• Parasites, Hosts and Diseases
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• 제53권2호
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• pp.227-232
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• 2015

Genetic polymorphisms of pvdhfr and pvdhps genes of Plasmodium vivax were investigated in 83 blood samples collected from patients in the Philippines, Bangladesh, and Nepal. The SNP-haplotypes of the pvdhfr gene at the amino acid positions 13, 33, 57, 58, 61, 117, and 173, and that of the pvdhps gene at the positions 383 and 553 were analyzed by nested PCR-RFLP. Results suggest diverse polymorphic patterns of pvdhfr alone as well as the combination patterns with pvdhps mutant alleles in P. vivax isolates collected from the 3 endemic countries in Asia. All samples carried mutant combination alleles of pvdhfr and pvdhps. The most prevalent combination alleles found in samples from the Philippines and Bangladesh were triple mutant pvdhfr combined with single mutant pvdhps allele and triple mutant pvdhfr combined with double wild-type pvdhps alleles, respectively. Those collected from Nepal were quadruple mutant pvdhfr combined with double wild-type pvdhps alleles. New alternative antifolate drugs which are effective against sulfadoxine-pyrimethamine (SP)-resistant P. vivax are required.

• BMB Reports
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• 제50권7호
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• pp.373-378
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• 2017

The Jun activation-domain binding protein 1 (Jab1) induces p53 nuclear export and cytoplasmic degradation, but the underlying mechanism is poorly understood. Here, we show that phosphorylation at the threonine 155 residue is essential for Jab1-mediated p53 nuclear export. Jab1 stimulated phosphorylation of p53 at T155 was inhibited by curcumin, an inhibitor of COP9 signalosome (CSN)-associated kinases. The T155E mutant, which mimics phosphorylated p53, exhibited spontaneous cytoplasmic localization in the absence of Jab1. This process was prevented by leptinomycin B (LMB), but not by curcumin. The substitution of threonine 155 for valine (T155V) abrogated Jab1-mediated p53 nuclear export, indicating that phosphorylation at this site is essential for Jab1-mediated regulation of p53. Although T155E can be localized in the cytoplasm in the absence of Mdm2, the translocation of T155E was significantly enhanced by ectopic Hdm2 expression. Our data suggests that Jab1-mediated phosphorylation of p53 at Thr155 residue mediates nuclear export of p53.

• Asian Pacific Journal of Cancer Prevention
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• 제14권7호
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• pp.4295-4299
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• 2013

Background: Pretreatment serum p53 and epidermal growth factor receptor (EGFR) were assessed using enzyme-linked immunosorbent assay (ELISA) in patients with acute leukemia to analysis their roles in characterization of different subtypes of the disease. Materials and Methods: Serum samples from thirty two patients with acute myeloid leukemia (AML) and fourteen patients with acute lymphoid leukemia (ALL) were analysed, along with 24 from healthy individuals used as a control group. Results: The results demonstrated a significant increase of serum p53 and EGFR in patients with AML (p<0.0001) compared to the control group. Also, the results showed a significant increase of both markers in patients with ALL (p<0.05, p<0.0001 respectively). Sensitivities and specificities for these variables were 52% and 100% for p53, and 73.9%, 95.8% for EGFR. Serum p53 and EGFR could successfully differentiate between M4 and other AML subtypes, while these variables failed to discriminate among ALL subtypes. A positive significant correlation was noted between p53 and EGFR. Negative significant correlations were observed between these variables and both of hemoglobin (Hg) content and RBC count. Conclusions: Mutant p53 and EGFR are helpful serological markers for diagnosis of patients with AML or ALL and can aid in characterization of disease. Moreover, these markers may reflect carcinogenesis mechanisms.

• Animal cells and systems
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• 제6권3호
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• pp.263-269
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• 2002

The p53 tumor suppressor gene is among the most frequently mutated and studied genes in human cancer, but the mechanisms by which it sur presses tumor formation remain unclear. DNA damage regulates both the protein levels of p53 and its affinity for specific DNA sequences. Stabilization of p53 in response to DNA damage is caused by its dissociation from Mdm2, a downstream target gene of p53 and a protein that targets p53 for degradation in the proteosome. Recent studies have suggested that phosphorylation of human p53 at Ser20 is important for stabilizing p53 in response to DNA damage through disruption of the interaction between Mdm2 and p53. We generated mice with an allele encoding changes at Ser20, known to be essential for p53 accumulation following DNA damage, to enable analyses of p53 stabilization in vivo. Our data showed that the mutant p53 was clearly defective for full stabilization of p53 in response to DNA damage. We concluded that Ser20 phosphorylation is critical for modulating the negative regulation of p53 by Mdm2, probably through phosphorylation-dependent inhibition of p53-Mdm2 interaction in the physiological context.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1994년도 춘계학술대회 and 제3회 신약개발 연구발표회
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• pp.178-178
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• 1994

Hepatitis B Virus (HBV) is a DNA virus with a 3.2kb partially double-stranded genome. The life cycle of the virus involves a reverse transcription of the greater than genome length 3.5kb mRNA. This pegenomic RNA contains all the genetic information encoded by the virus and functions as an intermediate in viral replication. Tumor suppressor p53 has previously been shown to interact with the X-gene product of the HBV, which led us to hypothesize that p53 may act as a negative regulator of HBV replication and the role of the X-gene product is to overcome the p53-mediated restriction. As a first step to prove the above hypothesis, we tested whether p53 represses the propagation of HBV in in vitro replication system. By transient cotransfection of the plasmid containing a complete copy of the HBV genome and/or the plasmid encoding p53, we found that the replication of HBV is specifically blocked by wild-type p53. The levels of HBV DNA, HBs Ag and HBc/e Ag secreted in cell culture media were dramatically reduced upon coexpresion of wild-type p53 but not by the coexpression of the mutants of p53 (G154V and R273L). Furthermore, levels of RNAs originated from HBV genome were repressed more than 10 fold by the cotransfection of the p53 encoding plasmid. These results clearly states that p53 is a nesative regulator of the HBV replication. Next, to addresss the mechanism by which p53 represses the HBV replication, we performed the transient transfection experiments employing the pregenomic/core promoter-CAT(Chloramphenicol Acetyl Transferase) construct as a reporter. Cotransfection of wild-type p53 but not the mutant p53 expression plasmids repressed the CAT activity more than 8 fold. Integrating the above results, we propose that p53 represses the replication of HBV specifically by the down-regulation of the pregenomic/core promoter, which results in the reduced DNA synthesis of HBV. Currently, the mechanism by which HBV overcomes the observed p53-mediated restriction of replication is tinder investigation.

• 대한기관식도과학회:학술대회논문집
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• 대한기관식도과학회 1993년도 제27차 학술대회 초록집
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• pp.84-84
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• 1993

17번 염색체의 단완에 위치하여 종양의 억제에 관여하는 p53유전자의 돌연변이로 인한 불활성 p53단백의 과도발현이 유방암과 대장암을 비롯한 인체의 여러 암에 있어서 보고되었으며 일부 암에서는 예후인자로서의 가능성이 제시되었다. 저자들은 비교적 완만히 성장하지만 국소재발 및 원격전이가 잦은 두경부 영역의 선양낭포암에 있어서 p53 단백 발현 양상 및 예후 인자로의 가능성을 알아보고자 1982년부터 1991년 사이에 서울대학병원에서 병리조직학적으로 진단받은 환자 중 추적관찰이 가능하였고 병리조직을 얻을 수 있었던 23례에 대해 p53 단백에 대한 단클론항체를 이용하여 면역 조직화학적 염색을 시행하여 다음과 같은 결과를 얻었다. 총 23례 중 p53단백의 양성 발현을 보인 경우는 7fP(34%)이었으며, p53단백의 양성 발현과 국소재발 또는 원격전이와의 사이에 유의한 상관관계는 없었다.

