• Biomolecules & Therapeutics
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• 제27권2호
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• pp.231-239
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• 2019

Suppressor of Variegation 3-9 Homolog 2 (SUV39H2) methylates the lysine 9 residue of histone H3 and induces heterochromatin formation, resulting in transcriptional repression or silencing of target genes. SUV39H1 and SUV39H2 have a role in embryonic development, and SUV39H1 was shown to suppress cell cycle progression associated with Rb. However, the function of human SUV39H2 has not been extensively studied. We observed that forced expression of SUV39H2 decreased cell proliferation by inducing $G_1$ cell cycle arrest. In addition, SUV39H2 was degraded through the ubiquitin-proteasomal pathway. Using yeast two-hybrid screening to address the degradation mechanism and function of SUV39H2, we identified translationally controlled tumor protein (TCTP) as an SUV39H2-interacting molecule. Mapping of the interacting regions indicated that the N-terminal 60 amino acids (aa) of full-length SUV39H2 and the C-terminus of TCTP (120-172 aa) were critical for binding. The interaction of SUV39H2 and TCTP was further confirmed by co-immunoprecipitation and immunofluorescence staining for colocalization. Moreover, depletion of TCTP by RNAi led to up-regulation of SUV39H2 protein, while TCTP overexpression reduced SUV39H2 protein level. The half-life of SUV39H2 protein was significantly extended upon TCTP depletion. These results clearly indicate that TCTP negatively regulates the expression of SUV39H2 post-translationally. Furthermore, SUV39H2 induced apoptotic cell death in TCTP-knockdown cells. Taken together, we identified SUV39H2, as a novel target protein of TCTP and demonstrated that SUV39H2 regulates cell proliferation of lung cancer cells.

• BMB Reports
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• 제44권6호
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• pp.369-374
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• 2011

MHC-I molecules play a critical role in immune surveillance against viruses by presenting peptides to cytotoxic T lymphocytes. Although the mechanisms by which MHC-I molecules assemble and acquire peptides in the ER are well characterized, how MHC-I molecules traffic to the cell surface remains poorly understood. To identify novel proteins that regulate the intracellular transport of MHC-I molecules, MHC-I-interacting proteins were isolated by affinity purification, and their identity was determined by mass spectrometry. Among the identified MHC-I-associated proteins was Tmp21, the human ortholog of yeast Emp24p, which mediates the ER-Golgi trafficking of a subset of proteins. Here, we show that Tmp21 binds to human classical and non-classical MHC-I molecules. The Tmp21-MHC-I complex lacks ${\beta}_2$-microglobulin, and the number of the complexes is increased when free MHC-I heavy chains are more abundant. Taken together, these results suggest that Tmp21 is a novel protein that preferentially binds to ${\beta}_2$-microglobulin-free MHC-I heavy chains.

• Radiation Oncology Journal
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• 제35권3호
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• pp.281-288
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• 2017

Purpose: The serum carcinoembryonic antigen (CEA) level has been recognized as a prognostic factor in colorectal cancer, and associated with response of rectal cancer to radiotherapy. This study aimed to identify CEA-interacting proteins in colon cancer cells and observe post-irradiation changes in their expression. Materials and Methods: CEA expression in colon cancer cells was examined by Western blot analysis. Using an anti-CEA antibody or IgG as a negative control, immunoprecipitation was performed in colon cancer cell lysates. CEA and IgG immunoprecipitates were used for liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Proteins identified in the CEA immunoprecipitates but not in the IgG immunoprecipitates were selected as CEA-interacting proteins. After radiation treatment, changes in expression of CEA-interacting proteins were monitored by Western blot analysis. Results: CEA expression was higher in SNU-81 cells compared with LoVo cells. The membrane localization of CEA limited the immunoprecipitation results and thus the number of CEA-interacting proteins identified. Only the Ras-related protein Rab-6B and lysozyme C were identified as CEA-interacting proteins in LoVo and SNU-81 cells, respectively. Lysozyme C was detected only in SNU-81, and CEA expression was differently regulated in two cell lines; it was down-regulated in LoVo but up-regulated in SNU-81 in radiation dosage-dependent manner. Conclusion: CEA-mediated radiation response appears to vary, depending on the characteristics of individual cancer cells. The lysozyme C and Rab subfamily proteins may play a role in the link between CEA and tumor response to radiation, although further studies are needed to clarify functional roles of the identified proteins.

