Progress 09/01/13 to 08/31/18
Outputs
Target Audience:Our target audience is agricultural scientists and cotton growers, along with high school biology teachers and students. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?Over the course of this project, two PhD students were trained, one Postdoc was trained, one Research technician was engaged, and four undergraduates were given the opporutnity to enagage in scientific research How have the results been disseminated to communities of interest?Students, postdocs and the PI made presentations on this work at several different scientific meetings over the course of this project, and each year at outreach events in Virginia high schools. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Objective 1. Characterization of genes Involved in the P80 senescence regulation complex. We identified genes in two major gene families including the Protein 80 (P80) gene family, and the Ubiquitin Binding Protein 3 (UBP3) gene family. Identifying these genes was important as they are the chief regulators of early senescence in our model plant system. We needed to identify and clone these from cotton to be able to attempt turning them off in cotton, which we hoped would stimulate early senescence. Objective 2. Manipulation of the P80 and InsP network to alter senescence in cotton. We used Viral Induced Gene Silencing (VIGS) to target P80 and UBP3 genes in cotton plants. We established that our VIGS conditions work, and that downregulationof either type of gene family could be accomplished. For both genes, turning them off resulted in plants that hadsevere responses to the virus that we used for this approach. This supports the idea that both types of genes are important forhost defenses against viruses. We think this is an important discovery that creates new genetic targets for engineering hostresistance pathways. In the second portion of this objective we identified two new inositol phosphate kinase (IPK) genefamilies that encode enzymes required to make important signaling molecules. We found these genes are regulated atdifferent times and places in cotton plants. Further, we obtained data linking production of one of the signaling molecules to aplant hormone named jasmonic acid. This is important as jasmonic acid turns on known host defense responses. In thefuture we can use this information in strategies to make plants more resilient to insect damage. Objective 3. Profiling of higher InsPs in cotton and related plants. We analyzed the production of important inositol phosphate (InsP) signaling molecules in various cotton tissues. This allowed us to conclude that cotton produces higher InsPs in a very unique way. Specifically, cotton makes much more of one unique InsP as compared to most other plants. Close cotton relatives, like Hibiscus, have a similar change in which higher InsPs they accmulate. We think this unique production of signaling molecules in cotton and related plants may be linked to stress tolerance.
Publications
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Progress 09/01/16 to 08/31/17
Outputs
Target Audience:Our target audience is agricultural scientists and cotton growers. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?This project engages the PI, a postdoc, a technician, a graduate student and two undergraduate researchers. The Postdoc travelled to three meetings this year and presented her work (1 international, 1 national, and one regional meeting). She also took the lead in organizing and implementing a 3 day science outreach event that engages 200-300 high school Biology students. The graduate student succesfully defended his Preliminary Exam proposal, an important milestone in his graduate career. The two undergraduate researchers were engaged in weekly lab meetings, and each gave a presentation at our undergraduate research symposiun on campus. One of these students gave a presentation at one of our Agriculture, Research, Extension Centers (ARECs) in Suffolk, Va, and performed research for 5 weeks at another AREC on the Eastern Shore of VA. These experiences help expand this student's knowledge and expertise in translational approaches important for agriculture. How have the results been disseminated to communities of interest? Research presentations have been given atnational meetings: theMid-Atlantic Plant Molecular Biology meeting in Laurel, Maryland in August 2017; theASBMB meeting in Chicago in April 2017. Research has been presented at the 2017NC1200 Photosynthesis Multi-state project meeting in Bozeman, MT. Two undergraduate presentations were made at a local symposium. One undergraduate presentation was made at the Tidewater Agriculture Research Station in Suffolk, VA. What do you plan to do during the next reporting period to accomplish the goals? 1. Producing and characterizing new cotton transgenic lines.We are continuing to screen our transgenic seed populations to develop new cotton transgenic lines. DuringAugust 2017 to February 2018: we will complete pistil drip transformation work, submit detailed protocol and chimera results in a manuscript to Plant Molecular Biology journal. Chimera work will facilitate part B described below. We will prepare existing P80-VIGS and UBP3-VIGS work for publication. FromDec 2018 to August 2018: we will perform detailed molecular and developmental analysis of UBP3, SnRK1.1 and IPK1 transgenic cotton plants from existing T1 seed stocks. 2. Testing the viral host defense function of P80.FromAugust 2017 to Dec 2018: We will characterize cotton P80 and IPK1 overexpressors or transgenic plants. We will isolate UBP3, SnRK1.1 and IPK1 transgenic cotton plants from existing T1 seed stocks. We will prepare IPK1-VIGS work for publication. FromDec 2017 to June 2018: we will challenge P80 plants with virus to determine if P80 overexpression interferes with viral responses. We will examine impact of P80 overexpression on senescence. fromJune 2018-August 2018: we will prepare P80 viral work for publication.
