• Title/Summary/Keyword: Rab3A

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Identification of CEA-interacting proteins in colon cancer cells and their changes in expression after irradiation

  • Yoo, Byong Chul;Yeo, Seung-Gu
    • Radiation Oncology Journal
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    • v.35 no.3
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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.

Deciphering the role of a membrane-targeting domain in assisting endosomal and autophagic membrane localization of a RavZ protein catalytic domain

  • Park, Jui-Hee;Lee, Seung-Hwan;Park, Sang-Won;Jun, Yong-Woo;Kim, Kunhyung;Jeon, Pureum;Kim, Myungjin;Lee, Jin-A;Jang, Deok-Jin
    • BMB Reports
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    • v.54 no.2
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    • pp.118-123
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    • 2021
  • The bacterial effector protein RavZ from a pathogen can impair autophagy in the host by delipidating the mammalian autophagy-related gene 8 (mATG8)-phosphatidylethanolamine (PE) on autophagic membranes. In RavZ, the membrane-targeting (MT) domain is an essential function. However, the molecular mechanism of this domain in regulating the intracellular localization of RavZ in cells is unclear. In this study, we found that the fusion of the green fluorescent protein (GFP) to the MT domain of RavZ (GFP-MT) resulted in localization primarily to the cytosol and nucleus, whereas the GFP-fused duplicated-MT domain (GFP-2xMT) localized to Rab5- or Rab7-positive endosomes. Similarly, GFP fusion to the catalytic domain (CA) of RavZ (GFP-CA) resulted in localization primarily to the cytosol and nucleus, even in autophagy-induced cells. However, by adding the MT domain to GFP-CA (GFP-CA-MT), the cooperation of MT and CA led to localization on the Rab5-positive endosomal membranes in a wortmannin-sensitive manner under nutrient-rich conditions, and to autophagic membranes in autophagy-induced cells. In autophagic membranes, GFP-CA-MT delipidated overexpressed or endogenous mATG8-PE. Furthermore, GFP-CA△α3-MT, an α3 helix deletion within the CA domain, failed to localize to the endosomal or autophagic membranes and could not delipidate overexpressed mATG8-PE. Thus, the CA or MT domain alone is insufficient for stable membrane localization in cells, but the cooperation of MT and CA leads to localization to the endosomal and autophagic membranes. In autophagic membranes, the CA domain can delipidate mATG8-PE without requiring substrate recognition mediated by LC3-interacting region (LIR) motifs.

Functional characterization of ABA signaling components using transient gene expression in rice protoplasts

  • Song, In-Sik;Moon, Seok-Jun;Kim, Jin-Ae;Yoon, Insun;Kwon, Taek-Ryoun;Kim, Beom-Gi
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2017.06a
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    • pp.109-109
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    • 2017
  • The core components of ABA-dependent gene expression signaling have been identified in Arabidopsis and rice. This signaling pathway consists of four major components; group A OsbZIPs, SAPKs, subclass A OsPP2Cs and OsPYL/RCARs in rice. These might be able to make thousands of combinations through interaction networks resulting in diverse signaling responses. We tried to characterize those gene functions using transient gene expression for rice protoplasts (TGERP) because it is instantaneous and convenient system. Firstly, in order to monitor the ABA signaling output, we developed reporter system named pRab16A-fLUC which consists of Rab16A promoter of rice and luciferase gene. It responses more rapidly and sensitively to ABA than pABRC3-fLUC that consists of ABRC3 of HVA1 promoter in TGERP. We screened the reporter responses for over-expression of each signaling components from group A OsbZIPs to OsPYL/RCARs with or without ABA in TGERP. OsbZIP46 induced reporter most strongly among OsbZIPs tested in the presence of ABA. SAPKs could activate the OsbZIP46 even in the ABA independence. Subclass A OsPP2C6 and -8 almost completely inhibited the OsbZIP46 activity in the different degree through the SAPK9. Lastly, OsPYL/RCAR2 and -5 rescued the OsbZIP46 activity in the presence of SAPK9 and OsPP2C6 dependent on ABA concentration and expression level. By using TGERP, we could characterize successfully the effects of ABA dependent gene expression signaling components in rice. In conclusion, TGERP represents very useful technology to study systemic functional genomics in rice or other monocots.

