• BMB Reports
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• 제47권11호
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• pp.643-648
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• 2014

Stanniocalcin (STC), a glycoprotein hormone originally discovered in fish, has been implicated in calcium and phosphate homeostasis. While fishes and mammals possess two STC homologs (STC1 and STC2), the physiological roles of STC2 are largely unknown compared with those of STC1. In this study, we identified Ran-binding protein M (RanBPM) as a novel binding partner of STC2 using yeast two-hybrid screening. The interaction between STC2 and RanBPM was confirmed in mammalian cells by immunoprecipitation. STC2 enhanced the RanBPM-mediated transactivation of liganded androgen receptor (AR), but not thyroid receptor ${\beta}$, glucocorticoid receptor, or estrogen receptor ${\beta}$. We also found that AR interacted with RanBPM in both the absence and presence of testosterone (T). Furthermore, we discovered that STC2 recruits RanBPM/AR complex in T-dependent manner. Taken together, our findings suggest that STC2 is a novel RanBPM-interacting protein that promotes AR transactivation.

• Biomolecules & Therapeutics
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• 제17권1호
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• pp.12-16
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• 2009

One of the WHSC1/MMSET/NSD2 variant RE-IIBP is a histone H3-K27 methyltransferase with transcriptional repression activity. Overexpression of RE-IIBP in various types of leukemia suggests it's role in leukemogenesis. Here we identify two proteins, KRAB zinc finger protein ZNF 350 and enolase-1 as RE-IIBP interacting proteins by yeast two-hybrid screening and confirmed direct interaction in vivo and in vitro. Both proteins have been known for their role in transcriptional repression. Reporter assays using transient transfection demonstrated that both ZNF 350 and enolase-1 proteins synergistically repressed transcription with RE-IIBP, respectively. These results indicate both proteins have roles in RE-IIBP mediated transcriptional repression by involving co-repressor complex.

• 미생물학회지
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• 제50권2호
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• pp.95-100
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• 2014

Hrq1은 곰팡이 유전체에서 생물정보분석에 의해 발견된 새로운 RecQ helicase이다. 이 단백질은 인간의 RECQL4와 가장 상동성이 높으며 최근의 유전학적 생화학적 연구를 통해서 유전체 안정성을 유지하는데 어떤 역할을 할 것으로 예상되었다. 본 연구에서는 RECQL4와 상호작용하는 것으로 알려진 인간 유전자들과 상동성이 있는 효모 유전자들이 Hrq1과 상호작용하는지를 yeast two-hybrid assay를 이용하여 조사하였다. 총 11개의 유전자를 조사한 결과, nucleotide excision repair (NER) 인자 중의 하나인 Rad14이 Hrq1과 상호작용하는 것을 발견하였다. 또한 정제한 단백질을 이용한 pull-down assay로 Hrq1과 Rad14 사이의 직접적인 상호작용을 확인하였다. Hrq1과 Rad14 사이의 yeast two-hybrid 상호작용은 4-nitroquinoline-1-oxide에 의한 DNA 손상으로 더욱 증가하였으며, 이러한 상호작용의 증가는 또 다른 NER 인자인 Rad4에 의존적이었다. 이러한 결과들은 Hrq1이 Rad14과의 상호작용을 통하여 NER 과정에 어떤 역할을 할 가능성을 제시하고 있다.

• Journal of Microbiology and Biotechnology
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• 제22권3호
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• pp.378-384
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• 2012

In aquatic invertebrates, particularly marine gastropods, organotin compounds induce irreversible sexual abnormality in females, which is termed imposex, at very low concentrations. Organotin compounds are agonists for nuclear receptors such as RXRs and $PPAR{\gamma}$. However, the imposex phenomenon has not been reported to act as an antagonist on estrogen receptors in other species, including vertebrates and invertebrates. In order to gain insights into the antagonistic activity of organotin compounds on estrogen receptors (ERs), we examined the inhibitive effect of these compounds on estradiol-dependent ${\beta}$-galactosidase activity using the yeast two-hybrid detection system consisting of a combination of the human estrogen receptor ($hER{\beta}$) ligand-binding domain and the co-activator steroid receptor co-activator-1 (SRC1). Tributyltin-hydroxide (TBT-OH) and triphenyltin-chlorine (TPT-Cl) exhibited an inhibitive effect on $E_2$-dependent transcriptional activity, similar to antagonistic chemicals such as 4-hydroxytamoxifen (OHT) or ICI 182,780, at a very low concentration of $10^{-14}$ M TBT or $10^{-10}$ M TPT, respectively. The yeast growth and transcriptional activity with transcriptional factor GAL4 did not exhibit any effect at the tested concentration of TBT or TPT. Moreover, the yeast two-hybrid system using the interaction between p53 and the T antigen of SV40 large did not describe any effect at the tested concentration of OHT or ICI 182,780. However, the interaction between p53 and T antigen was inhibited at a TBT or TPT concentration of $10^{-9}$ M, respectively. These results indicate that TBT and TPT act as inhibitors of ER-dependent reporter gene transcriptional activation and of the interaction between $hER{\beta}$ LBD and the co-activator SRC1 in the yeast two-hybrid system. Consequently, our data could partly explain the occurrence of organotin compound-induced imposex on the endocrine system of mammals, including humans.