• 생명과학회지
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• 제28권2호
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• pp.170-175
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• 2018

파킨슨병은 두번째로 많이 발병하는 퇴행성 신경질환이며 약 5-10%는 유전된다. Leucine-rich repeat kinase 2(LRRK2)는 그 돌연변이의 일부가 파킨슨병을 일으키는 유전자이다. LRRK2에는 인산화효소와 GTPase 기능이 있는 도메인과 함께 단백질 상호작용에 관여하는 Leucine-rich repeat (LRR), WD40 도메인이 존재하여, LRRK2와 상호작용하는 단백질이 파킨슨병 발병에 중요한 역할을 함을 암시한다. 우리는 이러한 LRRK2와 상호작용하는 단백질을 규명하여 그 단백질의 세포내 기능을 통해 역으로 LRRK2의 기능을 밝히고자 하였다. NIH3T3 세포 용해물을 LRRK2 항체와 IgG로 각각 면역침강하여 LRRK2 항체 침강반응에서만 특이적으로 나타나는 단백질 밴드를 질량 분석한 결과, methionyl-tRNA synthetase (MRS)로 나타났다. LRRK2와 MRS의 상호작용은 면역침강반응과 GST-pull down assay를 통해 확인됐다. 병을 유발하는, LRRK2의 돌연변이인 G2019S가 인산화효소 활성을 증가시키므로 LRRK2가 MRS를 인산화하는 지를 조사한 결과, LRRK2재조합단백질은 MRS 단백질을 인산화 하지 않았다. 또한 이들 두 단백질의 각각의 양 증가가 상대 단백질의 양 증가, 즉 안정성에 영향을 미치는 지를 조사하였으나 안정성의 변화를 관찰하지 못하였다. 결론적으로, MRS는 LRRK2와 상호작용을 하지만 LRRK2 인산화효소의 기질은 아니다.

• Journal of Microbiology
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• 제34권4호
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• pp.311-315
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• 1996

The retroviral Gag polyprotein directs the assembly of virion particles and plays an important role in some events after entry into a host cell. The Gag polyprotein of a virus mixture is responsible for inducing murine acquired immunodeficiency syndrome (MAIDS) when injected into susceptible strains of mice. In order to identify the host cellular proteins which interact with the MAIDS virus Gag proteins and possibly mediate the function of the Gag proteins, mouse T-cell leukemic cDNA expression library was screened using the yeast GAL4 two hybrid system. Of 11 individual positive clones, the clone Y1 was selected for the study of protein-protein interaction. Its DNA sequence revealed that it was an exact match to the murine SH3 domain-containing protein SH3P8. It is expressed as 2.4 kbp transcripts in testis at higher levels and in various tissues tested at lower levels. Glutathione S-transferase-Y1 fusion protein binds tightly to $Pr60^{def-gag}$ as well as $Pr65^{eco-gag}$.

• 대한의생명과학회지
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• 제18권2호
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• pp.96-103
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• 2012

Kv 4.2 ion channel protein has an ability to open at subthreshold membrane potentials and to recover quickly from inactivation. That is very important for neuronal signal transmission in vertebrate brain. In order to purify Kv 4.2 protein, the novel purification methods were experimented. The purification procedure utilized chromatography on DE-52 ion exchange column and affinity chromatography on a WGA-Sepharose 4B, and Kv 4.2 affinity column chromatography. It was found that 0.5% (wt./vol.) Triton X-100 detergent in lysis buffer worked well for Kv 4.2 protein solubilization from rat brain membrane. Protein quantitative determination was conducted by BCA method at 562 nm for each purification step to avoid determination interference of protein at 280 nm by detergent. The confirmation of Kv 4.2 existence and amount is performed using by SDS-PAGE/immunoblotting or 96-well dot blotting. The Kv 4.2 without interacting protein that contains carbohydrate, was purified from novel biochemical 3-steps purification method for further research.