Impacts
What was accomplished under these goals?
Objective 1. Characterization of Genes Involved in the P80 senescence Regulation Complex in Cotton.As stated previously, our plan was to utilize previously identifiedgenes to manipulate the timing of senescence in cotton, which could provide alternatives for defoliation and dessicant chemicals used in farming. To facilitate this, we identified and cloned cDNAs for SnRK1.1, UBP3, IPK1 and P80 from Gossypium hirsutum. We ultimately identified gene families in Gossypium hirsutum for P80 (5 genes), SnRK1.1 (3 genes), UBP3 (3 genes), IPK1 (4 genes) and IPK2 (6 genes). We used RT-PCR to identify the major expressed cDNAs for these genes, and have focused on the isoforms expressed in leaf and early vegetative development for our work. P80, SnRK1.1, UBP3 and IPK1 genes have been used to produce viral-induced gene silencing (VIGS) constructs for use in cotton. Objective 2. Manipulation of the P80 and InsP network to alter senescence in cotton. Over the past year we have completed multiple VIGS experiments in Deltapine90 using the bipartite geminivirus Cotton leaf crumple virus (CLCrV) system, delivered by Agrobacterium infiltration (Tuttle et al. 2008, Tuttle et al. 2012).The most important finding is that suppression of P80 in cotton does not alter the timing of senescence as predicted. Instead, our data show that suppression of P80 facilitates increased viral success, as shown by classical viral symptoms and increased presence of viral DNA.It is important to note that our control GFP-VIGS and ChlI-VIGS never show this response. In addition, we have consulted with R. Tuttle who developed this VIGS system to provide input on these phenotypes. In eight different individual VIGS experiments, we have seen that suppression of P80 increases viral success. We conclude that the cotton P80 gene functions as a host defense gene whose expression helps to limit geminiviral spread. In concordance with this hypothesis, we have shown that geminivirus infection of cotton results in increased P80 gene expression. We also characterized UBP3-VIGS plants and our data show that UBP3 suppression also increases viral success. The implications of these findings is that the P80/UBP3 genes may be useful targets to manipulate viral resistance in plants. We note that geminiviruses are important pathogens, especially when global agriculture is considered. We also have identified important impacts on growth from silencing SnRK1.1 and IPK1 that differ from the increased viral success phenotypes described above. Not surprisingly, suppression of SnRK1.1 greatly impacted overall cotton growth. We have developed transgenic cotton SnRK1.1 overexpressors, which may increase growth, biomass, or stress tolerance as was reported for Arabidopsis (Baena-Gonzalez et al. 2007).As part of our previous work sought to characterize the interplay of InsPs with senescence, we also used VIGS to suppress cotton IPK1 expression. The 500 bp IPK1 fragment used is predicted to suppress expression from two of the four cotton IPK1 genes (named IPK1A and B). We hypothesized that IPK1 silencing would greatly decrease InsP6, as occurs in Arabidopsis ipk1 mutants. Given the unique elevation of Ins(1,2,4,5,6)P5 in cotton, we also suspected that levels of this signaling molecule might decrease as well, since IPK1 is known to phosphorylate Ins(1,4,5,6)P4. We found that suppression of IPK1 reduced growth overall, and levels of Ins(1,2,4,5,6)P5. Our conclusion is that Ins(1,2,4,5,6)P5 is required for optimal growth and development of cotton, and underscores the importance this molecule plays in cotton. The fact that IPK1-VIGS reduces Ins(1,2,4,5,6)P5, but not InsP6, strongly implicates alternative regulation of the InsP pathway in cotton. Manipulation of the P80/InsP network via stable transformation/transgenics: Wehave completed work using the cotton pistil drip transformation process (Zhang, 2012). We have now obtained transgenic AtP80:GFP cotton plants in the T2 generation. We have three transgenic lines in hand, and are screening for more lines. Our recovery rate for transgenics is 1 transgenic plant per 233 seed screened. Using pistil drip transformation we have obtained 2,000-3,000 seed potentially carrying an AtP80-GFP, SnRK1.1 