• 한국산학기술학회논문지
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• 제9권6호
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• pp.1795-1799
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• 2008

hnRNP K 단백질은 hnRNP 복합체를 구성하는 핵단백질들 중의 하나이며 시토신이 많은 RNA/DNA sequence에 잘 결합한다. 이 단백질은 핵 내에서만 머무르지 않고 핵과 세포질을 왕복하는 특징을 지니고 있다. hnRNP K의 기능을 조사하기 위하여 우선 hnRNP K와 상호 결합하는 세포내 단백질을 찾아내고자 하였다. 이를 위하여 본 연구에서는 이스트 two-hybrid 시스템을 사용하여 HeLa CDNA librar를 탐색하였다. 그 결과 얻은 클론들 중에는 사람의 hnRNP E1 (poly(rC) binding protein 1) cDNA (GenBank accession number XM_031585) 클론이 포함되어 있었다. 본 논문에서는 이스트 two-hybrid 시스템과 in vitro에서의 생화학적 실험을 통하여 hnRNP E1은 hnRNP K와 특이적으로 상호 결합한다는 것을 밝혔다.

• Animal cells and systems
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• 제12권3호
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• pp.143-148
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• 2008

Heterogeneous nuclear ribonucleoprotein E1(hnRNP E1) is one of the primary pre-mRNA binding proteins in human cells. It consists of 356 amino acid residues and harbors three hnRNP K homology(KH) domains that mediate RNA-binding. The hnRNP E1 protein was shown to play important roles in mRNA stabilization and translational control. In order to enhance our understanding of the cellular functions of hnRNP E1, we searched for interacting proteins through a yeast two-hybrid screening while using HeLa cDNA library as target. One of the cDNA clones was found to be human ribosomal protein L18a cDNA(GenBank accession number BC071920). We demonstrated in this study that human ribosomal protein L18a, a constituent of ribosomal protein large subunit, interacts specifically with hnRNP E1 in the yeast two-hybrid system. Such an interaction was observed for the first time in this study, and was also verified by biochemical assay.

• 한국산학기술학회논문지
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• 제18권12호
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• pp.575-580
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• 2017

본 연구는 hnRNP A1 단백질에 포함되어 있는 RGG 상자가 이 단백질의 세포내 위치에 미치는 영향 및 이 단백질의 안정화에 미치는 영향을 분석하는 것을 목적으로 하며 2014년 10월부터 약 3년 동안 수행되었다. 이를 위해 먼저, RGG상자 내의 6개의 아르기닌을 라이신으로 돌연변이를 만든 다음 pcDNA1-HA-hnRNP A1(6R/K)를 재조합하였다. 다음, 이 플라스미드 DNA를 HeLa 세포에 형질주입하여 HA-hnRNP A1(6R/K) 단백질의 세포내 위치에 미치는 영향을 면역형광현미경법으로 분석하였다. 그 결과 hnRNP A1(6R/K) 단백질은 (wt 단백질처럼 핵에 위치하지 못하고) 핵과 세포질 모두에 퍼져 있는 것을 확인하였다. hnRNP A1(6R/K)의 안정성을 조사하기 위해서는 pcDNA1-HA-hnRNP A1(6R/K)를 HeLa 세포에 형질 주입시킨 다음 발현된 단백질을 웨스턴 블랏 실험으로 분석하였는데, 그 결과 HA-hnRNP A1(6R/K)는 (잘려서) 크기가 작아진 것을 확인할 수 있었다. 이 결과들로부터 HA-hnRNP A1(6R/K)가 핵과 세포질 모두에 퍼져 있는 것은 6R/K 돌연변이가 hnRNP A1의 핵 위치 능력에 영향을 미쳤기 때문이 아니라 6R/K 돌연변이가 일어난 부위 또는 그 부근이 절단되어 hnRNP A1의 핵 위치 신호(nuclear localization singal)인 M9 도메인이 들어있는 C-말단 부위를 잃었기 때문이라는 점을 확인 할 수 있었다. 본 연구의 결과들은 hnRNP A1에 있는 RGG 상자의 아르기닌이 hnRNP A1의 안정성에 중요한 역할을 한다는 것을 제시한다. 세균에서 발현시켜서 정제된 hnRNP A1(6R/K)의 RNA-결합 능력에 대한 영향 분석은 추후 과제로 남겨둔다.

• BMB Reports
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• 제50권6호
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• pp.318-322
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• 2017

Brain cytoplasmic 200 RNA (BC200 RNA) is a neuron-specific non-coding RNA, implicated in the inhibition of local synaptodendritic protein synthesis, and is highly expressed in some cancer cells. Although BC200 RNA has been shown to inhibit translation in vitro, the cellular location of this inhibition is unknown. In this study, we used a BC200 RNA-recognizing antibody to identify the cellular locations of BC200 RNA in HeLa cervical carcinoma cells. We observed punctate signals in both the cytoplasm and nucleus, and further discovered that BC200 RNA co-localized with the p-body decapping enzyme, DCP1A, and the heterogeneous nuclear ribonucleoprotein E2 (hnRNP E2). The latter is a known BC200 RNA-binding partner protein and a constituent of p-bodies. This suggests that BC200 RNA is localized to p-bodies via hnRNP E2.

• Journal of Veterinary Science
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• 제22권5호
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• pp.62.1-62.13
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• 2021

Background: Canine mammary gland tumor (MGT) is the most common cancer in aged female dogs. Although it's important to identify reliable metastasis or prognostic factors by evaluating related to cell division, adhesion, and cancer stem cell-related transcription factor (TF) in metastasis-induced canine MGT, but there are limited studies. Objectives: We aimed to identify metastasis prognostic factors and cancer stem cell-TFs in canine MGTs. Methods: Age-matched female dogs diagnosed with MGT only were classified into metastatic and non-metastatic groups by histopathological staining of MGT tissues. The mRNA levels of cancer prognostic metastasis molecular factors (E-cadherin, ICAM-1, PRR14, VEGF, HPRT1, RPL4 and hnRNP H) and cancer stem cell-related TFs (Oct4, Sox2, and Nanog) were compared between metastatic and non-metastatic canine MGT tissues using qRT-PCR analysis. Results: The mRNA levels of ICAM-1, PRR14, VEGF, hnRNP H, Oct4, Sox2, and Nanog in metastatic MGT group were significantly higher than those in non-metastatic MGT group. However, mRNA level of RPL4 was significantly lower in metastatic MGT group. Loss of E-cadherin and HPRT1 was observed in the metastatic MGT group but it was not significant. Conclusions: Consistent expression patterns of all metastasis-related factors showing elevation in ICAM-1, PRR14, VEGF, hnRNP H, Oct4, Sox2, and Nanog, but decreases in RPL4 levels occurred in canine MGT tissues, which was associated with metastasis. Thus, these cancer prognostic metastasis factors and TFs of cancer stem cells, except for E-cadherin and HPRT1, can be used as reliable metastasis factors for canine MGT and therapeutic strategy.

• Molecules and Cells
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• 제27권6호
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• pp.657-665
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• 2009

The ovomucoid pre-mRNA has been folded into mini-hairpins adaptable for the RNA recognition motif (RRM) protein binding. The number of mini-hairpins were 372 for pre-mRNA and 83-86 for mature mRNA. The spatial arrangements are, in average, 16 nucleotides per mini-hairpin which includes 7 nt in the stem, 5.6 nt in the loop and 3.7 nt in the inter-hairpin spacer. The constitutive splicing system of ovomucoid-pre-mRNA is characterized by preferred order of intron removal of 5/6 > 7/4 > 2/1 > 3. The 5' splice sites (5'SS), branch point sequences (BPS) and 3' splice sites (3'SS) were identified and free energies involved have been estimated in 7 splice sites. Thermodynamic barriers for splice sites from the least (|lowest| -Kcal) were 5, 4, 7, 6, 2, 1, and 3; i.e., -18.7 Kcal, -20.2 Kcal, -21.0 Kcal, -24.0 Kcal, - 25.4 Kcal, -26.4 Kcal and -28.2 Kcal respectively. These are parallel to the kinetic data of splicing order reported in the literature. As a result, the preferred order of intron removals can be described by a consideration of free energy changes involved in the spliceosomal assembly pathway. This finding is consistent with the validity of hnRNP formation mechanisms in previous reports.