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Early Preterm Infant Feeding Behaviors in Adolescent and Adult Mothers

  • Kanhadilok, Supannee;Brown, Lisa
    • Asian Journal for Public Opinion Research
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    • v.4 no.1
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    • pp.51-72
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    • 2016
  • Background: Adolescent mothers may find the transition to motherhood to be overwhelming when compared to adult mothers. The adaptiveness of mother and infant feeding behaviors may greatly influence the quality of adolescent mothers' feeding interactions when compared to adult mothers. Purpose: To examine the adaptiveness of early maternal and infant feeding behavior between adolescent mothers and adult mothers. Method: In this secondary analysis, six adolescent mothers (< 20 years old) and six adult mothers were videotaped while feeding. The mothers were observed 3 times: when the infants were in the hospital, 1 month old, and 4 months old. The videotapes were coded using four scales of the Parent-Child Early Relational Assessment (PCERA). The two maternal scales were Positive Affective Involvement and Sensitivity/Responsiveness (PAISR) and Regulation of Affect and Behavior (RAB). The two infant scales were Infant Positive Affect, Communication, and Social Skills (IPACS) and Infant Regulation of Affect and Behavior (IRAB). The dyads were videotaped just before hospital discharge and at one and four months corrected age. The Wilcoxon signed-rank test was used to examine the difference in the PCERA between the two groups. Results: Adolescent mothers had a significantly lower score on the PAISR before hospital discharge (p-value 0.005) and at one month corrected age (p-value 0.008) than adult mothers. Adolescent mothers also had a significantly lower score on the RAB before hospital discharge, 1 and 4 months corrected age than adult mothers. There were no significant differences in IPACS and IRAB of infants of adolescent and adult mothers. Conclusion: Clinicians need to be aware of the differences in sensitivity that occur.

P. aeruginosa EMS1의 mutagen 처리를 통한 고기능 유화제 생산 균주 개발 및 유전자 확인

  • 이근희;한창민;이준훈;이경민;차미선;이상준
    • Proceedings of the Korean Environmental Sciences Society Conference
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    • 2001.05a
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    • pp.244-245
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    • 2001
  • 산폐유의 농도를 l~8% 까지 단계별로 조정하여 P. aeruginosa EMS1과 mutant strain A34의 유화활성 및 생육도, 표면장력 등을 서로 비교하였다. P. aeruginosa의 경우 1%의 산폐유가 첨가되었을 때 유화활성이 가장 높았으며, 1~3%까지 유화활성이 높게 나타났으며, mutant strain의 경우 2%에서 가장 높은 유화활성 값을 나타내었고, 1~3%까지 유화활성이 높게 유지되었다. P. aeruginosa EMS1 및 mutant strain A34 모두 4% 농도에서 가장 높게 나타났다. 표면장력과 biosurfactant의 상대적인 양을 나타내는 Fcmc의 경우 P. aeruginosa EMS1 보다 mutant strain A34가 우수하게 나타났다. PCR 결과 P. aeruginosa EMS1은 rhlamnolipid의 coding 유전자인 rhlRAB gene을 가지는 것으로 확인되었다.

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Proteomic Analysis and the Antimetastatic Effect of N-(4methyl)phenyl-O-(4-methoxy) phenyl-thionocarbamate-Induced Apoptosis in Human Melanoma SK-MEL-28 cells