or IPK1 transgene. Given our preliminary results of 1/233 recovery of transgenics, we expect to establish at least 10 founder lines for each construct. Another important finding from our work is that the pistil drip procedure likely leads to T0 chimera embryos, as we found only a portion of T1 plants were transgenic. Objective 3. Profiling of higher InsPs in cotton and related plants.To better understand developmental regulation of various InsPs, we tested whether cotton IPK1 and IPK2 genes are regulated throughout development in cotton. We have three primer sets that amplify 2 IPK2 genes each that we call IPK2AB, IPK2CD and IPK2EF. Our analysis shows that these genes are developmentally regulated, with IPK2CD being most abundant in leaves. We have a set of gene primers that amplify the IPK1A and B genes, and a second set that amplify the IPK1 C and D genes. Our analyses show that IPK1AB CD and are also developmentally regulated, with IPK1AB being the predominantly expressed isoforms in cotton leaves. We next tested whether wounding, MeJA or heat stress induces these genes. We found that IPK1AB is upregulated by wounding, MeJA treatment, and heat stress, while IPK2CD is upregulated by MeJA, and heat, and IPK2AB is induced by heat. Importantly, we found that the 1 hour gene expression induction of IPK1AB by wounding is followed at 1.5 hours by elevated Ins(1,2,4,5,6,)P5 levels. This links Ins(1,2,4,5,6)P5 and IPK1AB gene transcription to a stress response. Further, it suggests that IPK1AB proteins might act mostly or exclusively to convert Ins(1,4,5,6)P4 to Ins(1,2,4,5,6)P5.
Publications
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Progress 09/01/15 to 08/31/16
Outputs
Target Audience:Our target audience is cotton growers and other life scientists interested in the mechanisms of senescence and energy-sensing in plants. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?One graduate student, one scientist, and two undergraduate researchers have been engaged in this project. A postdoc and technician funded by Virignia Tech have also contributed to this project. The PI nad postdoc attended an internatinal meeting and presetned results from this project. A minority undergraduate researcher was engaged on this project and presented his work at our annual summer research symposium on campus. How have the results been disseminated to communities of interest?Research presentations have been given at internaitonal meetings, the NIFA investigator meeting, and a local research symposium. An outreach project to high school students in Virginia conitnued with involvmenet by the postdoc working on this project. What do you plan to do during the next reporting period to accomplish the goals?1. Producing and characterizing new cotton transgenic lines. We are continuing to screen our transgenic seed populations to develop new cotton transgenic lines. We are preparing a manuscript on our refinement of this important technique that could induce more basic science plant researchers to work with cotton. We expect this to make a novel contribution in the literature in that screening procedures and numbers of expected transformants are still not adequately described. Given our preliminary results of 1/233 recovery of transgenics, we expect to establish at least 10 founder lines for each construct. We will use SnRK1.1 and IPK1 lines to test whether productivity/biomass and stress tolerance can be manipulated with these genes, as suggested from VIGS experiments and previous work on SnRK1.1. The IPK1 lines will also be valuable in terms of future work to delineate the function of InsPs in JA, stress and phosphate signaling. The questions we will address over the next year are: Does overexpression of IPK1 lead to changes in InsP6 levels as expected? Further, does this alter growth or physiology of cotton? 2. Testing the viral host defense function of P80. Given the importance of emerging geminiviruses, we will determine whether the P80 gene can be used to protect plants against geminiviruses. We will use the AtP80-GFP cotton plants to test whether overexpression of P80 interferes with geminiviral infection. We will also construct cotton RNAi P80 transgenic plants to further explore the role of P80 in defense pathways.
