ABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression variation and significance of Skp2 and p27(kip1) during the proliferation of lymphoma cell line Jurkat cells.</p><p><b>METHODS</b>The binding of p27(kip1) and Skp2 in Jurkat cells were detected by immunoprecipitation. Jurkat cells were treated with serum starvation and release synchronization. The expression variation and subcellular localization of p27(kip1) and Skp2 were detected by subcellular fractionation, Western blot and double immunofluorescence labelling.</p><p><b>RESULTS</b>The results of immunoprecipitation suggested that p27(kip1) and Skp2 could bind each other in Jurkat cells. During the proliferation of Jurkat cells, the protein expression of p27(kip1) decreased and intranuclear p27(kip1) decreased significantly, while the Skp2 protein increased and cytoplasmic Skp2 increased significantly.</p><p><b>CONCLUSION</b>During the proliferation of Jurkat cells, the increased cytoplasmic synthesis of Skp2 may speed up p27(kip1) degradation via the ubiquitin-proteasome pathway, then intranuclear p27(kip1) decreases significantly, leading to an increased cell cycling activity.</p>
Subject(s)
Humans , Cell Nucleus , Metabolism , Cell Proliferation , Cyclin-Dependent Kinase Inhibitor p27 , Metabolism , Cytoplasm , Metabolism , Jurkat Cells , Lymphoma, B-Cell , Metabolism , Pathology , Protein Binding , S-Phase Kinase-Associated Proteins , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression of nuclear export factor CRM1, Ser10-phosphorylated p27 and p27 in human gliomas.</p><p><b>METHODS</b>The expression of CRM1, Ser10-phosphorylated p27 and p27 were investigated in 70 cases of human gliomas and 10 specimens of the normal brain tissue by immunohistochemical technique and Western blot.</p><p><b>RESULTS</b>There were significant differences on the expression levels of CRM1, Ser10-phosphorylated p27 and p27 among normal brain tissue, gliomas of grades II and gliomas of grades III plus IV (P < 0.01). The expression of CRM1 in gliomas was inversely correlated with the expression of p27 (r(s) = -0.727, P < 0.01) and positively correlated with the expression of Ser10-phosphorylated p27 (r(s) = 0.954, P < 0.01) and Ki-67 (r(s) = 0.799, P < 0.01). Moreover, the expression of Ser10-phosphorylated p27 was inversely correlated with p27 (r(s) = -0.744, P < 0.01) and positively correlated with Ki-67 (r(s) = 0.785, P < 0.01).</p><p><b>CONCLUSIONS</b>CRM1, through recognizing and binding with Ser10-phosphorylated p27, may promote moving of p27CRM1 from its original locating sites; act as a critical signaling component in the proliferative process of glioma cells and then, plays an important role in the development of gliomas.</p>
Subject(s)
Adolescent , Adult , Aged , Child , Humans , Middle Aged , Young Adult , Active Transport, Cell Nucleus , Genetics , Brain Neoplasms , Genetics , Metabolism , Cyclin-Dependent Kinase Inhibitor p27 , Genetics , Metabolism , Gene Expression Regulation, Neoplastic , Glioma , Genetics , Metabolism , Nuclear Export Signals , Genetics , Phosphorylation , PrognosisABSTRACT
<p><b>OBJECTIVE</b>To study the effect of all-trans retinoic acid (ATRA) on U937 cell growth and its mechanism.</p><p><b>METHODS</b>Cell cycle was detected by flow cytometry (FCM), expressions of cell cycle associated protein and the p27 related protein were detected by Western blot. The binding of P27 and Skp2 was detected by immunoprecipitation.</p><p><b>RESULTS</b>FCM displayed that ATRA could inhibit the proliferation of U937 cells. At 72 h on 1 micromol/L ATRA treatment, 72% of the cells were arrested at G0/G1 phase. Western blot displayed that ATRA could decrease the expression of cyclin A, up-regulate the expression of p21 and p27, and down-regulate the expression of p27 related proteins Skp2. p27 could bind with Skp2 in U937 cells as detected by immunoprecipitation.</p><p><b>CONCLUSION</b>ATRA may arrest the proliferation of U937 cells through the reduction of Skp2 expression, and finally the induction of the accumulation of p27.</p>
Subject(s)
