casein-kinase-ii and benzyloxycarbonylleucyl-leucyl-leucine-aldehyde

HDAC3 is involved in deacetylation of histone and non-histone proteins, having a key role in the regulation of gene transcription and also in the process of tumorgenesis. However, how HDAC3 is regulated in cancer remains largely unclear. Here, we showed that PIWIL2 can interact with HDAC3, leading to stabilization of HDAC3 from ubiquitin-mediated degradation by competitive association with E3 ubiquitin ligase Siah2. Furthermore, we found that expression of PIWIL2 enhanced HDAC3 activity via CK2α. PIWIL2 facilitated the interaction between HDAC3 and CK2α, thus exhibiting a promotion on the HDAC3 phosphorylation by CK2α. Further work showed that PIWIL2 could promote cell proliferation and suppress cell apoptosis via regulating HDAC3. Our present study firstly revealed that PIWIL2 can play a role in HDAC3-mediated epigenetic regulation on cancer cell proliferation and apoptosis. These findings provide a novel insight into the roles of PIWIL2 in tumorigenesis.

Topics: Antibodies; Apoptosis; Argonaute Proteins; Casein Kinase II; Cell Line, Tumor; Female; Histone Deacetylases; Humans; Leupeptins; Nuclear Proteins; Phosphorylation; Protein Binding; Proteolysis; RNA Interference; RNA, Small Interfering; Ubiquitin-Protein Ligases; Ubiquitination; Uterine Cervical Neoplasms

Pancreatic cancer (PC) evades immune destruction by favoring the development of regulatory T cells (Tregs) that inhibit effector T cells. The transcription factor Ikaros is critical for lymphocyte development, especially T cells. We have previously shown that downregulation of Ikaros occurs as a result of its protein degradation by the ubiquitin-proteasome system in our Panc02 tumor-bearing (TB) mouse model. Mechanistically, we observed a deregulation in the balance between Casein Kinase II (CK2) and protein phosphatase 1 (PP1), which suggested that increased CK2 activity is responsible for regulating Ikaros' stability in our model. We also showed that this loss of Ikaros expression is associated with a significant decrease in CD4+ and CD8+ T cell percentages but increased CD4+CD25+ Tregs in TB mice. In this study, we evaluated the effects of the dietary flavonoid apigenin (API), on Ikaros expression and T cell immune responses. Treatment of splenocytes from naïve mice with (API) stabilized Ikaros expression and prevented Ikaros downregulation in the presence of murine Panc02 cells in vitro, similar to the proteasome inhibitor MG132. In vivo treatment of TB mice with apigenin (TB-API) improved survival, reduced tumor weights and prevented splenomegaly. API treatment also restored protein expression of some Ikaros isoforms, which may be attributed to its moderate inhibition of CK2 activity from splenocytes of TB-API mice. This partial restoration of Ikaros expression was accompanied by a significant increase in CD4+ and CD8+ T cell percentages and a reduction in Treg percentages in TB-API mice. In addition, CD8+ T cells from TB-API mice produced more IFN-γ and their splenocytes were better able to prime allogeneic CD8+ T cell responses compared to TB mice. These results provide further evidence that Ikaros is regulated by CK2 in our pancreatic cancer model. More importantly, our findings suggest that API may be a possible therapeutic agent for stabilizing Ikaros expression and function to maintain T cell homeostasis in murine PC.

Topics: Animals; Apigenin; Casein Kinase II; Cell Line, Tumor; Down-Regulation; Female; Homeostasis; Ikaros Transcription Factor; Leupeptins; Lymphocyte Culture Test, Mixed; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Pancreatic Neoplasms; Proteasome Inhibitors; Protein Phosphatase 1; T-Lymphocytes; Tumor Suppressor Proteins

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has emerged as a promising antineoplastic agent because of its ability to selectively kill tumoral cells. However, some cancer cells are resistant to TRAIL-induced apoptosis. We have previously demonstrated that in endometrial carcinoma cells such resistance is caused by elevated FLICE-inhibitory protein (FLIP) levels. The present study focuses on the mechanisms by which FLIP could be modulated to sensitize endometrial carcinoma cells to TRAIL-induced apoptosis. We find that inhibition of casein kinase (CK2) sensitizes endometrial carcinoma cells to TRAIL- and Fas-induced apoptosis. CK2 inhibition correlates with a reduction of FLIP protein, suggesting that CK2 regulates resistance to TRAIL and Fas by controlling FLIP levels. FLIP downregulation correlates with a reduction of mRNA and is prevented by addition of the MG-132, suggesting that CK2 inhibition results in a proteasome-mediated degradation of FLIP. Consistently, forced expression of FLIP restores resistance to TRAIL and Fas. Moreover, knockdown of either FADD or caspase-8 abrogates apoptosis triggered by inhibition of CK2, indicating that CK2 sensitization requires formation of functional DISC. Finally, because of the possible role of both TRAIL and CK2 in cancer therapy, we demonstrate that CK2 inhibition sensitizes primary endometrial carcinoma explants to TRAIL apoptosis. In conclusion, we demonstrate that CK2 regulates endometrial carcinoma cell sensitivity to TRAIL and Fas by regulating FLIP levels.

