The Safflower bHLH Transcription Factor CtbHLH41 Negatively Regulates SA-induced Leaf Senescence Through Interaction with CtCP1

Abstract BackgroundSalicylic acid (SA) plays an important role in regulating leaf senescence. However, the molecular mechanism of leaf senescence of safflower (Carthamus tinctorius) is still elusive. In this study we found that the bHLH transcription factor (TF) CtbHLH41 in Carthamus tinctorius significantly delayed leaf senescence and inhibited the expression of senescence-related genes.ResultsIn order to explore how CtbHLH41 promotes leaf senescence, we carried out yeast two-hybrid screening. In this study, by exploring the mechanism of CtbHLH41 regulating CtCP1, it was found that CtCP1 promoted the hydrolysis of CtbHLH41 protein, accelerated the transcriptional activities of salicylic acid-mediated senescence-related genes CtSAG12 and CtSAG29, chlorophyll degradation genes CtNYC1 and CtNYE1, and accelerated leaf senescence. We found a negative SA regulator CtANS1, which interacts with CtbHLH41 and regulates its stability, thereby inhibiting CtCP1-mediated leaf senescence.ConclusionsIn short, our results provide a new insight into the mechanism of CtbHLH41 actively regulating the senescence of safflower leaves induced by SA.

Download Full-text

Identification of the Novel Host Protein Interacting With the Structural Protein VP1 of Chinese Sacbrood Virus by Yeast Two-Hybrid Screening

Frontiers in Microbiology ◽

10.3389/fmicb.2019.02192 ◽

2019 ◽

Vol 10 ◽

Cited By ~ 2

Author(s):

Xiyan Zhang ◽

Dongliang Fei ◽

Li Sun ◽

Ming Li ◽

YueYu Ma ◽

...

Keyword(s):

Host Protein ◽

The Novel ◽

Structural Protein ◽

Yeast Two Hybrid ◽

Sacbrood Virus ◽

Novel Host ◽

Two Hybrid ◽

Hybrid Screening

Download Full-text

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

Molecular Biology of the Cell ◽

10.1091/mbc.e07-03-0278 ◽

2007 ◽

Vol 18 (11) ◽

pp. 4317-4326 ◽

Cited By ~ 39

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.

Download Full-text

The apple C2H2-type zinc finger transcription factor MdZAT10 positively regulates JA-induced leaf senescence by interacting with MdBT2

Horticulture Research ◽

10.1038/s41438-021-00593-0 ◽

2021 ◽

Vol 8 (1) ◽

Author(s):

Kuo Yang ◽

Jian-Ping An ◽

Chong-Yang Li ◽

Xue-Na Shen ◽

Ya-Jing Liu ◽

...

Keyword(s):

Mass Spectrometry ◽

Transcription Factor ◽

Liquid Chromatography ◽

Zinc Finger ◽

Leaf Senescence ◽

Transcriptional Activity ◽

Molecular Mechanisms ◽

Liquid Chromatography Mass Spectrometry ◽

Zinc Finger Transcription Factor ◽

Insight Into

AbstractJasmonic acid (JA) plays an important role in regulating leaf senescence. However, the molecular mechanisms of leaf senescence in apple (Malus domestica) remain elusive. In this study, we found that MdZAT10, a C2H2-type zinc finger transcription factor (TF) in apple, markedly accelerates leaf senescence and increases the expression of senescence-related genes. To explore how MdZAT10 promotes leaf senescence, we carried out liquid chromatography/mass spectrometry screening. We found that MdABI5 physically interacts with MdZAT10. MdABI5, an important positive regulator of leaf senescence, significantly accelerated leaf senescence in apple. MdZAT10 was found to enhance the transcriptional activity of MdABI5 for MdNYC1 and MdNYE1, thus accelerating leaf senescence. In addition, we found that MdZAT10 expression was induced by methyl jasmonate (MeJA), which accelerated JA-induced leaf senescence. We also found that the JA-responsive protein MdBT2 directly interacts with MdZAT10 and reduces its protein stability through ubiquitination and degradation, thereby delaying MdZAT10-mediated leaf senescence. Taken together, our results provide new insight into the mechanisms by which MdZAT10 positively regulates JA-induced leaf senescence in apple.

Download Full-text

The Catenin p120ctnInteracts with Kaiso, a Novel BTB/POZ Domain Zinc Finger Transcription Factor

Molecular and Cellular Biology ◽

10.1128/mcb.19.5.3614 ◽

1999 ◽

Vol 19 (5) ◽

pp. 3614-3623 ◽

Cited By ~ 263

Author(s):

Juliet M. Daniel ◽

Albert B. Reynolds

Keyword(s):

Transcription Factor ◽

Monoclonal Antibodies ◽

Cell Adhesion ◽

Mammalian Cells ◽

Yeast Two Hybrid ◽

Amino Terminal ◽

Juxtamembrane Region ◽

Carboxy Terminal ◽

Two Hybrid ◽

E Cadherin

ABSTRACT p120 ctn is an Armadillo repeat domain protein with structural similarity to the cell adhesion cofactors β-catenin and plakoglobin. All three proteins interact directly with the cytoplasmic domain of the transmembrane cell adhesion molecule E-cadherin; β-catenin and plakoglobin bind a carboxy-terminal region in a mutually exclusive manner, while p120 binds the juxtamembrane region. Unlike β-catenin and plakoglobin, p120 does not interact with α-catenin, the tumor suppressor adenomatous polyposis coli (APC), or the transcription factor Lef-1, suggesting that it has unique binding partners and plays a distinct role in the cadherin-catenin complex. Using p120 as bait, we conducted a yeast two-hybrid screen and identified a novel transcription factor which we named Kaiso. Kaiso’s deduced amino acid sequence revealed an amino-terminal BTB/POZ protein-protein interaction domain and three carboxy-terminal zinc fingers of the C2H2 DNA-binding type. Kaiso thus belongs to a rapidly growing family of POZ-ZF transcription factors that include the Drosophila developmental regulators Tramtrak and Bric à brac, and the human oncoproteins BCL-6 and PLZF, which are causally linked to non-Hodgkins’ lymphoma and acute promyelocytic leukemia, respectively. Monoclonal antibodies to Kaiso were generated and used to immunolocalize the protein and confirm the specificity of the p120-Kaiso interaction in mammalian cells. Kaiso specifically coprecipitated with a variety of p120-specific monoclonal antibodies but not with antibodies to α- or β-catenin, E-cadherin, or APC. Like other POZ-ZF proteins, Kaiso localized to the nucleus and was associated with specific nuclear dots. Yeast two-hybrid interaction assays mapped the binding domains to Arm repeats 1 to 7 of p120 and the carboxy-terminal 200 amino acids of Kaiso. In addition, Kaiso homodimerized via its POZ domain but it did not heterodimerize with BCL-6, which heterodimerizes with PLZF. The involvement of POZ-ZF proteins in development and cancer makes Kaiso an interesting candidate for a downstream effector of cadherin and/or p120 signaling.

Download Full-text

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

Biological Chemistry ◽

10.1515/bc.2008.149 ◽

2008 ◽

Vol 389 (10) ◽

Cited By ~ 1

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.

Download Full-text

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

Steroid Receptor Methods ◽

10.1385/1-59259-115-9:227 ◽

2003 ◽

pp. 227-248 ◽

Cited By ~ 3

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

Download Full-text

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

Circulation ◽

10.1161/circ.118.suppl_18.s_449 ◽

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.

Download Full-text