Humans , Cell Cycle , Cell Proliferation , Cyclin-Dependent Kinase Inhibitor p27 , Metabolism , S-Phase Kinase-Associated Proteins , Metabolism , Tretinoin , Pharmacology , U937 CellsABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression and relationship of p27(kip1) and its related molecules Jab1 and CRM1 during proliferation of lymphoma cells U937.</p><p><b>METHODS</b>U937 cells were treated with serum starvation and release, and the effects of these treatments on the cell growth was tested with cell number counting. The expression and localization of p27(kip1), Jab1 and CRM1 in U937 cells were detected by Western blot, double immunolabelling and laser scanning confocal microscopy.</p><p><b>RESULTS</b>The growth of U937 cells was blocked by serum starvation. The total protein of p27(kip1) was increased while Ser10-phosphorylated p27(kip1) -related molecules Jab1 and CRM1 were decreased. Meanwhile, the location of p27(kip1) was changed from cytoplasm into nuclei. After serum release, the location of p27(kip1) expression reappeared in the cytoplasm again.</p><p><b>CONCLUSION</b>During the proliferation process of lymphoma U937 cells, Jab1 and CRM1 may influence the location and expression of p27kip1, and may participate in regulation of growth of NHL cells.</p>
Subject(s)
Humans , COP9 Signalosome Complex , Cell Culture Techniques , Cell Nucleus , Metabolism , Cell Proliferation , Culture Media, Serum-Free , Pharmacology , Cyclin-Dependent Kinase Inhibitor p27 , Metabolism , Cytoplasm , Metabolism , Intracellular Signaling Peptides and Proteins , Metabolism , Karyopherins , Metabolism , Peptide Hydrolases , Metabolism , Receptors, Cytoplasmic and Nuclear , Metabolism , U937 CellsABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression and relationship of p27(kip1) and its nuclear export factor Jab1 during proliferation process of lymphoma cell.</p><p><b>METHODS</b>Jurkat and Raji cells were treated with serum starvation and then serum release. The protein and mRNA expression of p27(kip1), Jab1 in the cells were detected by Western blot and RT-PCR respectively. LMB were used for stimulating Jurkat cells during their proliferation process, and then the expression changes of p27(kip1) and Jab1 were detected. An eukaryotic expression plasmid(pcDNA3. 1-myc) containing Jab1 was constructed. Jurkat cell were transfected in vitro with or without pcDNA3. 1-myc-Jab1. Double immunolabelling was used to identify the localization of p27(kip1). Immunoprecipitation was used to detect the combination of p27(kip1) and Jab1.</p><p><b>RESULTS</b>The growth of Jurkat and Raji cells were blocked by serum starvation. The total protein amount of p27(kip1) increased while that of Jab1 decreased. The reverse changes were happened after serum release, but the mRNA expression of p27(kip1) has no significant change. LMB could inhibit the cell proliferation caused by serum release. The expression of p27(kip1) was up-regulated and Jab1 down-regulated when Jurkat cells were treated with LMB. After pcDNA3. 1-myc-Jab1 infected Jurkat cells for 48 h, the distribution of p27(kip1) was translocated from nucleus into cytoplasma. p27(kip1) and Jab1 could form compound in Jurkat and Raji cells detected by Immunoprecipitation.</p><p><b>CONCLUSION</b>Jab1 may influence the location and expression of p27(kip1) through integrating with p27(kip1), and then participates in regulating the growth of NHL cell through interfering with the function of p27(kip1).</p>
Subject(s)
Humans , COP9 Signalosome Complex , Cyclin-Dependent Kinase Inhibitor p27 , Metabolism , Intracellular Signaling Peptides and Proteins , Metabolism , Jurkat Cells , Peptide Hydrolases , Metabolism , Plasmids , RNA, Messenger , Metabolism , TransfectionABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression and correlation of Skp2 and p27kipl in non-Hodgkin's lymphoma.</p><p><b>METHODS</b>The expression of Skp2, p27(kip1) and Ki-67 (the proliferation index)were detected in sections of 92 cases of non-Hodgkin's lymphoma (NHL) and 14 cases of reactive lymph nodes by immunohistochemistry and histopathology. The expression of Skp2 and p27(kip1) in 4 NHL cell lines were detected by Western blot.</p><p><b>RESULTS</b>The expression of Skp2 in NHL cases were significantly higher than that in reactive lymph nodes (except the germinal centers), positively correlated with proliferation activity, and an increasing tumor aggressiveness was associated with the increased expression of Skp2. The expression of p27(kip1) protein in NHL cases were significantly lower than that in reactive lymph nodes (except the germinal centers), negatively correlated with proliferation activity, and an increasing tumor aggressiveness was associated with decreased expression of p27(kip1). The statistical analysis indicated that there was no obvious correlation between Skp2 and p27(kip1) expression in NHL tissues.</p><p><b>CONCLUSION</b>The higher expression of Skp2 and lower expression of p27(kip1) in NHL tissues may play a role in the tumorigenesis and development of NHL.</p>