  • Choi Su-La;Choi Yun-Sil;Kim Young-Kwan;Sung Nack-Do;Kho Chang-Won;Park Byong-Chul;Kim Eun-Mi;Lee Jung-Hyung;Kim Kyung-Mee;Kim Min-Yung;Myung Pyung-Keun
    • Archives of Pharmacal Research
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    • v.29 no.3
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    • pp.224-234
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    • 2006
  • We employed human SK-MEL-28 cells as a model system to identify cellular proteins that accompany N-(4-methyl)phenyl-O-(4-methoxy)phenyl-thionocarbamate (MMTC)-induced apoptosis based on a proteomic approach. Cell viability tests revealed that SK-MEL-28 skin cancer cells underwent more cell death than normal HaCaT cells in a dose-dependent manner after treatment with MMTC. Two-dimensional electrophoresis in conjunction with matrixassisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry analysis or computer matching with a protein database further revealed that the MMTC-induced apoptosis is accompanied by increased levels of caspase-1, checkpoint suppressor-1, caspase-4, NF-kB inhibitor, AP-2, c-Jun-N-terminal kinase, melanoma inhibitor, granzyme K, G1/S specific cyclin D3, cystein rich protein, Ras-related protein Rab-37 or Ras-related protein Rab-13, and reduced levels of EMS (oncogene), ATP synthase, tyrosine-phosphatase, Cdc25c, 14-3-3 protein or specific structure of nuclear receptor. The migration suppressing effect of MMTC on SK-MEL-28 cell was tested. MMTC suppressed the metastasis of SK-MEL-8 cells. It was also identified that MMTC had little angiogenic effect because it did not suppress the proliferation of HUVEC cell line. These results suggest that MMTC is a novel chemotherapeutic and metastatic agents against the SK-MEL-28 human melanoma cell line.

Identification of Differentially Expressed Genes by TCDD in Human Bronchial Cells: Toxicogenomic Markers for Dioxin Exposure

  • Park, Chung-Mu;Jin, Kyong-Suk;Lee, Yong-Woo
    • Biomedical Science Letters
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    • v.18 no.1
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    • pp.1-9
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    • 2012
  • Differentially expressed genes by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) were identified in order to evaluate them as dioxin-sensitive markers and crucial signaling molecules to understand dioxin-induced toxic mechanisms in human bronchial cells. Gene expression profiling was analyzed by cDNA microarray and ten genes were selected for further study. They were cytochrome P450, family 1, subfamily B, polypeptide 1 (CYP1B1), S100 calcium binding protein A8 (calgranulin A), S100 calcium binding protein A9 (calgranulin B), aldehyde dehydrogenase 1 family, member A3 (ALDH6) and peroxiredoxin 5 (PRDX5) in up-regulated group. Among them, CYP1B1 was used as a hallmark for dioxin and sharply increased by TCDD exposure. Down-regulated genes were IK cytokine, interferon-induced protein with tetratricopeptide repeats 1 (IFIT1), nuclease sensitive element binding protein 1 (NSEP1), protein tyrosine phosphatase type VI A, member 1 (PTP4A1), ras oncogene family 32 (RAB32). Although up-regulated 4 genes in microarray were coincided with northern hybridization, down-regulated 5 genes showed U-shaped expression pattern which is sharply decreased at lower doses and gradually increased at higher doses. These results introduce some of TCDD-responsive genes can be sensitive markers against TCDD exposure and used as signaling cues to understand toxicity initiated by TCDD inhalation in pulmonary tissues.

Gene Expression Analysis of Hepatic Response Induced by Gentamicin in Mice

  • Oh, Jung-Hwa;Park, Han-Jin;Hwang, Ji-Yoon;Jeong, Sun-Young;Lim, Jung-Sun;Kim, Yong-Bum;Yoon, Seok-Joo
    • Molecular & Cellular Toxicology
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    • v.3 no.1
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    • pp.60-67
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    • 2007
  • Gentamicin is a broad-spectrum aminoglycoside antibiotic used in the treatment of bacterial infection. Although side effects of gentamicin such as nephrotoxicity and ototoxicity have been investigated, the information on the hepatic effects of gentamicin is still limited. In the present study, gene expression profiles were analyzed in the liver of gentamicin treated mice using Affymetrix GeneChip$^{(R)}$ Mouse Expression 430A 2.0 Array. Totally, 400 genes were identified as being either up- or down-regulated over 1.5-fold changes (P<0.01) in the liver of gentamicin treated mice. Among these deregulated genes, 16 up-regulated genes mainly involved in transport (Kif5b, Pex14, Rab14, Clcn3, and Necap1) and 20 down-regulated genes involved in lipid and other metabolisms (Hdlbp, Gm2a, Uroc1, and Dak) were selected using k-means clustering algorithm. The functional classification of differentially expressed genes represented that several stress-related genes were regulated in the liver by gentamicin treatment. This data may contribute in understanding the molecular mechanism in the liver of gentamicin treated mice.

