UCR

Center for Plant Cell Biology



REU 2011


REU Students and their Summer 2011 Research Programs

Undergraduate students were invited to apply to the Center for Plant Cell Biology (CEPCEB) to pursue individual research projects in the areas of cellular and molecular biology of plants and their pathogens. In 2011, the following twelve students were accepted to this ongoing 10-week residential summer program.  Please click on the following student links to see photos and read about their Summer 2010 research programs in CEPCEB laboratories.

An REU Poster Session was held in the Genomics lobby at the end of the ten-week program, where students discussed their projects.

REU Student
College/University
CEPCEB Faculty
Johnathon Blahut University of California, Riverside Judelson Lab
Emma Britain Caraway

Wellesley College, MA

Bailey-Serres Lab

Jenna Browning-Kamins

Amherst College, MA

Smith Lab

Apolonio I. Huerta

University of California, Riverside

Judelson Lab

Matt Kennedy

Tufts University, MA

Reddy Lab

Penelope Lindsay
New College of Florida
Walling Lab

Matthew Lefebvre

St. Olafs College, MN

Chen Lab

Matthew Martin

Gustavus Adolphus College, MN

Ma Lab

Daniel McNelis

New York University

Smith Lab

Maxine Nanthavong

Riverside Community College, CA

Roper Lab

Sarah Rommelfanger

Southwestern College, KS

Jin Lab

Patrick Salveson

Riverside Community College, CA

Larive Lab

 

JOHNATHON BLAHUT
University of California, Riverside
Johnathan Blahut

Phytophthora infestans is an oomycete pathogen responsible for the tomato and potato late blight disease. Its economic significance has prompted research on its genetics and Dr. Judelson’s lab at UC Riverside is at the forefront of this research. The lab studies transcription factors expressed during the organism’s asexual reproductive cycle with an ultimate goal of inhibiting this reproduction. Previously the lab found a transcription factor called Myb2R1 to be of particular interest since it may be involved in inducing the stage of asexual reproduction called sporulation. Their experiments found that transformants with Myb2R1 overexpression were unstable and frequently reverted to wild type sporulation levels. This suggests that the transformed plasmid was lost and this summer my research project will help address this instability. The study will apply techniques such as Southern blotting and qRT-PCR to track the overexpression plasmid and the expression of Myb2R1, respectively. A second study will investigate how Myb2R1 regulates sporulation during infection. Here sporulation patterns will be observed in tomato leaves infected with wild type and overexpressing strains. Then qRT-PCR will be performed to monitor the overexpression status of Myb2R1. In summary, the study of transcription factors in P. infestans may yield valuable information on how to combat the late blight disease.

 

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EMMA BRITAIN CARAWAY
Wellesley College, MA
Emma Britain Caraway

Little is known about the genetic and molecular basis of flood tolerance in Zea mays L. spp mays (maize). All maize varieties commercially grown in the United States are susceptible to complete submergence from flooding. As global climate change, farmers are at risk of losing entire fields due to uncontrolled floods. Work with rice and Arabidopsis has shown that a subgroup of the large family of transcription factors called Ethylene Response Factors (ERFs) are important mediators in flooding and hypoxic conditions. Interestingly, protein half-life of some of these ERFs appears to be regulated by oxygen availability in Arabidopsis. Maize also possesses related ERF genes and my project is to examine the expression of one of these, Sub1A Like 1 (Sbl1), under submergence and in a low oxygen environment. I will also be investigating whether or not SBL1 protein accumulation is regulated in a similar manner as related Arabidopsis proteins. We hypothesize that SBL1 will be turned over under low oxygen conditions, possibly contributing to maize's intolerance to flooding. This may provide important observations for understanding and determining ways to improve submergence tolerance in maize.

 

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JENNA BROWNING-KAMINS
Amherst College, MA
Jenna Browning-Kamins

Alternate bearing or biennial bearing is a poorly understood process that occurs in most fruit and nut bearing tree crops, including the avocado (Persea americana). Experimental studies indicate that the fruit inhibits the growth and development of the shoot. A reduction in growth and flowering by the fruit ultimately effects the production of the tree. Further, climatic events that result in the abscission of flowers and fruits will synchronize most of the shoots in fruit/nut tree crops in large regions. The synchronization of trees in orchards over a large growing region creates annual fluctuation in fruit/nut markets, resulting in monetary strain for growers. My research project is focused on identifying genes that inhibit shoot growth in response to fruit load in avocado. The Ancestral Angiosperm Genome Project has recently sequenced cDNAs from shoots of avocado. I will select avocado genes that are homologous to key hormone signaling and dormant responsive genes in other plants and design gene specific primers. Next, apical buds from shoots with and without fruit will be collected at Farm ACW. After isolating mRNA from tissue samples, I will examine expression levels of candidate hormone/dormancy genes to identify fruit load responsive genes. In addition, I will determine if the lack of shoot growth is due to a reduction in photosynthates at the apices of shoots bearing fruit. Taken together, the goal of my research project is to provide a framework that can be used to shed light on how fruits inhibit growth of the shoot in avocado.

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APOLONIO I. HUERTA
University of California, Riverside
Apolonio I. Huerta

Phytophthora infestans is a plant pathogen responsible for the devastating late blight disease of potatoes and tomatoes worldwide. Despite being often referred to as a fungus, P. infestans is actually a member of the Oomycete class of eukaryotes. P. infestans uses both sexual and asexual reproduction as a means by which to proliferate. Zoospores (biflagellar spores), formed by specialized structures called sporangia during asexual reproduction, serve as the main vector by which infection takes place. Further germination of these zoospores and the formation of an appressorium allow P. infestans to penetrate and infect the host plant. My project involves the study of a particular family of transcription factors named bZIP, which are found in all eukaryotes. bZIP transcription factors have two conserved domains which are key to their function: a basic DNA binding domain and a leucine-zipper domain responsible for dimerization. This family of transcription factors has been shown to regulate the expression of genes important for development, metabolism, and stress responses in eukaryotes such as fungi and plants. In order to understand the role that bZIP’s play in asexual reproduction, particularly infection, of P. infestans I will knockdown the expression of several bZIP genes through the use of RNA interference. Concurrently, I will determine bZIP expression profiles at several stages of P. infestans' asexual reproduction cycle, quantifying gene expression through the use of RT qPCR. By performing these experiments we will better understand the role of bZIP transcription factors in P. infestans asexual development, including infection.

 

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MATT KENNEDY
Tufts University, MA
Matt Kennedy

The shoot apical meristem is responsible for all above ground plant structures which originate from an undifferentiated pool of cells known as stem cells. The Reddy laboratory is currently researching the gene networks regulating this critical group of plant cells. The protein WUSCHEL(WUS) is a mobile transcription factor that regulates the stem cell population through both repression and activation. The mobility and differential regulation is likely modulated through protein-protein interactions. In my project, I will investigate the interactions between WUS and other proteins by using Bi-molecular Fluorescence Complementation (BiFC). This process uses a pair of engineered plasmids in which each plasmid translates one half of the fluorescent protein YFP fused to the protein of interest. When the two halves of YFP are brought together by the interacting pair of proteins, a fluorescent signal is produced. Over the summer, I will transform E. coli with these plasmids, use the bacterial machinery to amplify the genes, and isolate the DNA through extraction techniques. Afterward, the gene gun is used to shoot DNA coated gold particles into onion epidermal cells. If the bombarded onion cells fluoresce, then WUS interacts with the protein of interest in vivo. The information gained about these protein-protein interactions will lead to further understanding of WUS and regulation of the stem cell population.

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PENELOPE LINDSAY
New College of Florida
Penelope Lindsay

LAP-A, a protein found in Solanum lycopersicum (tomato), has been found to affect the late wound response signaling pathway downstream of jasmonic acid when wounded mechanically, as well as when consumed by the caterpillar Maduca sexta. When LAP-A is removed from the plant, it is found to drastically hinder the plant’s immune response, making it much more susceptible to herbivory. A related protein, LAP-N, is found constitutively in many plants in low levels, and due to its genetic similarity, it has been silenced and expressed when LAP-A is silenced. The focus of the project is to determine whether or not LAP-N works in conjunction with LAP-A in the late wound-response pathway. We will analyze transplastomic plants that express only LAP-A in the chloroplast. This way, a definitive analysis of the effect of LAP-A can be determined. First, LAP-A protein levels in the chloroplast line will be analyzed through 2D-PAGE protein analysis. The 2D-PAGE immunoblots will determine whether or not the LAP-A protein is actually being expressed in the plant. Second, seeds from the chloroplast lines will be grown and wounded at the three-leaf stage to determine the levels of the LapA, pin1, pin2, and LapN RNAs using qPCR. The RNA analysis will show whether or not the transplastomic lines restore wound signaling, with an increase in LapA mRNA, as well as other late response genes similar to the wild-type control levels indicating that it is restored.

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MATTHEW LEFEBVRE
St. Olafs College, MN
Matthew Lefebvre

Micro-RNAs (miRNA) play a pivotal role in regulation of gene expression in eukaryotes. The best characterized miRNA mediated expression pathway involves post-transcriptional degradation of mRNAs based on sequence homology with 21-24 nucleotide miRNAs. However, miRNAs can also regulate gene expression by other means, potentially including post-translational mechanisms involving protein stability. A candidate protein involved in post-translational gene regulation by miRNAs in arabidopsis has been identified in the Chen laboratories. My work involves characterizing protein-protein interactions that the candidate protein is involved in so as to better understand the miRNA mediated post-translational pathway. To initially identify protein-protein interactions a yeast two hybrid screen will be employed with the candidate protein acting as the “bait” and an Arabidopsis c-DNA library acting as a high throughput screen of all other potential “prey” proteins in the Arabidopsis genome. Potential interacting proteins will be further characterized by fluorescent live-cell imaging (Bif-C) and co-immunoprecipitation experiments.

 

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MATTHEW MARTIN
Gustavus Adolphus College, MN
Matthew Martin

Phytophthora sojae is an oomycete plant pathogen responsible for causing stem and root rot in soybeans and a $1- 2 billion annual cost worldwide (Tyler 2007; Wang et al. 2011). Oomycetes are resistant to fungicides because, though they are fungus-like organisms, they belong to the Kingdom Chromista, and are most closely related to photosynthetic eukaryotic microorganisms such as brown algae and diatoms (Lamour et al. 2007; Hein et al. 2009). To date, no applicable “oomycete-icide” is available, making these pathogens extremely relevant and important to research. Preliminary research has revealed that oomycetes produce several hundreds of effector proteins, which likely function to promote infection of plant tissue (Kamoun 2001; Tyler 2007). The aim of my research project is to elucidate the virulence functions of two P. sojae effectors. To accomplish this goal, I will conduct two experiments: 1) Introduce both proteins into a bacterial pathogen, Pseudomonas syringae, and observe whether the transformed strains can better infect soybeans. 2) Overexpress effector proteins in soybean roots to see whether the transgenic roots are more susceptible to P. sojae infection. The outcome of my research will advance our understanding of P. sojae pathogenicity, which will help develop novel control strategies against the persistently devastating oomycete diseases.

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DANIEL MCNELIS
New York University
Daniel McNelis

In plants, aboveground organs and shoots derive from the shoot apical meristem (SAM), a site of continuous growth and development. This requires a balance of stem cell perpetuation and organogenesis, called meristem maintenance. Additionally, the plant hormones auxin and cytokinin play crucial roles in regulating meristem homeostasis. Research indicates specific KNOTTED1-LIKE HOMEOBOX (KNOX) and BELL1-LIKE HOMEOBOX (BLH) transcription factors form heterodimers to regulate this process. Our lab has shown two related Arabidopsis BLH genes, PENNYWISE (PNY) and POUNDFOOLISH (PNF), preserve the integrity of the central zone (CZ), an SAM region harboring stem cells. My goal is to elucidate how PNY and PNF accomplish meristem maintenance. My first objective is to investigate a CZ marker gene, CLAVATA3 (CLV3), that may be regulated by PNY and PNF. In pny pnf double mutants, the expression pattern of CLV3, whose 3’ enhancer contains predicted KNOX-BLH binding sites, is expanded throughout the SAM. By performing GUS staining I will test if these binding sites are required for the native expression patterns of CLV3. Next I will determine if PNY/PNF regulate auxin and cytokinin signaling in the SAM. Using mRNA in situ hybridization, I will compare expression patterns of known cytokinin and auxin signaling genes in wild-type and pny pnf plants. Finally, I will conduct a yeast 2-hybrid to determine whether PNY/PNF directly bind to known transcription factors that mediate auxin response in the SAM. My results this summer will hopefully reveal more about the function of PNY and PNF in meristem maintenance.p>

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MAXINE NANTHAVONG
Riverside Community College, CA
Maxine Nanthavong

Pantoea stewartii subsp. stewartii is a gram-negative bacterium responsible for Stewart’s wilt of sweet corn. The bacteria preferentially colonizes the xylem and causes severe wilting in young seedlings. Biofilm formation plays a critical role in the onset of Stewart’s wilt because it blocks water flow in the xylem, ultimately leading to plant wilting and death. The goal of this project is to determine proteins responsible for the initial cell-surface attachment of the bacterium, a critical step in successful biofilm formation. The mechanisms of how P. stewartii attaches to surfaces remains unknown. Evidence shows that OxyR, a transcriptional factor involved in the oxidative stress response, is also involved in biofilm formation. An OxyR knockout mutant (ΔoxyR) exhibits a hyper-attaching phenotype suggesting that OxyR may be negatively regulating one or more surface adhesins. We will take advantage of the increased attachment phenotype of the ΔoxyR mutant (Roper, 2011) and create a random mutant bank in the ΔoxyR genetic background using random transposon mutagenesis. Following this, we will screen for mutants that exhibit a decrease in cell-surface attachment as compared to ΔoxyR using a Crystal Violet-based attachment assay (O‘Toole & Kolter, 1998). Transposon mutants that are compromised in cell-surface attachment will be isolated and the site of transposon insertion will be identified to determine which genes are responsible for cell-surface attachment. Identifying proteins involved in attachment could serve useful in designing target specific chemicals that could render xylem-dwelling bacteria inefficient in causing plant diseases like Stewart’s wilt.

 

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SARAH ROMMELFANGER
Southwestern College, KS
Sarah Rommelfanger

Central to control of gene expression in almost all eukaryotes are a variety of post-transcriptional modification factors, including micro-RNAs (miRNA), and small-interfering RNAs (siRNAs). These specialized double-stranded RNA molecules are the main components of a process called RNAi, or RNA interference, whose key role is to regulate steps in mRNA maturation. RNAi-regulated gene silencing evolved as an antiviral defense mechanism in plants and animals. Though much work has been done determining the role of RNAi in plant-bacteria interactions, much less is known about RNAi function in plant-fungus relationships. Dr. Hailing Jin’s lab is focusing on elucidating the roles of RNAi during plant-bacteria and plant-fungus interactions, measuring its effects on pathogenicity and host plant resistance, with an ultimate goal of utilizing results to optimize plant immunity. The discovery of a bacteria-induced siRNA in the Jin lab provided the first example of the regulatory role siRNAs play in plant immunity. Botrytis cinerea, commonly known as grey mold, is a necrotrophic fungus prominent in a variety of produce. My role in the Jin lab will be to assist in defining the important role of RNAi in the plant-fungus interactions of B. cinerea with Arabidopsis thaliana and tomato. Techniques used will include plant and fungal mutants in infection assays, and fungal biomass quantification of infected plant tissue by real-time PCR. These methods will allow disease assessment of Botrytis RNAi mutants and susceptibility of Arabidopsis RNAi mutants.

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PATRICK SALVESON
Riverside Community College, CA
Patrick Salveson

The aim of my project is to develop a unified extraction method for plant metabolomics measurements by nuclear magnetic resonance (NMR) and liquid chromatography – mass spectrometry (LC-MS). Plant metabolomics is a growing field of research and is of increasing interest to plant biologists due to the insights it provides into the metabolome, genome, and transcriptome. Because of their sessile nature, plants are believed to produce approximately 200,000 different metabolites representing a variety of chemical classes. Due to the chemical diversity of the plant metabolome, metabolite extraction methods must be optimized to maximize the information obtained from a metabolomics experiment. Several sample preparation steps are required, including grinding the plant material, extraction of the small molecule metabolites, removal of interfering compounds, concentration of the extract to increase sensitivity, and finally dissolution in a solvent system compatible with the analytical platform. I am growing Arabidopsis and Maize to use in the evaluation of extraction procedures. My experiments will test three solvent compositions, two different mixing methods, and the effect of temperature on metabolite extraction. The efficiency of each extraction method will be judged by results obtained from LC-MS and NMR. This project will evaluate the amount and type of metabolites extracted as well as the effects of each extraction method on the quality of data collected. A unified extraction method would allow for cross platform analysis using a single extract of a plant sample, decreasing the analytical variance while increasing the amount of information gained from each sample.


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University of California, Riverside
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Riverside, CA 92521
Tel: (951) 827-1012

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Center for Plant Cell Biology
Botany & Plant Sciences Department
2150 Batchelor Hall

Tel: (951) 827-7177
Fax: (951) 827-5155
E-mail: genomics@ucr.edu

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