Impacts
What was accomplished under these goals?
Cotton is the major source of fiber used by humans, and production of cotton has a $3.86 billion dollar impact on the U.S. economy. Because farmers currently use several chemical treatments in the field to maximize yield by inducing senescence, we have sought to develop genetic resources that can be used as a replacement to chemical defoliants. The work centers around a complex of proteins we have shown are involved in regulating energy status and senescence in Arabidopsis. One big surprise was the finding that silencing cotton P80 genes increases viral success, as opposed to turning on senescence. Thus we have identified the cotton P80 genes as potential targets for altering viral infection of cotton. This is important because viruses have a huge economic impact and very few mitigation strategies currently exist. Objective 1. Characterization of Genes Involved in the P80 senescence Regulation Complex in Cotton. We previously identified a novel protein complex in a yeast two hybrid screens inclduing 1) P80, a WD40-containing protein predicted to be involved in regulating protein stability, and 2) Sucrose non-fermenting related kinase 1.1 (SnRK1.1), a protein kinase that is a central regulator of energy metabolism and stress signaling pathways (Williams et al., 2014). We characterized another P80 interactor, called UBP3, that encodes a deubiquinase, and this suggested the mechanism of regulation involves the de-ubiquination of target proteins required for growth and development. As stated previously, our plan was to utilize these genes to manipulate the timing of senescence in cotton. To facilitate this, we identified and cloned cDNAs for SnRK1.1, UBP3, IPK1 and P80 from Gossypium hirsutum. We ultimately identified gene families in Gossypium hirsutum for P80 (5 genes), SnRK1.1 (3 genes), UBP3 (3 genes), and an inositol phosphate kinase 1 gene family (IPK1) (4 genes). We used RT-PCR to identify the major expressed cDNAs for these genes, and have focused on the isoforms expressed in leaf and early vegetative development for our work. Expression studies to determine whether individual isoforms are regulated by stress are on-going. Each of these genes has been used to produce viral-induced gene silencing (VIGS) constructs for use in cotton. Objective 2. Manipulation of the P80 and InsP network to alter senescence in cotton. Over the past 3 years we have completed multiple VIGS experiments in Deltapine90 using the bipartite geminivirus Cotton leaf crumple virus (CLCrV) system, delivered by Agrobacterium infiltration. We use CLCrV constructs carrying the magnesium chelatase subunit I gene (ChlI) or GFP as controls to provide a visible marker for VIGS (ChlI), and a negative control (GFP). Following onset of VIGS symptoms, as measured by bleaching in VIGS-ChlI plants, we routinely used real-time PCR with PCR primers outside of the 500 bp VIGS region to assess whether suppression has occurred. For all genes except IPK1 we have used a 500 bp fragment predicted to silence all expressed isoforms. The most important finding is that suppression of P80 in cotton does not alter the timing of senescence as predicted. Instead, our data show that suppression of P80 facilitates increased viral success, as shown by classical viral symptoms and increased presence of viral DNA. It is important to note that our control GFP-VIGS and ChlI-VIGS never show this response. We also characterized UBP3-VIGS plants and preliminary data indicate UBP3 suppression also increases viral success. The implications of these findings is great in that the P80/UBP3 genes may be useful targets to manipulate viral resistance in plants. We note that geminiviruses are important pathogens, especially when global agriculture is considered. We also have identified important impacts on growth from silencing SnRK1.1and IPK1 that differ from the increased viral success phenotypes described above. Not surprisingly, suppression of SnRK1.1 greatly impacted overall cotton growth (Fig. 3). We are following this finding with development of transgenic cotton SnRK1.1 overexpressors, which may increase growth, biomass, or stress tolerance as was reported for Arabidopsis (Desai et al. 2014). As part of our previous work sought to characterize the interplay of InsPs with senescence, we also used VIGS to suppress cotton IPK1 expression. The 500 bp IPK1 fragment used is predicted to suppress expression from two of the four cotton IPK1 genes. We hypothesized that IPK1 silencing would greatly decrease InsP6, as occurs in Arabidopsis ipk1 mutants. We found that suppression of IPK1 reduced growth overall. The vegetative apex, especially was suppressed in growth. Importantly, we also measured the impact on Ins(1,2,4,5,6)P5 levels in leaves from IPK1-VIGS plants (Fig. 4). Surprisingly, we found levels of InsP6 were not impacted, but levels of Ins(1,2,4,5,6)P5 were decreased. Our conclusion is that Ins(1,2,4,5,6)P5 is required for optimal growth and development of cotton, and underscores the importance this molecule plays in cotton. The fact that IPK1-VIGS reduces Ins(1,2,4,5,6)P5, but not InsP6, strongly implicates alternative regulation of the InsP pathway in cotton. Manipulation of the P80/InsP network via stable transformation/transgenics: We proposed and have completed work using the cotton pistil drip transformation process (Zhang, 2012). To our knowledge we are the first group in North America to use this technique successfully.We have three transgenic lines in hand, and are screening for more lines. Our recovery rate for transgenics is 1 transgenic plant per 233 seed screened. Though the pistil drip procedure worked well to generate potential transgenic seed, one major hurdle we encountered was finding an easy, effective screen for kanamycin resistant seedlings. The literature is lacking in this regard and we found that published protocols for screening of in vitro regenerated plants was not suitable for screening hundreds of seedlings. In the end, we used leaf painting with 2% kanamycin at the 1 leaf stage yielded reproducible yellow spots that can be used to differentiate between transgenic and non-transgenic seedlings. Using pistil drip transformation we have obtained 2,000-3,000 seed potentially carrying an AtP80-GFP, SnRK1.1 or IPK1 transgene. Objective 3. Profiling of higher InsPs in cotton and related plants. In 2015 we published a paper detailing that cotton, and some close relatives, contain elevated Ins(1,2,4,5,6)P5 levels. Intriguingly, cotton (G. hirsutum, raimondii, and barbadense), multiple types of Hibiscus, but not other Malvaceae, contain much higher levels of Ins(1,2,4,5,6)P5 as compared to InsP6. In fact, within these plants, InsP5 is the predominant InsP and this ratio is unusual in that most plants surveyed to date store more phosphate in InsP6 than InsP5. We have also used radiolabeling with 3H-inositol and HPLC to detect higher InsPs, where we can detect three separate peaks of InsP5. Both our mass assay and our radiolabeling techniques allow us to gather quantitative data on InsP levels. It is worth noting that while some lower InsPs may be more abundant than InsP6 in cotton vegetative tissues, we verified that InsP5 was the most abundant labelled InsP in cotton seedling shoot, root and leaves (Phillippy et al. 2015). Each of these tissues also contained higher Ins(1,2,4,5,6)P5 as compared to InsP6.
Publications
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Progress 09/01/14 to 08/31/15
Outputs
Target Audience:Our target audience is cotton growers and other life scientists interested in the mechanisms of senescence and energy-sensing in plants. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?One graduate student, one post-bac researcher, and one undergraduate researcher have been engaged in this project. Both the graduate student and post-bac researcher attended a national meeting and presented data. The technician working on this project also attended the same meeting and gave an invited talk. The graduate student also organized the Gordon Research Conference on Plant Lipids in 2015. A minority undergraduate researcher was engaged on this project and presented his work at our annual summer research symposium on campus. This student is a Biochemistry major who plans to attend law school and exposure to this project has helped cement an interest in a patent law career. How have the results been disseminated to communities of interest?Two posters and one invited talk were given at an international meeting (ASBMB 2015 in Boston). Two lectures on this work were given at universities. One research article and one review were published. What do you plan to do during the next reporting period to accomplish the goals?Obj. 1 is complete. For Obj. 2. (Manipulation of P80 and InsP network in cotton) we will continue to refine data on P80 suppression and begin knocking down other members of the P80 complex. Our hypothesis is that suppression of UBP3 and SnRK1.1 will also lead to changes in host defenses to geminiviruses. Further, we will investigate whether loss or gain-of-function in P80 in Arabidopis leads to changes in the response to a geminivirus (cabbage leaf curl virus : CaLC virus). This is an important model system in which to verify our cotton results and to test whether P80 can be used as an anti-viral resistance gene. In addition, we are transforming cotton with tagged version of the Arabidopsis P80 gene to produce material to test for resistance in cotton. We have also established a collaboration to test whether other viruses are more successful in Arabidopsis p80 mutants, or alternatively, whether the phenomena we are