Subunit Of CK2 Research Articles

Rational drug-design approach supported with thermodynamic studies \u2014 a peptide leader for the efficient bi-substrate inhibitor of protein kinase CK2

Numerous inhibitors of protein kinases act on the basis of competition, targeting the ATP binding site. In this work, we present a procedure of rational design of a bi-substrate inhibitor, complemented with biophysical assays. The inhibitors of this type are commonly engineered by combining ligands carrying an ATP-like part with a peptide or peptide-mimicking fragment that determines specificity. Approach presented in this paper led to generation of a specific system for independent screening for efficient ligands and peptides, by means of thermodynamic measurements, that assessed the ability of the identified ligand and peptide to combine into a bi-substrate inhibitor. The catalytic subunit of human protein kinase CK2 was used as the model target. Peptide sequence was optimized using peptide libraries [KGDE]-[DE]-[ST]-[DE]3–4-NH2, originated from the consensus CK2 sequence. We identified KESEEE-NH2 peptide as the most promising one, whose binding affinity is substantially higher than that of the reference RRRDDDSDDD peptide. We assessed its potency to form an efficient bi-substrate inhibitor using tetrabromobenzotriazole (TBBt) as the model ATP-competitive inhibitor. The formation of ternary complex was monitored using Differential Scanning Fluorimetry (DSF), Microscale Thermophoresis (MST) and Isothermal Titration Calorimetry (ITC).

The effect of ionizing radiation on the subcellular localization and kinase activity of protein kinase CK2 in human non-small cell lung cancer cells

Protein kinase CK2 is a ubiquitously expressed kinase in eukaryotes, which is known to phosphorylate many protein substrates. Because CK2 is involved in the regulation of various signaling pathways, we wondered whether CK2 participated in the regulation of ionizing radiation (IR) induced biological process. In this study, we investigated the effect of IR on the subcellular localization and kinase activity in human non-small cell lung cancer (NSCLC) cells. Immunofluorescent results showed that CK2 subunits shuttle into the nucleus mostly beginning 1 h after IR and lasting more than 6 h. We also conducted in vitro kinase assay and observed an increase in CK2 kinase activity at 6 h after IR. Furthermore, an increase in S phase was observed at 6 h after IR. Colony formation assay results demonstrated that CK2 inhibitor CX-4945 significantly enhanced the effect of irradiation in NSCLC cells. These results indicated that CK2 may be implicated in the regulation of IR-induced biological process.

Abstract 240: Therapeutic targeting of protein kinase ck2 in mycn-driven neuroblastoma

Abstract Amplification of the MYCN oncogene occurs in about 30-40% of advanced neuroblastoma cases and is the most reliable predictor of poor prognosis. Protein kinase CK2 (casein kinase II) is a highly conserved serine/threonine kinase that can be effectively inhibited by small molecules. In fact, CX-4945, a potent, specific, ATP-competitive CK2 inhibitor, is currently tested in clinical trials to treat multiple cancers. However, the role of CK2 in MYCN-driven neuroblastoma remains unknown. To determine the relevance of CK2 in MYCN-driven neuroblastoma pathogenesis, we analyzed published microarray data and compared the expression of the CK2 subunits in primary neuroblastoma patients samples. We found that both CK2α and CK2α’ are upregulated in neuroblastoma patients samples with MYCN amplification, compared to the ones without MYCN amplification. To determine the effect of CK2 inhibition on neuroblastoma cells, we treated a panel of human neuroblastoma cell lines with the CK2 inhibitor CX-4945 and short hairpin RNA specifically targeting CK2α. CK2 inhibitor CX-4945 suppressed human neuroblastoma cell growth, mainly through inducing apoptosis, especially in the MYCN-amplified cells. More over, CK2α knockdown also induced cell apoptosis. In-vitro assay indicated that CK2 inhibition through either genetic or pharmacological approaches effectively reduced cell growth of human neuroblastoma cells primarily through induction of apoptosis. 14 days in-vivo treatment, CX-4945 significantly decreased tumor growth in mouse xenografts by subcutaneously implanting human KELLY neuroblastoma cells, but didn’t affect their body weight. These studies characterize the role of CK2 in neuroblastoma development, and provide valuable information about the relevance of CK2 as a target in MYCN-driven neuroblastoma. Note: This abstract was not presented at the meeting. Citation Format: Haiwei Lian, Ning Shen, Yun Zhou, Dun Li, Hui Feng. Therapeutic targeting of protein kinase ck2 in mycn-driven neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 240.