• 대한의생명과학회지
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• 제14권2호
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• pp.77-81
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• 2008

Testis brain RNA-binding protein (TB-RBP) is a DNA/RNA binding protein. TB-RBP is mainly expressed in testis and brain and highly conserved protein with several functions, including chromosomal translocations, DNA repair, mitotic cell division, and mRNA transport, stabilization, and storage. In our previous study, we identified TB-RBP as an interacting partner for the catalytic subunit $(C{\alpha})$ of protein kinase A(PKA) and verified their interaction with several biochemical analyses. Here, we confirmed interaction between $C{\alpha}$. and TB-RBP in mammalian cells and determined the effect of $C{\alpha}$. on the function of TB-RBP. The activation of $C{\alpha}$. increased the TB-RBP function as a DNA-binding protein. These results suggest that the function of TB-RBP can be modulated by PKA and provide insights into the diverse role of PKA.

• 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.

• Bulletin of the Korean Chemical Society
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• 제35권1호
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• pp.83-86
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• 2014

Poly(A) polymerase (PAP) play an essential role for maturation of mRNA by adding the adenylate residues at the 3' end. PAP functions are regulated through protein-protein interaction at its C-terminal region. In this study, cyclophilin A (CypA), a member of the peptidyl-prolyl cis-trans isomerase family, was identified as a partner protein interacting with the C-terminal region PAP. The interaction between PAP and CypA was inhibited by the immunosuppressive drug cyclosporine A. Deletion analysis revealed that the N-terminal 56 residues of CypA are sufficient for the interaction with PAP. Interestingly, we observed that PAP and CypA colocalize in the nucleus during SDF-1-induced chemotaxis, implying that CypA could be involved in the regulation of polyadenylation by PAP in the chemotactic cells.

• 정보처리학회논문지B
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• 제13B권7호
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• pp.679-688
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• 2006

대용량 실험으로부터 산출된 단백질 상호작용 데이터는 위양성(false positive) 데이터의 비율이 높다는 단점을 가지고 있다. 본 논문에서는 오류가 섞여있는 단백질 상호작용 데이터를 입력으로 받아 각 단백질 상호작용의 신뢰도를 검증하는 시스템을 제안하고 구현하였다. 제안 시스템은 단백질 상호작용 데이터에 상호작용의 근거로서 사용될 수 있는 다양한 생물학적 특징들에 관한 데이터를 통합하고 특징 선택 방법을 사용하여 통합된 속성들 중 위양성 여부를 판별하는데 가장 적합한 특징들을 선택한 후 데이터 마이닝 분류 알고리즘을 적용하여 대용량 실험으로부터 산출된 단백질 상호작용 데이터의 신뢰도를 평가한다. 특징 선택의 결과와 분류 기법의 성능은 데이터 특성에 매우 의존하므로, 제안시스템에 가장 적합한 속성 부분집합과 가장 좋은 성능을 내는 분류 알고리즘을 찾기 위해 다양한 특징 선택 방법과 데이터 마이닝 분류 알고리즘들을 적용하고 그 성능을 다각적으로 비교분석 하였다. 실험 결과, 특징 선택 방법과 분류 알고리즘을 결합시킨 제안 시스템은 오류 데이터가 섞여있는 단백질 상호작용 데이터에서 실제로 상호작용하는 단백질 쌍을 골라내는 작업에 있어 기존 연구들에 비해 매우 뛰어난 성능을 보여줬다. 또한 본 연구를 통해 단백질 상호작용 데이터의 신뢰도를 검증함에 있어서 다양한 특징 선택 방법들과 분류 알고리즘들이 성능에 미치는 영향에 관해서도 정리할 수 있었다.