observing is specific to DNA viruses. A new area of work will being studies to identify the most-likely target of P80 deubiquination. We suspect this target is what alters viral resistance. We have good candidates for this target, and will work over the next year to confirm whether these candidates are linked to the new viral defense pathway we have identified in this project. For Obj 2: Manipulation of the InsP network via stable transformation: During the next year we expect to have IPK1 overexpressing cotton plants that can serve as material for InsP profiling work. The questions we will address are: Does overexpression of IPK1 lead to changes in InsP6 levels as expected? Further, does this alter growth or physiology of cotton? Does it alter JA signaling, as expected? In addition, we expect to screen for P80 and SnRK1.1 overexpression in cotton, and perform beginning analyses on these plants. Obj. 3. Profiling of higher InsPs in cotton and related plants. Our finding linking energy status to levels of higher InsPs cotton will be followed up on. Specifically, we will initiate experiments to carefully correlate both circadian rhythm and energy status with InsP levels. We are especially interested in whether an extended night period can further elevate InsP levels. On another project we are investigating Arabidopsis InsP levels, and will work over the next year to compare Arabidopsis and cotton. We suspect that parallel changes are occurring in both plants. Lastly, we will profile all transgenic and VIGS plants created in this project, to explore the connections between viral resistance and InsP levels.
Impacts
What was accomplished under these goals?
Obj. 1: Characterization of genes Involved in the P80 senescence regulation complex. We have identified and cloned genes for SnRK1.1, UBP3, IPK1 and various P80 genes. Each of these genes has been used to produce VIGS constructs for use in cotton. In general, the work in this objective is complete, and we have all required genes in hand for manipulating the complex in cotton. Obj. 2. Manipulation of P80 in cotton via VIGS: We have completed four separate VIGS experiments with appropriate controls that indicate that suppression of the cotton P80 genes results in growth and developmental phenotypes. It is interesting and important to note this phenotype is suggestive of increased viral movement throughout the plant. PCR experiments show that indeed, suppression of P80 gene expression correlates with increased geminivirus movement throughout treated plants. Our tentative conclusion is that P80 functions as a host defense gene whose expression helps to limit geminivirus spread. The implications of this finding could be very large in that the P80 gene may be a useful target to manipulate viral resistance in plants. We note that geminiviruses are important pathogens, especially when global agriculture is considered. We are currently refining data on this for publication, and pursuing VIGS suppression of the other members of the complex. We are pursuing collaborations with virologists who specialize in geminiviruses and describe our plans for future work to follow up on this important finding in the relevant section. Manipulation of the InsP network via stable transformation: We have completed a large pistil drip experiment that should yield transgenic cotton overexpressing the IPK1 gene. We are currently screening transformants and so do not yet have reliable data on transformation efficiencies with this approach. We have also undertaken large pistil drip transformation to overexpress the P80 gene and the SnRK1.1 gene in Deltapine 60 cotton. Obj. 3. Profiling of higher InsPs in cotton and related plants. We completed profiling of higher InsPs in leaves of cotton and cotton relatives, and published this data in 2015. We also initiated profiling work on different developmental stages of cotton, to discern whether cotton-specific elevation of certain InsPs was present in all tissues, or is specific for certain stages/ tissues. Our preliminary data indicate that one key regulator of InsP levels in cotton is the energy status of the tissue. In cotton seedlings, when we limit light, or measure InsPs at the end of the night period (both are low energy conditions), we see elevations in higher InsPs. In contrast, when ample light is present or measurements are made at the mid-point of the day, levels of higher InsPs are lower. This is a key finding that links energy status to levels of higher InsPs. Further, it suggests that our efforts to manipulate InsP levels may lead to effective routes for altering plant biomass and growth rates.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2015
Citation:
Williams SP, Gillaspy GE, Perera IY. Biosynthesis and possible functions of inositol pyrophosphates in plants.Front Plant Sci. 2015 Feb 12;6:67.
Phillippy, BQ, Perera, IY, Donahue, JL, and Gillaspy, GE. Certain Malvaceae Plants Have a Unique Accumulation of myo-Inositol 1,2,4,5,6-Pentakisphosphate. Plants 2015, 4, 267-283; doi:10.3390/plants4020267
- Type:
Journal Articles
Status:
Published
Year Published:
2015
Citation:
Phillippy, BQ, Perera, IY, Donahue, JL and Gillaspy, GE. Plants 2015, 4, 267-283; doi:10.3390/plants4020267
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Progress 09/01/13 to 08/31/14
Outputs
Target Audience: The target audiences are the life sciences and agricultural research communities. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? Two PhD students and one undergraduate are receiving training. One PhD student attended an international meeting and gave a presentation. Two seminars were given at academic institutions (NC State University and the IGC in Lisbon, Portugal). The PI attended a symposium on systems biology and agriculture, and one student, the PI and the lab technician attended a lab retreat to meet with the NC State collaborators. The PI engaged in sabbatical research both in Lisbon, Portugal and at NC State University. How have the results been disseminated to communities of interest? Three scientific publications have been published. Over 300 elementary school students were engaged in a plant DNA extraction in a University-sponsored research day. What do you plan to do during the next reporting period to accomplish the goals? During the next year we will investigate the mechanism by which the P80 gene dramatically alters cotton growth and development. This will lead to a better understanding of how to manipulate cotton senescence. We will also alter the expression of other genes in cotton plants, both transiently, and by making stable, transgenic cotton plants. The purpose of this work is to identify alternate targets for modulating cotton senescence. Lastly, we will examine levels of InsPs in transgenic cotton, to discern whether engineered changes in InsPs are likely to result in desired traits, such as hardiness and/or stress tolerance.
Impacts
What was accomplished under these goals?
New genes from cotton were identified and cloned to address our first objective. These new genes are potential regulators of energy sensing and senescence in plants. These types of genes are important because their manipulation may lead to a new way to regulate senescence in cotton. Every year farmers use chemicals to induce early senescence in cotton, which is needed to facilitate harvesting of cotton. If the new genes can be used to regulate senescence in cotton, then farmers could save time and money, and less chemicals would be used, resulting in less environmental pollutants. To address Objective 2, we have developed a protocol that allows us to silence the identified genes in cotton plants. Our results indicate that silencing these genes dramtically alters plant development. This is important because it lays groundwork for future manipulation of cotton senescence. To understand which molecules inside of cotton may be important for regulating growth and stress tolerance pathways, we have also developed methods to detect and manipulate specific signaling molecules called inositol phosphates (InsPs). Our work this year has shown that cotton and other related plants have a unique composition of InsPs. We are working towards altering this composition to test whether changes in InsP molecules will allow other plants to be as hardy, or stress tolerant, as cotton.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Williams SP, Rangarajan P, Donahue JL, Hess JE, Gillaspy GE. (2014) Regulation of Sucrose non-Fermenting Related Kinase 1 genes in Arabidopsis thaliana. Front Plant Sci. J0;5:324.
- Type:
Journal Articles
Status:
Awaiting Publication
Year Published:
2014
Citation:
Nourbakhsh, A., Collakova, E. and Gillaspy, G. Characterization of the Inositol Monophosphatase Gene Family in Arabidopsis. Front Plant Sci.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Desai M, Rangarajan P, Donahue JL, Williams SP, Land ES, Mandal MK, Phillippy BQ, Perera IY, Raboy V, Gillaspy GE. (2014) Two inositol hexakisphosphate kinases drive inositol pyrophosphate synthesis in plants. Plant J. 80(4):642-53.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2014
Citation:
Williams, S.P., Gillaspy, G.E., Perera, I. The role of inositol pyrophosphate signaling molecules in plants. Front Plant Sci.
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