Topics: Antineoplastic Agents; Apoptosis; Casein Kinase II; CASP8 and FADD-Like Apoptosis Regulating Protein; Cell Line; Drug Resistance, Neoplasm; Endometrial Neoplasms; fas Receptor; Female; Gene Expression Regulation, Neoplastic; Humans; Leupeptins; Neoplasm Proteins; Proteasome Endopeptidase Complex; TNF-Related Apoptosis-Inducing Ligand; Tumor Cells, Cultured

The PML tumor suppressor controls key pathways for growth suppression, induction of apoptosis, and cellular senescence. PML loss occurs frequently in human tumors through unknown posttranslational mechanisms. Casein kinase 2 (CK2) is oncogenic and frequently upregulated in human tumors. Here we show that CK2 regulates PML protein levels by promoting its ubiquitin-mediated degradation dependent on direct phosphorylation at Ser517. Consequently, PML mutants that are resistant to CK2 phosphorylation display increased tumor-suppressive functions. In a faithful mouse model of lung cancer, we demonstrate that Pml inactivation leads to increased tumorigenesis. Furthermore, CK2 pharmacological inhibition enhances the PML tumor-suppressive property in vivo. Importantly, we found an inverse correlation between CK2 kinase activity and PML protein levels in human lung cancer-derived cell lines and primary specimens. These data identify a key posttranslational mechanism that controls PML protein levels and provide therapeutic means toward PML restoration through CK2 inhibition.

Topics: Amino Acid Sequence; Amino Acid Substitution; Animals; Apoptosis; Carcinoma, Non-Small-Cell Lung; Casein Kinase II; Cell Line; Cell Line, Transformed; Cell Line, Tumor; Enzyme Activation; Enzyme Inhibitors; Genes, Tumor Suppressor; Green Fluorescent Proteins; Hemagglutinins; Humans; Leupeptins; Lung Neoplasms; Mice; Mice, Transgenic; Molecular Sequence Data; Neoplasm Proteins; NIH 3T3 Cells; Nuclear Proteins; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Promyelocytic Leukemia Protein; Proteasome Endopeptidase Complex; Protein Structure, Tertiary; Protein Subunits; RNA, Small Interfering; Sequence Deletion; Serine; Sorbitol; Transcription Factors; Transcriptional Activation; Triazoles; Tumor Suppressor Proteins; Ubiquitin

In eukaryotic cells, protein kinase CKII is required for progression through the cell division cycle. We recently reported that CKBBP1/SAG/ROC2/Rbx2 associates with the beta-subunit of CKII and is phosphorylated by purified CKII in the presence of ATP in vitro. In this report, we demonstrate that CKBBP1 is efficiently phosphorylated in vitro by purified CKII in the presence of GTP and by heparin-sensitive protein kinase in HeLa cell extract. Mutational analysis indicates that CKII phosphorylates threonine at residue 10 within CKBBP1. Furthermore, CKBBP1 is phosphorylated in vivo and threonine to alanine mutation at residue 10 abrogates the phosphorylation of CKBBP1 observed in vivo, indicating that CKII is a major kinase that is responsible for in vivo phosphorylation of CKBBP1. As compared with the wild-type CKBBP1 or CKBBP1T10E (in which threonine 10 is replaced by glutamate), overexpression of nonphosphorylatable CKBBP1 (CKBBP1T10A) results in accumulation of IkappaBalpha and p27Kip1. Experiments using proteasome inhibitor MG132 and CKII inhibitor 5,6-dichloro-1-beta-d-ribofuranosylbenzimidazole suggest that the accumulation of IkappaBalpha and p27Kip1 results primarily from the reduction of proteasomal degradation in cells expressing CKBBP1T10A, and that CKII-mediated CKBBP1 phosphorylation is required for efficient degradation of IkappaBalpha and p27Kip1. Overexpression of CKBBP1T10A in HeLa cells suppresses cell proliferation and causes accumulation of G1/G0 peak of the cell cycle. Taken together, our results indicate that CKII may control IkappaBalpha and p27Kip1 degradation and thereby G1/S phase transition through the phosphorylation of threonine 10 within CKBBP1.

Topics: Alanine; Casein Kinase II; Cell Cycle Proteins; Cell Division; Cyclin-Dependent Kinase Inhibitor p27; Cysteine Endopeptidases; Enzyme Inhibitors; G1 Phase; Gene Expression; Guanosine Triphosphate; HeLa Cells; Humans; I-kappa B Proteins; Leupeptins; Multienzyme Complexes; Mutagenesis, Site-Directed; NF-KappaB Inhibitor alpha; Phosphorylation; Proteasome Endopeptidase Complex; Protein Serine-Threonine Kinases; RNA; RNA-Binding Proteins; S Phase; Structure-Activity Relationship; Threonine; Transfection; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases