scholarly journals Tumor Suppressor SMAR1 Activates and Stabilizes p53 through Its Arginine-Serine-rich Motif

2005 ◽  
Vol 280 (16) ◽  
pp. 16019-16029 ◽  
Author(s):  
Archana Jalota ◽  
Kamini Singh ◽  
Lakshminarasimhan Pavithra ◽  
Ruchika Kaul-Ghanekar ◽  
Shahid Jameel ◽  
...  
Keyword(s):  
Cell Fate ◽  
The Novel ◽  
Nuclear Retention ◽  
Post Translational Modifications ◽  
Rich Domain ◽  
Dna Damaging Agents ◽  
Proliferation And Apoptosis ◽  
Interfering Rna ◽  
The Guardian

Various stresses and DNA-damaging agents trigger transcriptional activity of p53 by post-translational modifications, making it a global regulatory switch that controls cell proliferation and apoptosis. Earlier we have shown that the novel MAR-associated protein SMAR1 interacts with p53. Here we delineate the minimal domain of SMAR1 (the arginine-serine-rich domain) that is phosphorylated by protein kinase C family proteins and is responsible for p53 interaction, activation, and stabilization within the nucleus. SMAR1-mediated stabilization of p53 is brought about by inhibiting Mdm2-mediated degradation of p53. We also demonstrate that this arginine-serine (RS)-rich domain triggers the various cell cycle modulating proteins that decide cell fate. Furthermore, phenotypic knock-down experiments using small interfering RNA showed that SMAR1 is required for activation and nuclear retention of p53. The level of phosphorylated p53 was significantly increased in the thymus of SMAR1 transgenic mice, showingin vivosignificance of SMAR1 expression. This is the first report that demonstrates the mechanism of action of the MAR-binding protein SMAR1 in modulating the activity of p53, often referred to as the “guardian of the genome.”

scholarly journals Inhibiting lysine 353 oxidation of GRP78 by a hypochlorous probe targeting endoplasmic reticulum promotes autophagy in cancer cells

2019 ◽  
Vol 10 (11) ◽  
Author(s):  
Junya Ning ◽  
Zhaomin Lin ◽  
Xuan Zhao ◽  
Baoxiang Zhao ◽  
Junying Miao
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Growth ◽  
Cancer Cells ◽  
Cell Fate ◽  
Hypochlorous Acid ◽  
Dependent Manner ◽  
Autophagy Flux ◽  
Post Translational Modifications

Abstract The level of hypochlorous acid (HOCl) in cancer cells is higher than that in non-cancer cells. HOCl is an essential signal for the regulation of cell fate and works mainly through the protein post-translational modifications in cancer cells. However, the mechanism of HOCl regulating autophagy has not been clarified. Here we reported that a HOCl probe named ZBM-H targeted endoplasmic reticulum and induced an intact autophagy flux in lung cancer cells. Furthermore, ZBM-H promoted the binding of GRP78 to AMPK and increased the phosphorylation of AMPK in a dose- and time-dependent manner. GRP78 knockdown inhibited ZBM-H-induced AMPK phosphorylation and ZBM-H-stimulated autophagy. In addition, mass spectrometry combined with point mutation experiments revealed that ZBM-H increased GRP78 activity by inhibiting HOCl-induced lysine 353 oxidation of GRP78. Following ZBM-H treatment in vitro and in vivo, cell growth was significantly inhibited while apoptosis was induced. Nevertheless, exogenous HOCl partially reversed ZBM-H-inhibited cell growth and ZBM-H-induced GRP78 activation. In brief, we found that an endoplasmic reticulum-targeted HOCl probe named ZBM-H, acting through attenuating HOCl-induced GRP78 oxidation, inhibited tumor cell survival by promoting autophagy and apoptosis. Overall, these data demonstrated a novel mechanism of hypochlorous acid regulating autophagy by promoting the oxidation modification of GRP78.

scholarly journals Identification of the Candida albicans Cap1p Regulon

2009 ◽  
Vol 8 (6) ◽  
pp. 806-820 ◽  
Author(s):  
Sadri Znaidi ◽  
Katherine S. Barker ◽  
Sandra Weber ◽  
Anne-Marie Alarco ◽  
Teresa T. Liu ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Transcriptional Activation ◽  
Leucine Zipper ◽  
Target Genes ◽  
Open Reading Frames ◽  
Nuclear Retention ◽  
Tiling Microarrays ◽  
Rich Domain ◽  
C Terminus

ABSTRACT Cap1p, a transcription factor of the basic region leucine zipper family, regulates the oxidative stress response (OSR) in Candida albicans. Alteration of its C-terminal cysteine-rich domain (CRD) results in Cap1p nuclear retention and transcriptional activation. To better understand the function of Cap1p in C. albicans, we used genome-wide location profiling (chromatin immunoprecipitation-on-chip) to identify its transcriptional targets in vivo. A triple-hemagglutinin (HA3) epitope was introduced at the C terminus of wild-type Cap1p (Cap1p-HA3) or hyperactive Cap1p with an altered CRD (Cap1p-CSE-HA3). Location profiling using whole-genome oligonucleotide tiling microarrays identified 89 targets bound by Cap1p-HA3 or Cap1p-CSE-HA3 (the binding ratio was at least twofold; P ≤ 0.01). Strikingly, Cap1p binding was detected not only at the promoter region of its target genes but also at their 3′ ends and within their open reading frames, suggesting that Cap1p may associate with the transcriptional or chromatin remodeling machinery to exert its activity. Overrepresented functional groups of the Cap1p targets (P ≤ 0.02) included 11 genes involved in the OSR (CAP1, GLR1, TRX1, SOD1, CAT1, and others), 13 genes involved in response to drugs (PDR16, MDR1, FLU1, YCF1, FCR1, and others), 4 genes involved in phospholipid transport (PDR16, GIT1, RTA2, and orf19.932), and 3 genes involved in the regulation of nitrogen utilization (GST3, orf19.2693, and orf19.3121), suggesting that Cap1p has other cellular functions in addition to the OSR. Bioinformatic analyses of the bound sequences suggest that Cap1p recognizes the DNA motif 5′-MTKASTMA. Finally, transcriptome analyses showed that increased expression generally accompanies Cap1p binding at its targets, indicating that Cap1p functions as a transcriptional activator.

scholarly journals Effect of SOCS1 silencing on proliferation and apoptosis of melanoma cells: An in vivo and in vitro study

Tumor Biology ◽  
2017 ◽  
Vol 39 (5) ◽  
pp. 101042831769431
Author(s):  
Sheng-Jia Yu ◽  
Zi-Wen Long
Keyword(s):  
Messenger Rna ◽  
Small Interfering Rna ◽  
Melanoma Cells ◽  
Janus Kinase ◽  
Extracellular Signal ◽  
Proliferation And Apoptosis ◽  
Protein Expressions ◽  
Interfering Rna

This study aimed to investigate the effect of SOCS1 silencing on the proliferation and apoptosis of melanoma cells by in vivo and in vitro studies. Immunohistochemical staining was used to detect SOCS1 expression in melanoma tissues and pigmented nevi. Quantitative real-time polymerase chain reaction and western blotting were applied to detect the messenger RNA and protein expressions of SOCS1 in primary human melanocytes and malignant melanoma cell lines (A375, SK-MEL-5, M14, and MV3). Melanoma cells were assigned into mock, negative small interfering RNA, and SOCS1-small interfering RNA groups. The proliferation, cell cycle and apoptosis, and messenger RNA expression of SOCS1 in MV3 and A375 cells were detected using MTT assay, flow cytometry, and quantitative real-time polymerase chain reaction, respectively. The expressions of SOCS1 protein, extracellular signal–regulated kinase, and janus kinase signal transduction and activators of transcription signaling pathways–related proteins were detected using western blotting. After the establishment of subcutaneous xenograft tumor models in nude mice, the latent period, size, volume and growth speed of xenograft tumors in the mock, negative small interfering RNA, and SOCS1-small interfering RNA groups were examined and compared. The results indicated that positive expression rate of SOCS1 was higher in malignant melanoma tissues than in pigmented nevi. MV3 cells had the highest messenger RNA and protein expressions of SOCS1, followed by A357 cells. Compared with the mock and negative small interfering RNA groups, SOCS1-small interfering RNA group showed lower cell viability, elevated cell apoptosis, more cells in G0/G1 phase and less cells in S and G2/M phases, and decreased messenger RNA and protein expressions of SOCS1, p-ERK1/2, p-JAK2, p-STAT1, and p-STAT3. Compared with the mock and negative small interfering RNA groups, the SOCS1-small interfering RNA group showed longer latent period of tumor, smaller tumor size and volume, and smoother tumor growth curve. To conclude, SOCS1 silencing can inhibit proliferation and induce apoptosis of MV3 and A357 melanoma cells in vivo and in vitro by inhibiting extracellular signal–regulated kinase and janus kinase signal transduction and activators of transcription signaling pathways.

scholarly journals iPSC-Based Phenomic Screen Revealed an Impact of Uncontrolled NFkB Activity at the Initiating Stages of AML1-ETO Related Leukemia

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3911-3911
Author(s):  
Akira Niwa ◽  
Tatsutoshi Nakahata ◽  
Megumu K Saito
Keyword(s):  
Cell Fate ◽  
Conflicts Of Interest ◽  
Genetic Alterations ◽  
The Novel ◽  
Early Stages ◽  
Immature Cells ◽  
Colony Forming Assay ◽  
Functional Knockdown

Abstract Onset of acute myeloid leukemia (AML) has been accounted for by cooperation between multiple genetic alterations which induce abnormal control of various cellular pathways. Among the previously listed leukemogenic lesions, AML1-ETO fusion (AE) generated by translocation (8;21) (q22;q22) is one of the common mutations observed in 20-40% of patients. AE affects transcriptional regulation associated with hematopoietic differentiation, while 60% of AE-positive AML cases are shown to have together other types of mutation of genes involved in cell proliferation, such as receptor tyrosine kinase (RTK) c-kit and FLT3. However, the detailed mechanisms of how they work in the very early stages of leukemogenesis and what unknown "cooperative " cues function in those periods. From this viewpoint, in order to identify novel cellular molecules involved in the acquisition of leukemic phenotypes, we have conducted the gene-trap strategy-based phenomic screen in the use of pluripotent stem cell (PSC)-derived hematopoietic culture. Through our screening, we found that the functional knockdown of a gene NSFL1c, also known as p47, enhanced the activities of hematopoietic progenitor cells harboring AE to show leukemic phenotype both in vivo and in vitro: Cells differentiated from AE-PSCs which have additionally the poly A trapping sequence inserted in NSFL1c locus showed doubled efficiency in engraftment into immunodeficient NOG mice than cells without trapping (3.1% v.s. 1.3%, 16 weeks after intra bone marrow transplantation), and also showed the significantly higher colony replating efficacy in methylcellulose colony forming assay. In order to clear what lineages of cells were most responsible for those phenomena, we next performed those colony forming experiments after sorting of CD34+CD43+CD13- immature cells, CD34+/-CD43+CD13+ myeloid cells and CD71+CD41+ erythro-megakaryocytic subpopulations of cells. As a result, CD34+/-CD43+CD13+ myeloid precursors showed the strongest tendencies to emerge highly proliferative clones followed by CD34+CD43+CD13- immature progenitors in the presence of NSF1c trapping. Interestingly, those activities were cancelled in the absence of AE expression. NSFL1c is one of the component of NEMO complex which binds to ubiquitinated NEMO and induces its degradation, resulting in reduced IKK and elevated NFkB activities. In AML, elevated NF-κB pathways have been detected in more than 30% of patients. Although NF-κB signaling networks have proved induced by inflammatory and immune signals, and previous studies in vivo and in vitro showed their abnormal activities make leukemia cells escape from cell death and go into abnormal proliferation, the detailed mechanisms of how they are involved in the pathogenesis, in particular at the very early stages of the leukemogenesis, are well not defined. Our data therefore reflect the novel mechanisms behind the deviation of progenitor cell fate from normal to abnormal pathway leading to the emergence of leukemic initiating cells, and suggested the myeloid-biased leukemogenic potentials. Disclosures No relevant conflicts of interest to declare.

scholarly journals Loss of peptidase D binding restores the tumor suppressor functions of oncogenic p53 mutants

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Lu Yang ◽  
Yun Li ◽  
Arup Bhattacharya ◽  
Yuesheng Zhang
Keyword(s):  
Tumor Suppressor ◽  
Cell Fate ◽  
Posttranslational Modifications ◽  
Tumor Suppressor P53 ◽  
Cancer Cell Growth ◽  
Rich Domain ◽  
Cancer Mutation ◽  
Peptidase D

AbstractTumor suppressor p53, a critical regulator of cell fate, is frequently mutated in cancer. Mutation of p53 abolishes its tumor-suppressing functions or endows oncogenic functions. We recently found that p53 binds via its proline-rich domain to peptidase D (PEPD) and is activated when the binding is disrupted. The proline-rich domain in p53 is rarely mutated. Here, we show that oncogenic p53 mutants closely resemble p53 in PEPD binding but are transformed into tumor suppressors, rather than activated as oncoproteins, when their binding to PEPD is disrupted by PEPD knockdown. Once freed from PEPD, p53 mutants undergo multiple posttranslational modifications, especially lysine 373 acetylation, which cause them to refold and regain tumor suppressor activities that are typically displayed by p53. The reactivated p53 mutants strongly inhibit cancer cell growth in vitro and in vivo. Our study identifies a cellular mechanism for reactivation of the tumor suppressor functions of oncogenic p53 mutants.