• Journal of Korean Neurosurgical Society
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• 제37권1호
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• pp.48-53
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• 2005

Objective: The purpose of this study is to elucidate in vitro responses to combined gamma knife irradiation and p53 gene transfection on human malignant glioma cell lines. Methods: Two malignant human glioma cell lines, U87MG (p53-wild type) and U373MG (p53-mutant) were transfected with an adenoviral vector containing p53 (MOI of 50) before and after applying 20Gy of gamma irradiation. Various assessments were performed, including, cell viability by MTT assay; apoptosis by annexin assay; and cell cycle by flow cytometry, for the seven groups: mock, p53 only, gamma knife (GK) only, GK after LacZ, LacZ after GK, GK after p53, p53 after GK. Results: Cell survival decreased especially, in the subgroup transfected with p53 after gamma irradiation. Apoptosis tended to increase in p53 transfected U373 MG after gamma irradiation (apoptotic rate, 38.9%). The G2-M phase cell cycle arrest markedly increased by transfecting with p53, 48 hours after gamma knife irradiation in U373 MG (G2-M phase, 90.8%). Conclusion: These results suggest that the in vitro effects of combined gamma knife irradiation and p53 gene transfection is an augmentation of apoptosis and G2-M phase cell cycle arrest, which are more exaggerated in U373 MG with p53 transfection after gamma knife irradiation.

• Tuberculosis and Respiratory Diseases
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• 제41권4호
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• pp.339-353
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• 1994

연구배경 : p53 유전자는 염색체 17p에 존재하는 종양억제유전자인데, 돌연변이가 있는 경우의 mutant protein은 그 반감기가 4~8시간으로 wild type protein의 반감기 6~20분에 비하여 현저한 증가를 보이게 된다. 결과적으로 돌연변이가 있는 경우 p53 단백질이 과축적되어 과발현이 유발되는데, 이러한 기전을 이용하여 p53 면역조직화학염색은 p53유전자의 돌연변이의 검색 방법으로 많이 이용되고 있다. 유방암에서는 p53 핵단백질의 과발현이 있는 경우에 없을 때보다 무병생존기간 및 전체 생존기간 모두에서 유의하게 나쁜 임상 경과를 취함이 보고되고 있지만 폐암에서는 p53 유전자의 돌연변이 유무, 또는 p53단백의 과발현 유무가 예후에 미치는 영향은 아직까지 확실하지 않다. 저자 등은 한국인의 폐암에서 p53 종양억제 유전자의 돌연변이 빈도를 확인하고 이들이 폐암 환자의 임상 경과에 미치는 영향을 분석하며, 면역조직화학염색 및 PCR-SSCP 분석의 감수성과 특이성을 염기서열분석과 비교하여 확인하고자 본 연구를 시행하였다. 방법 : 근치적 폐절제술 후 임상경과를 추적중인 원발성폐암환자 75예를 대상으로 하였으며, 면역조직화학검사 및 분자생물학적 연구를 위하여는 이들의 종양조직(paraffin block)을 이용하였다. p53 단백질의 면역조직화학검사를 위한 일차항체로는 DO7(Novocastra, U.K.)을 사용하였고, PCR-SSCP 및 염기서열분석 등을 위하여는 파라핀 포매조직에서 암조직을 선택적으로 박절하여 DNA를 추출하여 이용하였다. 결과: 1) 전체 75예 폐암환자의 면역조직화학염색 결과 27예(36%)에서 p53단백질의 과발현을 보였다(Table 2.3, Fig 1). 2) 전체 환자의 p53 핵단백질 과발현 여부에 따른 중앙전체생존기간은 p53 음성군과 양성군 모두 25개월, 무병생존기간은 두군 모두 13개월로 동일하였다(Fig. 2). 3) PCR-SSCP 분석에 의한 p53 유전자의 mobility shift는 시험한 58예중 16예(27.6%)에서 확인되었다(Fig 3). Mobility shift 유무에 따른 중앙전체생존기간은 shift가 있는 군과 없는 군에서 각각 27개월과 20개월, 무병생존기간은 각각 8개월, 10개월로 유의한 차이가 없었다. 4) 염기서열분석을 통한 p53 유전자의 돌연변이는 시험한 29예중 10예(34.5%)에서 확인되었다(Fig 4). 염기서열분석 결과 돌연변이 유무에 따른 중앙전체생존기간은 돌연변이가 있었던 군과 없었던 군에서 각각 27개월, 22개월이었으며, 무병생존기간은 각각 20개월과 10개월로 유의한 차이는 없었으나 돌연변이가 있는 경우 조기재발을 하는 경향을 보였다(Fig 5, 6). 5) 면역조직화학염색은 PCR-SSCP 결과를 기준으로 할 때 민감도 67.0%, 특이도 74.0%, 그리고 일치도는 62.5% 이었다. PCR-SSCP의 결과는 염기서열분석 결과를 기준으로 할 때 민감도 91.8%, 특이도 96.2%, 일치도는 95.3% 이었다. 결론 : p53 핵단백질의 과발현 정도, PCR-SSCP 및 염기서열분석상 돌연변이 유무는 비소세포폐암환자의 근치적수술후 예후의 예측지표로서는 그 효용성이 적었으나, 돌연변이가 있는 경우 조기재발을 하는 경향을 보였다. 또한 p53유전자의 돌연변이 검색법으로는 면역조직화학염색보다는 PCR-SSCP법이 우수하였다.

• Molecules and Cells
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• 제19권2호
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• pp.289-293
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• 2005

BAF53 is an actin-related protein that shuttles between nucleus and cytoplasm. In the nucleus, it constitutes an integral component of many chromatin-modifying complexes such as the SWI/SNF, TIP60, TRRAP, and TIP48/49 complexes. BAF53 is essential for growth, but its function remains elusive. BAF53 homologues from yeast to humans have a conserved N-terminal motif, MS_(G/A)(G/A)__(V/L)YGG, which is unique to these proteins. Previously we showed that over-expression of an N-terminal deletion mutant of BAF53 ($BAF53_-{\Delta}N$) reduced the viability of HEK293 and HeLa cells. When we replaced the serine 2 and tyrosine 6 of this N-terminal motif with alanine, over-expression of the alanine-replaced BAF53 strongly impaired the growth of HEK293 cells whereas replacement with aspartate/glutamate had no effect. The alanine-replaced BAF53 mutants also stimulated p53-dependent transcription, in which the SWI/SNF and TRRAP complexes are involved. Our results demonstrate that serine 2 and tyrosine 6 play important roles in BAF53 activity.

• Molecules and Cells
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• 제19권2호
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• pp.219-222
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• 2005

The a-subunit of Escherichia coli tryptophan synthase (${\alpha}TS$), a component of the tryptophan synthase ${\alpha}_2{\beta}_2$ complex, is a monomeric 268-residues protein (Mr = 28,600). ${\alpha}TS$ by itself catalyzes the cleavage of indole-3-glycerol phosphate to glyceraldehyde-3-phosphate and indole, which is converted to tryptophan in tryptophan biosynthesis. Wild-type and P28L/Y173F double mutant ${\alpha}$-subunits were overexpressed in E. coli and crystallized at 298 K by the hanging-drop vapor-diffusion method. X-ray diffraction data were collected to $2.5{\AA}$ resolution from the wild-type crystals and to $1.8{\AA}$ from the crystals of the double mutant, since the latter produced better quality diffraction data. The wild-type crystals belonged to the monoclinic space group C2 ($a=155.64{\AA}$, $b=44.54{\AA}$, $c=71.53{\AA}$ and ${\beta}=96.39^{\circ}$) and the P28L/Y173F crystals to the monoclinic space group $P2_1$ ($a=71.09{\AA}$, b=52.70, $c=71.52{\AA}$ and ${\beta}=91.49^{\circ}$). The asymmetric unit of both structures contained two molecules of ${\alpha}TS$. Crystal volume per protein mass ($V_m$) and solvent content were $2.15{\AA}^3\;Da^{-1}$ and 42.95% for the wild-type and $2.34{\AA}^3\;Da^{-1}$ and 47.52% for the double mutant.