A Pseudomonas syringae pv. tomato avrE1/hopM1 Mutant Is Severely Reduced in Growth and Lesion Formation in Tomato . MPMI , 99–111.
6.
Lindeberg, M. (2012, October 14). Pesudomonas-Plant Interaction . Retrieved
October 3, 2004, from P. syringae - Plant Interaction Resource: http://www.pseudomonas-syringae.org/Pto-plant-interaction.html#avrE
7.
Li-Qing Chen, B.-H. H.-Q.-J.-G. (2010, November). Sugar transporters for intercellular exchange and nutrition of pathogens. Nature , 527–532.
8.
iron en . . ( 1 . tomato ys receptor-like kinase promotes immunity and its kinase activity is inhibited by AvrPtoB . the Plant Journal ,
92-103.
9.
Mike Wilton, R. S. (2010). The type III effector HopF2Pto targets Arabidopsis
RIN4 protein to promote Pseudomonas syringae virulence. PNAS , 2349-54. year
1 st – 2 nd
2 nd – 3 rd
1 st – 2 nd
3 rd
1 st
2 nd – 3 rd
3 rd

10.
Park, P. J.-M.-G.-M. (2011). An Arabidopsis senescence-associated protein
SAG29 regulates cell viability under high salinity. Planta , 189-200.
11.
Robert B.Abramovitch, Y.-J. K. (2003). Pseudomonas type III effector AvrPtoB induces plant disease susceptibility by inhibition of host programmed cell death . EMBO Journal , 22 , 60-69.
12.
Sruti DebRoy, R. T.-B. (2004). A family of conserved bacterial effectors inhibits salicylic acid-mediated basal immunity and promotes disease necrosis in plants. PNAS , 99227-9932.
13.
Strange, R. N. (2005). Plant Disease: A Threat to Global Food Security. Annual
Review of Phytopathology , 43 , 83-116.
14.
Talbot, N. J. (2010). Living the Sweet Life: How Does a Plant Pathogenic
Fungus Acquire Sugar from Plants? . PLoS Biology , 8 (2), 1-3.
15.
Tingting Xiang, N. Z.-M. (2008). Pseudomonas syringae Effector AvrPto
Blocks Innate Immunity by Targeting Receptor Kinases. Current Biology , 74-
80.
16.
Yang B, S. A. (2006). Os8N3 is a host disease-susceptibility gene for bacterial blight of rice. Proc Natl Acad Sci USA , 10503-10508.
17.
Yao Luo, K. S. (2009). Proteolysis of a Negative Regulator of Innate Immunity
Is Dependent on Resistance Genes in Tomato and Nicotiana benthamiana and
Induced by Multiple Bacterial Effectors . The Plant Cell , 21 , 2458-2472.
18.
Yujing Wang, J. L.-M. (2010). A Pseudomonas syringae ADP-
Ribosyltransferase Inhibits Arabidopsis Mitogen-Activated Protein Kinase
Kinases. 2033-2044.

Project Budget Worksheet - Iowa State University of Science and Technology
Program Sponsor
Title
PI 3
Period of Performance
Deadline
1/1/2013-12/31/2017
Year 1
A
6
7
8
9
4
5
1
2
3
Key Personnel
B Other Personnel
1 Post Doc
2 Post Doc
3 Research Asst-Halftime
4 Research Asst-Halftime
5 Hourly Undergraduate student
6 Hourly Undergraduate student
7 P&S
8 P&S
9 Secretarial/Clerical
10 Secretarial/Clerical
11 Non-Student Hourly
12 Non-Student Hourly
Subtotal: Salaries and Wages
Monthly
$0
$0
$1,900
$0
$300
$0
$0
$0
$0
$0
$0
$0
Salary
Monthly
$5,500
$0
$0
$0
$0
$0
$0
$0
$0
C Fringe Benefits
0
0
0
0
0
0
0
0
0
Post Doc
Post Doc
Research Asst-Halftime
Research Asst-Halftime
Hourly Undergraduate student
Hourly Undergraduate student
P&S
P&S
Secretarial/Clerical
Secretarial/Clerical
Non-Student Hourly
Non-Student Hourly
30.5%
30.5%
30.5%
22.0%
22.0%
12.9%
12.9%
4.6%
4.6%
Rate
30.5%
30.5%
30.5%
30.5%
30.5%
30.5%
37.0%
37.0%
49.7%
49.7%
12.0%
12.0%
0.00
0.00
12.00
0.00
9.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Calendar
Months
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Calendar
Months
Academic
Months
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
2.00
0.00
1.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Summer
Months
3.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Number of persons
$16,500
$16,500
$0
$0
$0
$0
$0
$0
$0
$0
$48,300
$0
$0
$45,600
$0
$2,700
$0
$0
$0
$0
$0
$0
$0
$64,800
$11,039
$5,033
$5,882
$0
$124
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
Year 2
$49,749
$0
$0
$46,968
$0
$2,781
$0
$0
$0
$0
$0
$0
$0
$66,744
$16,995
$16,995
$0
$0
$0
$0
$0
$0
$0
$0
$11,370
$5,183
$6,059
$0
$128
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
Year 3
$51,241
$0
$0
$48,377
$0
$2,864
$0
$0
$0
$0
$0
$0
$0
$68,746
$17,505
$17,505
$0
$0
$0
$0
$0
$0
$0
$0
$11,711
$5,339
$6,241
$0
$132
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
Year 4
$52,779
$0
$0
$49,828
$0
$2,950
$0
$0
$0
$0
$0
$0
$0
$70,809
$18,030
$18,030
$0
$0
$0
$0
$0
$0
$0
$0
$12,063
$5,499
$6,428
$0
$136
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
Year 5
$54,362
$0
$0
$51,323
$0
$3,039
$0
$0
$0
$0
$0
$0
$0
$72,933
$18,571
$18,571
$0
$0
$0
$0
$0
$0
$0
$0
$12,425
$5,664
$6,621
$0
$140
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
Form updated 7-27-12
Eff. 7-1-12
Total c
$87,601
$87,601
$0
$0 Check
$0
$0
$0
$0
$0
$0 $87,600.74
$256,431
$0
$0
$242,097
$0
$14,335
$0
$0
$0
$0
$256,431.26
$344,032
$344,032.00
$58,608
$26,718
$31,230
$0
$659
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$0
$58,608.08
$0
$0
$0
$0
$75,839
$0
$78,114
$0
$80,458
$0
$82,871
$0
$85,358
$0
$402,640
$0 D
E
F
Subtotal: Salaries, Wages, and Benefits
Equipment (List Item and $ amount for each item > $5k)
1
2
Travel
1. Domestic Travel
2. Foreign Travel
Participant Support Cost
1. Stipend
2. Travel
3. Subsistence
4. Other
See notes below
G Other Direct Costs
1 Materials and Supplies
2 Publication cost
3 Computing support
4 Instrumentation facility
5 Subcontractor1 - Subject to IDC (first $25,000) See notes below
NOT subject to IDC (Amount over $25,000)
6 Subcontractor2 - Subject to IDC (first $25,000) See notes below
NOT subject to IDC (Amount over $25,000)
7 Tuition - Non-Engineering
8 Tuition - Engineering
(Click on "Tuition" sheet)
(Click on "Tuition" sheet)
9 Other
10 Other
Subtotal: Total Direct Costs (TDC)
Subtotal: Modified Total Direct Costs
[ MTDC = TDC - Tuition - Equipment - Participant Support Cost ]
H Indirect Costs
IDC on MTDC
Rate
48.0%
[ IDC = MTDC * Indirect Rate ]
I Total Project Costs
[ Total = TDC + IDC ]
$0
$0
$0
$2,000
$0
$2,000
$0
$0
$27,657
$10,000
$0
$3,000
$0
$0
$0
$0
$0
$14,657
$0
$0
$0
$105,496
$88,839
$12,000
$12,000
$117,496
$25,247
$10,000
$0
$0
$0
$0
$0
$0
$0
$15,247
$0
$0
$0
$103,361
$88,114
$0
$0
$0
$0
$0
$0
$0
$0
$12,000
$12,000
$115,361
$3,000
$3,000
$0
$2,000
$0
$2,000
$0
$0
$21,591
$10,000
$1,000
$0
$0
$0
$0
$0
$0
$10,591
$0
$0
$0
$107,049
$94,458
$12,000
$12,000
$119,049
$22,036
$10,000
$1,000
$0
$0
$0
$0
$0
$0
$11,036
$0
$0
$0
$104,908
$93,871
$0
$0
$0
$0
$0
$0
$0
$0
$12,000
$12,000
$116,908
$0
$0
$0
$2,000
$0
$2,000
$0
$0
$22,500
$10,000
$1,000
$0
$0
$0
$0
$0
$0
$11,500
$0
$0
$0
$109,857
$96,358
$12,000
$12,000
$121,857
$3,000
$3,000
$0
$6,000
$0
$6,000
$0
$0
$119,031
$50,000
$3,000
$3,000
$0
$0
$0
$0
$0
$63,031
$0
$0
$0
$530,671 $530,671.24
$461,640
$60,000
$590,671 $590,671.24