Blocking Notch1 signaling by RNA interference can induce growth inhibition in HeLa cells

2007 ◽  
Vol 17 (2) ◽  
pp. 511-516 ◽  
Author(s):  
H. Yu ◽  
X. Zhao ◽  
S. Huang ◽  
L. Jian ◽  
G. Qian ◽  
...  
Keyword(s):  
Rna Interference ◽  
Growth Inhibition ◽  
Notch Signaling ◽  
Hela Cells ◽  
Transmembrane Receptors ◽  
Proliferation And Apoptosis ◽  
Interfering Rna ◽  
Induce Growth Inhibition

The Notch proteins constitute a family of transmembrane receptors that play a pivotal role in cellular differentiation, proliferation, and apoptosis. RNA interference of Presenilin1 (PS1) and Notch1 was carried out in this research to determine whether it could block Notch signaling and induce growth inhibition in HeLa cells. We transfected synthesized target small interfering RNA (siRNA) into HeLa cells, and blocking of Notch signaling was detected by C-promoter binding factor-1 (CBF1) reporter. We then conducted cell proliferation assay. Cells transfected with PS1 and Notch1 siRNA showed great inhibition in proliferation compared to the controls in vitro and in vivo. We conclude that RNA interference of PS1 or Notch1 can block Notch signaling and consequently induce growth inhibition of HeLa cells.

Uptake, Internalization and Degradation of the Novel Plasminogen Activator Reteplase (BM 06.022) in the Rat

1995 ◽  
Vol 74 (06) ◽  
pp. 1501-1510 ◽  
Author(s):  
J Kuiper ◽  
H van de Bilt ◽  
U Martin ◽  
Th J C van Berkel
Keyword(s):  
Plasminogen Activator ◽  
Liver Cells ◽  
The Novel ◽  
Uptake Mechanism ◽  
Liver Uptake ◽  
Lysosomal Compartment ◽  
Β Phase ◽  
Α Phase ◽  
Parenchymal Liver

SummaryThe catabolism of the novel plasminogen activator reteplase (BM 06.022) was described. For this purpose BM 06.022 was radiolabelled with l25I or with the accumulating label l25I-tyramine cellobiose (l25I-TC).BM 06.022 was injected at a pharmacological dose of 380 μg/kg b.w. and it was cleared from the plasma in a biphasic manner with a half-life of about 1 min in the α-phase and t1/2of 20-28 min in the β-phase. 28% and 72% of the injected dose was cleared in the α-phase and β-phase, respectively. Initially liver, kidneys, skin, bones, lungs, spleen, and muscles contributed mainly to the plasma clearance. Only liver and the kidneys, however, were responsible for the uptake and subsequent degradation of BM 06.022 and contributed for 75% to the catabolism of BM 06.022. BM 06.022 was degraded in the lysosomal compartment of both organs. Parenchymal liver cells were responsible for 70% of the liver uptake of BM 06.022. BM 06.022 associated rapidly to isolated rat parenchymal liver cells and was subsequently degraded in the lysosomal compartment of these cells. BM 06.022 bound with low-affinity to the parenchymal liver cells (550 nM) and the binding of BM 06.022 could be displaced by t-PA (IC50 5.6 nM), indicating that the low-density lipoprotein receptor-related protein (LRP) could be involved in the binding of BM 06.022. GST-RAP, which is an inhibitor of LRP, could in vivo significantly inhibit the uptake of BM 06.022 in the liver.It is concluded that BM 06.022 is metabolized primarily in the liver and the kidneys. These organs take up and degrade BM 06.022 in the lysosomes. The uptake mechanism of BM 06.022 in the kidneys is unknown, while LRP is responsible for a low-affinity binding and uptake of BM 06.022 in parenchymal liver cells.

SiRNA technology, the gene therapy of the future?

2008 ◽  
Vol 149 (4) ◽  
pp. 153-159 ◽  
Author(s):  
Zsuzsanna Rácz ◽  
Péter Hamar
Keyword(s):  
Gene Therapy ◽  
Short Interfering Rna ◽  
The Future ◽  
Interfering Rna

A genetikában új korszak kezdődött 17 éve, amikor a petúniában felfedezték a koszuppressziót. Később a koszuppressziót azonosították a növényekben és alacsonyabb rendű eukariótákban megfigyelt RNS-interferenciával (RNSi). Bár a növényekben ez ősi vírusellenes gazdaszervezeti védekezőmechanizmus, emlősökben az RNSi élettani szerepe még nincs teljesen tisztázva. Az RNSi-t rövid kettős szálú interferáló RNS-ek (short interfering RNA, siRNS) irányítják. A jelen cikkben összefoglaljuk az RNSi történetét és mechanizmusát, az siRNS-ek szerkezete és hatékonysága közötti összefüggéseket, a célsejtbe való bejuttatás virális és nem virális módjait. Az siRNS-ek klinikai alkalmazásának legfontosabb akadálya az in vivo alkalmazás. Bár a hidrodinamikus kezelés állatokban hatékony, embereknél nem alkalmazható. Lehetőséget jelent viszont a szervspecifikus katéterezés. A szintetizált siRNS-ek ismert mellékhatásait szintén tárgyaljuk. Bár a génterápia ezen új területén számos problémával kell szembenézni, a sikeres in vitro és in vivo kísérletek reményt jelentenek emberi betegségek siRNS-sel történő kezelésére.

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