The Mediating Effect of Dementia Recognition on the Number of Chronic Diseases and Dementia Prevention Behaviors of Elders in Rural Communities (농촌 지역사회노인의 만성질병수와 치매예방행위에 미치는 치매인식의 매개효과)

  • Park, Pilnam
    • Journal of Korean Academy of Rural Health Nursing
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    • v.15 no.2
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    • pp.41-48
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    • 2020
  • Purpose: This study was a descriptive study to identify the factors affecting the dementia prevention behavior of elders in rural communities. Methods: The participants in this study were 125 elders aged 60 or older who lived in Eup or Myeon areas of P city. For data analysis, SPSS/WIN 22.0 was used to perform descriptive statistics, t-test, ANOVA, Pearson correlation, and linear multiple regression and mediated effects. Results: Scores for dementia recognition, dementia attitude and dementia prevention behavior averaged 5.6±2.50 points in the 0~11 range, 38.8±4.59 in 14~56 and 20.2±3.59 in 10~30 respectively. Dementia recognition (a), dementia attitude (b), dementia prevention behavior (c) and the number of chronic diseases of the elders (d) were positively or negatively correlated with each other (rab=.29, p<.01; rbc=.26, p<.01; rac=.36, p<.01; rad=-.29, p<.01; rcd=.19, p<.05). Factors affecting dementia prevention behavior were dementia recognition, dementia attitude, and degree of dementia interest. When the number of chronic diseases affects dementia prevention behavior, dementia recognition has a mediating effect. Conclusion: In order to prevent dementia among elders in rural areas, appropriate management of chronic diseases and provision of appropriate dementia-related education and information to enhance dementia recognition should be provided.

A WUSCHEL Homeobox Transcription Factor, OsWOX13, Enhances Drought Tolerance and Triggers Early Flowering in Rice

  • Minh-Thu, Pham-Thi;Kim, Joung Sug;Chae, Songhwa;Jun, Kyong Mi;Lee, Gang-Seob;Kim, Dong-Eun;Cheong, Jong-Joo;Song, Sang Ik;Nahm, Baek Hie;Kim, Yeon-Ki
    • Molecules and Cells
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    • v.41 no.8
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    • pp.781-798
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    • 2018
  • Plants have evolved strategies to cope with drought stress by maximizing physiological capacity and adjusting developmental processes such as flowering time. The WOX13 orthologous group is the most conserved among the clade of WOX homeodomain-containing proteins and is found to function in both drought stress and flower development. In this study, we isolated and characterized OsWOX13 from rice. OsWOX13 was regulated spatially in vegetative organs but temporally in flowers and seeds. Overexpression of OsWOX13 (OsWOX13-ov) in rice under the rab21 promoter resulted in drought resistance and early flowering by 7-10 days. Screening of gene expression profiles in mature leaf and panicles of OsWOX13-ov showed a broad spectrum of effects on biological processes, such as abiotic and biotic stresses, exerting a cross-talk between responses. Protein binding microarray and electrophoretic mobility shift assay analyses supported ATTGATTG as the putative cis-element binding of OsWOX13. OsDREB1A and OsDREB1F, drought stress response transcription factors, contain ATTGATTG motif(s) in their promoters and are preferentially expressed in OsWOX13-ov. In addition, Heading date 3a and OsMADS14, regulators in the flowering pathway and development, were enhanced in OsWOX13-ov. These results suggest that OsWOX13 mediates the stress response and early flowering and, thus, may be a regulator of genes involved in drought escape.