• 미생물학회지
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• 제53권1호
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• pp.49-54
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• 2017

Sus1 / ENY2는 진핵생물에 잘 보존된 작은 단백질로 염색사 리모델링과 mRNA 생성과정에 관여한다. Sus1 단백질은 전사 보조활성자인 SAGA 복합체와 핵공과 연관된 TREX2 복합체 등, 핵에 존재하는 2개 복합체의 구성 요소이다. 분열효모 Schizosaccharomyces pombe의 유전체 데이터에서 Sus1의 이종상동체를 찾아 기능을 분석하였다. 4분체 분석 결과 이 유전자는 생장에 필수적이 아니었으나, Sus1 유전자를 결실시키면 낮은 온도에서 생장이 느렸으며, $poly(A)^+$ RNA도 핵 안에 약간 축적되는 표현형을 보였다. 또한 Sus1-GFP 단백질은 주로 핵에 존재하였다. Yeast two-hybrid 분석과 공동면역침전 실험에서 Sus1 단백질은 TREX-2 복합체의 또 다른 구성인자인 Sac3와 상호작용을 하였다. 이와 같은 결과들은 S. pombe의 Sus1 단백질도 역시 TREX-2 복합체의 구성인자로 mRNA 방출에 관여하고 있음을 시사한다.

• BMB Reports
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• 제34권1호
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• pp.85-89
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• 2001

Both topoisomerase $II{\alpha}$ and $II{\beta}$ east as phosphoproteins in the cells. Recently it was reported that DNA topoisomerase $II{\alpha}$ associates with and is phosphorylated by the extracellular signal-regulated kinase 2 (ERK2). Also, ERK2 stimulates the activity of topoisomerase II by a phosphorylation-independent manner [Shapiro et al., (1999) Mol. Cell. Biol. 19, 3551-3560]. In this study, a yeast two-hybrid system was used to investigate the binding site between topoisomerase $II{\alpha}$ or $II{\beta}$ and ERK2. The two-hybrid test clearly showed that topoisomerase $II{\beta}$ residues 1099-1263, and topoisomerase $II{\alpha}$ residues 1078-1182, mediate the interaction with ERK2, and that the leucine zipper motifs of topoisomerase $II{\alpha}$ and $II{\beta}$ are not required for its physical binding to ERK2. Our results suggest that topoisomerase $II{\beta}$ residues 1099-1263, and topoisomerase $II{\alpha}$ residues 1078-1182, may be common binding sites for activator proteins.

• 미생물학회지
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• 제54권4호
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• pp.325-329
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• 2018

PCI 영역을 포함하고 있는 Thp1/PCID2 단백질은 mRNA 전사와 방출을 연결하는, 진화적으로 보존된 TREX-2 복합체의 구성요소이다. 분열효모인 Schizosaccharomyces pombe에서 pci2 (SPBC1105.07c) 유전자는 TREX-2 복합체의 구성요소인 Thp1 (출아효모)/PCID2 (사람)의 분열효모 이종상동체로 추정되는 PCI 영역을 갖는 단백질을 암호화하고 있다. pci2 발현을 억제하면 생장과 mRNA 방출이 모두 억제되었다. 그리고 pci2 유전자의 과발현 또한 생장을 늦추고 $poly(A)^+$ RNA를 핵 안에 약간 축적되게 하였다. 뿐만 아니라 Yeast two-hybrid와 공동침전(Co-immunoprecipitation) 분석 실험에서 Pci2는 TREX-2 복합체의 다른 구성요소인 Sac3, Dss1 단백질들과 물리적으로 상호작용하였다. 이와 같은 관찰들은 S. pombe의 Pci2 단백질도 TREX-2 복합체의 구성요소로서 mRNA 방출에 관여함을 의미한다.

• 대한의생명과학회지
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• 제13권2호
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• pp.127-133
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• 2007

Heterotrimeric GTP binding proteins (G proteins) mediate signal generated by neurotransmitter and hormones. Among all G proteins, Go is the most abundant in brain but its role in brain is not clearly understood. To determine the function of the alpha subunit of Go (Go$\alpha$), we search for the interacting partner of Go$\alpha$ in brain using yeast two-hybrid system. A resistant to inhibitor of cholinesterase (Ric-8B) was identified as a Go$\alpha$ interacting protein. We confirmed interaction between Go$\alpha$ and Ric-8b employing in vitro affinity binding assay and showed that the Ric-8b increased the function of Go$\alpha$. Our findings indicate that Ric-8b is possible guanine nucleotide exchange factor for Go$\alpha$.

• BMB Reports
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• 제37권1호
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• pp.45-52
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• 2004

The goal of interaction proteomics that studies the protein-protein interactions of all expressed proteins is to understand biological processes that are strictly regulated by these interactions. The availability of entire genome sequences of many organisms and high-throughput analysis tools has led scientists to study the entire proteome (Pandey and Mann, 2000). There are various high-throughput methods for detecting protein interactions such as yeast two-hybrid approach and mass spectrometry to produce vast amounts of data that can be utilized to decipher protein functions in complicated biological networks. In this review, we discuss recent developments in analytical methods for large-scale protein interactions and the future direction of interaction proteomics.

• 대한의생명과학회지
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• 제13권4호
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• pp.305-311
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• 2007

cAMP-dependent protein kinase A (PKA) is the best-characterized protein kinases and has served as a model of the structure and regulation of cAMP-binding protein as well as of protein kinases. To determine the function of PKA in development, we employed the yeast two-hybrid system to screen for catalytic subunit of PKA $(C\alpha)$ interacting partners in a cDNA library from mouse embryo. A Testis-brain RNA-binding protein (TB-RBP), specifically bound to $C\alpha$. This interaction was verified by several biochemical analysis. Our findings indicate that $C\alpha$ can modulate nucleic acid binding proteins of TB-RBP and provide insights into the diverse role of PKA.