scholarly journals Host interactors of effector proteins of the lettuce downy mildew Bremia lactucae obtained by yeast two-hybrid screening

PLoS ONE ◽  
2020 ◽  
Vol 15 (5) ◽  
pp. e0226540 ◽  
Author(s):  
Alexandra J. E. Pelgrom ◽  
Claudia-Nicole Meisrimler ◽  
Joyce Elberse ◽  
Thijs Koorman ◽  
Mike Boxem ◽  
...  
Keyword(s):  
Downy Mildew ◽  
Effector Proteins ◽  
Bremia Lactucae ◽  
Yeast Two Hybrid ◽  
Lettuce Downy Mildew ◽  
Two Hybrid ◽  
Hybrid Screening

scholarly journals Host targets of effectors of the lettuce downy mildew Bremia lactucae from cDNA-based yeast two-hybrid screening

2019 ◽  
Author(s):  
Alexandra J.E. Pelgrom ◽  
Claudia-Nicole Meisrimler ◽  
Joyce Elberse ◽  
Thijs Koorman ◽  
Mike Boxem ◽  
...  
Keyword(s):  
Downy Mildew ◽  
Pathogenic Bacteria ◽  
Plant Pathogens ◽  
Molecular Mechanisms ◽  
Effector Proteins ◽  
Bremia Lactucae ◽  
Yeast Two Hybrid ◽  
Target Proteins ◽  
Successful Infection ◽  
Two Hybrid

AbstractPlant pathogenic bacteria, fungi and oomycetes secrete effector proteins to manipulate host cell processes to establish a successful infection. Over the last decade the genomes and transcriptomes of many agriculturally important plant pathogens have been sequenced and vast candidate effector repertoires were identified using bioinformatic analyses. Elucidating the contribution of individual effectors to pathogenicity is the next major hurdle. To advance our understanding of the molecular mechanisms underlying lettuce susceptibility to the downy mildew Bremia lactucae, we mapped a network of physical interactions between B. lactucae effectors and lettuce target proteins. Using a lettuce cDNA library-based yeast-two-hybrid system, 61 protein-protein interactions were identified, involving 21 B. lactucae effectors and 46 unique lettuce proteins. The top ten targets based on the number of independent colonies identified in the Y2H and two targets that belong to gene families involved in plant immunity, were further characterized. We determined the subcellular localization of the fluorescently tagged target proteins and their interacting effectors. Importantly, relocalization of effectors or targets to the nucleus was observed for four effector-target pairs upon their co-expression, supporting their interaction in planta.

scholarly journals JAZ8 Interacts With VirE3 Attenuating Agrobacterium Mediated Root Tumorigenesis

2021 ◽  
Vol 12 ◽  
Author(s):  
Shijuan Li ◽  
Bingliang Xu ◽  
Xiaolei Niu ◽  
Xiang Lu ◽  
Jianping Cheng ◽  
...  
Keyword(s):  
Host Plants ◽  
Effector Proteins ◽  
Bimolecular Fluorescence Complementation ◽  
Yeast Two Hybrid ◽  
Defense Signaling ◽  
Wide Range ◽  
Crown Gall Tumors ◽  
F Box Protein ◽  
Two Hybrid ◽  
Hybrid Screening

Agrobacterium tumefaciens can cause crown gall tumors by transferring both an oncogenic piece of DNA (T-DNA) and several effector proteins into a wide range of host plants. For the translocated effector VirE3 multiple functions have been reported. It acts as a transcription factor in the nucleus binding to the Arabidopsis thaliana pBrp TFIIB-like protein to activate the expression of VBF, an F-box protein involved in degradation of the VirE2 and VIP1 proteins, facilitating Agrobacterium-mediated transformation. Also VirE3 has been found at the plasma membrane, where it could interact with VirE2. Here, we identified AtJAZ8 in a yeast two-hybrid screening with VirE3 as a bait and confirmed the interaction by pull-down and bimolecular fluorescence complementation assays. We also found that the deletion of virE3 reduced Agrobacterium virulence in a root tumor assay. Overexpression of virE3 in Arabidopsis enhanced tumorigenesis, whereas overexpression of AtJAZ8 in Arabidopsis significantly decreased the numbers of tumors formed. Further experiments demonstrated that AtJAZ8 inhibited the activity of VirE3 as a plant transcriptional regulator, and overexpression of AtJAZ8 in Arabidopsis activated AtPR1 gene expression while it repressed the expression of AtPDF1.2. Conversely, overexpression of virE3 in Arabidopsis suppressed the expression of AtPR1 whereas activated the expression of AtPDF1.2. Our results proposed a novel mechanism of counter defense signaling pathways used by Agrobacterium, suggesting that VirE3 and JAZ8 may antagonistically modulate the salicylic acid/jasmonic acid (SA/JA)-mediated plant defense signaling response during Agrobacterium infection.

scholarly journals Two LIM Domain Proteins and UNC-96 Link UNC-97/PINCH to Myosin Thick Filaments in Caenorhabditis elegans Muscle

2007 ◽  
Vol 18 (11) ◽  
pp. 4317-4326 ◽  
Author(s):  
Hiroshi Qadota ◽  
Kristina B. Mercer ◽  
Rachel K. Miller ◽  
Kozo Kaibuchi ◽  
Guy M. Benian
Keyword(s):  
Caenorhabditis Elegans ◽  
Binding Assays ◽  
Lim Domain ◽  
Yeast Two Hybrid ◽  
Lim Domains ◽  
Thick Filaments ◽  
Vitro Binding ◽  
Two Hybrid ◽  
Hybrid Screening

By yeast two-hybrid screening, we found three novel interactors (UNC-95, LIM-8, and LIM-9) for UNC-97/PINCH in Caenorhabditis elegans. All three proteins contain LIM domains that are required for binding. Among the three interactors, LIM-8 and LIM-9 also bind to UNC-96, a component of sarcomeric M-lines. UNC-96 and LIM-8 also bind to the C-terminal portion of a myosin heavy chain (MHC), MHC A, which resides in the middle of thick filaments in the proximity of M-lines. All interactions identified by yeast two-hybrid assays were confirmed by in vitro binding assays using purified proteins. All three novel UNC-97 interactors are expressed in body wall muscle and by antibodies localize to M-lines. Either a decreased or an increased dosage of UNC-96 results in disorganization of thick filaments. Our previous studies showed that UNC-98, a C2H2 Zn finger protein, acts as a linkage between UNC-97, an integrin-associated protein, and MHC A in myosin thick filaments. In this study, we demonstrate another mechanism by which this linkage occurs: from UNC-97 through LIM-8 or LIM-9/UNC-96 to myosin.

Specific inhibition of transcriptional activity of the constitutive androstane receptor (CAR) by the splicing factor SF3a3

2008 ◽  
Vol 389 (10) ◽  
Author(s):  
Hye Jin Yun ◽  
Jungsun Kwon ◽  
Wongi Seol
Keyword(s):  
Transcriptional Activity ◽  
Constitutive Androstane Receptor ◽  
Splicing Factor ◽  
Yeast Two Hybrid ◽  
Interacting Protein ◽  
Functional Studies ◽  
Reporter Activity ◽  
Inhibitory Function ◽  
Two Hybrid ◽  
Hybrid Screening

Abstract The constitutive androstane receptor (CAR) is a member of the nuclear receptor superfamily and plays an important role in the degradation of xenobiotics in the liver. Using yeast two-hybrid screening, we identified SF3a3, a 60-kDa subunit of the splicing factor 3a complex, as a specific CAR-interacting protein. We further confirmed their interaction by both co-immunoprecipitation and GST pull-down assay. Functional studies showed that overexpression of SF3a3 inhibited the reporter activity driven by a promoter containing CAR binding sequences by up to 50%, whereas reduced expression of SF3a3 activated the same reporter activity by approximately three-fold. The inhibitory function of SF3a3 is independent of the presence of TCPOBOP, a CAR ligand. These data suggest that SF3a3 functions as a co-repressor of CAR transcriptional activity, in addition to its canonical function.

Yeast Two-Hybrid Screening for Proteins that Interact with Nuclear Hormone Receptors

2003 ◽  
pp. 227-248 ◽  
Author(s):  
Bertrand Le Douarin ◽  
David M. Heery ◽  
Claudine Gaudon ◽  
Elmar vom Baur ◽  
Régine Losson
Keyword(s):  
Hormone Receptors ◽  
Nuclear Hormone Receptors ◽  
Yeast Two Hybrid ◽  
Two Hybrid ◽  
Hybrid Screening

Abstract 3600: HspA12B Promotes Angiogenesis through Suppressing AKAP12 and Up-Regulating VEGF Pathway

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Rebecca J Steagall ◽  
Fang Hua ◽  
Mahesh Thirunazukarasu ◽  
Lijun Zhan ◽  
Chuanfu Li ◽  
...  
Keyword(s):  
Cancer Metastasis ◽  
Yeast Two Hybrid ◽  
Vegf Expression ◽  
Tubule Formation ◽  
Interacting Protein ◽  
Over Expression ◽  
Vegf Pathway ◽  
Two Hybrid ◽  
Hybrid Screening

We have previously shown that HspA12B, a member of HspA70 family subfamily 12, is a novel angiogenesis regulator that is preferentially expressed in endothelial cells (ECs) and required for angiogenesis in vitro . The mechanism by which HspA12B regulates angiogenesis, however, is unknown. In this study we identified AKAP12/SSeCKS as a HSPA12B-interacting protein through a yeast two-hybrid screening and confirmed the interaction by co-immunoprecipitation and co-localization. We observed that HspA12B negatively regulated the expression of AKAP12/SSeCKS, a cancer metastasis repressor that inhibits VEGF expression and angiogen-esis. In HUVEC, HspA12B knockdown increased AKAP12 levels, decreased VEGF by more than 75%, and down-regulated Akt and pAkt; whereas HspA12B over expression decreased AKAP12 and more than doubled VEGF levels. We further identified a 32-AA domain in AKAP12 that was capable of interacting with HspA12B. Overexpression of this 32-AA domain in HUVEC disrupted the HspA12B-AKAP12 interaction and decreased VEGF expression by more than 70%, suggesting the importance of HspA12B-AKAP12 interaction in regulating VEGF. We also observed that HspA12B expression was increased more than 2 folds in ECs by hypoxia or shearing stress, and induced in ischemic rat heart. Inhibition of HspA12B abolished hypoxia-induced tubule formation. Adeno-HspA12B promoted angiogenesis in DIVAA assay. We concluded that this is the first evidence that HspA12B promotes angiogenesis through regulating VEGF by way of suppressing AKAP12. Our finding is the first example of an EC-specific molecular chaperone acting as the regulator of angiogenesis.

Yeast Two-Hybrid Screening for Identification of in

2021 ◽  
pp. 95-110
Author(s):  
Hazel McLellan ◽  
Miles R. Armstrong ◽  
Paul R. J. Birch
Keyword(s):  
Yeast Two Hybrid ◽  
Two Hybrid ◽  
Hybrid Screening

scholarly journals Insensitivity to the Fungicide Fosetyl-Aluminum in California Isolates of the Lettuce Downy Mildew Pathogen, Bremia lactucae

Plant Disease ◽  
2004 ◽  
Vol 88 (5) ◽  
pp. 502-508 ◽  
Author(s):  
Sarah Brown ◽  
Steven T. Koike ◽  
Oswaldo E. Ochoa ◽  
Frank Laemmlen ◽  
Richard W. Michelmore
Keyword(s):  
Quantitative Analysis ◽  
Downy Mildew ◽  
Mating Type ◽  
Control Measures ◽  
Bremia Lactucae ◽  
Widespread Occurrence ◽  
Lettuce Downy Mildew ◽  
Normal Field ◽  
Growth Room ◽  
Disease Pressure

Lettuce downy mildew, caused by Bremia lactucae, is the most important foliar disease of lettuce in California. In recent years, there were apparent failures of fungicides containing fosetyl-aluminum (Aliette) to control downy mildew in commercial lettuce fields in California. Consequently, we characterized 134 isolates collected over 2 years from throughout the coastal growing areas of California for insensitivity to the fungicides fosetyl-aluminum and maneb, pathotype, and mating type. Tests using seedlings in controlled growth room conditions demonstrated the widespread occurrence of insensitivity to fosetyl-aluminum in California populations of B. lactucae. Fifty percent of the isolates assayed sporulated profusely in the presence of fosetyl-aluminum applied at rates twice the normal field dosage, and an additional 40% showed moderate sporulation at this rate. Fosetyl-aluminum-insensitive isolates were detected from all regions sampled. Insensitivity was also observed in multiple pathotypes. Insensitivity was not complete, however, because quantitative analysis of the number of lesions on older plants revealed that applications of fosetyl-aluminum could reduce the levels of disease by 50%. Therefore, while fosetyl-aluminum may have utility under low disease pressure in the field, other control measures are required to provide control under conditions favorable to the disease.

Yeast Two-Hybrid Screening for Proteins Interacting With the Anti-Apoptotic Protein A20

2004 ◽  
pp. 223-242 ◽  
Author(s):  
Karen Heyninck ◽  
Sofie Van Huffel ◽  
Marja Kreike ◽  
Rudi Beyaert
Keyword(s):  
Yeast Two Hybrid ◽  
Apoptotic Protein ◽  
Two Hybrid ◽  
Hybrid Screening
Sign in / Sign up

Export Citation Format

Share Document