Magnaporthe oryzae CK2 Accumulates in Nuclei, Nucleoli, at Septal Pores and Forms a Large Ring Structure in Appressoria, and Is Involved in Rice Blast Pathogenesis.

Magnaporthe oryzae (Mo) is a model pathogen causing rice blast resulting in yield and economic losses world-wide. CK2 is a constitutively active, serine/threonine kinase in eukaryotes, having a wide array of known substrates, and involved in many cellular processes. We investigated the localization and role of MoCK2 during growth and infection. BLAST search for MoCK2 components and targeted deletion of subunits was combined with protein-GFP fusions to investigate localization. We found one CKa and two CKb subunits of the CK2 holoenzyme. Deletion of the catalytic subunit CKa was not possible and might indicate that such deletions are lethal. The CKb subunits could be deleted but they were both necessary for normal growth and pathogenicity. Localization studies showed that the CK2 holoenzyme needed to be intact for normal localization at septal pores and at appressorium penetration pores. Nuclear localization of CKa was however not dependent on the intact CK2 holoenzyme. In appressoria, CK2 formed a large ring perpendicular to the penetration pore and the ring formation was dependent on the presence of all CK2 subunits. The effects on growth and pathogenicity of deletion of the b subunits combined with the localization indicate that CK2 can have important regulatory functions not only in the nucleus/nucleolus but also at fungal specific structures such as septa and appressorial pores.

Diacritic Binding of an Indenoindole Inhibitor by CK2α Paralogs Explored by a Reliable Path to Atomic Resolution CK2α' Structures.

CK2α and CK2α′ are the two isoforms of the catalytic subunit of human protein kinase CK2, an important target for cancer therapy. They have similar, albeit not identical functional and structural properties, and were occasionally reported to be inhibited with distinct efficacies by certain ATP-competitive ligands. Here, we present THN27, an indeno[1,2-b]indole derivative, as a further inhibitor with basal isoform selectivity. The selectivity disappears when measured using CK2α/CK2α′ complexes with CK2β, the regulatory CK2 subunit. Co-crystal structures of THN27 with CK2α and CK2α′ reveal that subtle differences in the conformational variability of the interdomain hinge region are correlated with the observed effect. In the case of CK2α′, a crystallographically problematic protein so far, this comparative structural analysis required the development of an experimental strategy that finally enables atomic resolution structure determinations with ab initio phasing of potentially any ATP-competitive CK2 inhibitor and possibly many non-ATP-competitive ligands as well bound to CK2α′.

A proteomics analysis of CK2β(-/-) C2C12 cells provides novel insights into the biological functions of the non-catalytic β subunit.

The acronym CK2 (derived from the misnomer 'casein kinase-2') denotes a pleiotropic acidophilic protein kinase implicated in a plethora of cellular functions, whose abnormally high expression correlates with malignancy. CK2 holoenzyme is composed of two catalytic (α and/or α') and two noncatalytic β-subunits. The β-subunits are not responsible for either activation or inactivation of the catalytic ones. Hence, to gain additional information about the roles of the individual CK2 subunits, we have generated C2C12 myoblasts entirely devoid either of both catalytic subunits, or of the β-subunit. Here, we show that while CK2α/α'(-/-) cells grow similarly to wild-type cells, the growth of CK2β(-/-) cells is severely impaired, consistent with the hypothesis that not all cellular functions of the β-subunit are mediated by CK2 holoenzyme. To get a deeper insight into the functional implications of the β-subunit, a quantitative proteomics study of CK2β(-/-) cells was performed, leading to the identification and quantification of more than 1200 proteins. Of these, 187 showed a significantly altered expression (fold change ≥ 1.5 or ≤ -1.5) as compared to wild-type cells. A functional analysis of these proteins discloses the implication of CK2β in many processes, for example, cell cycle, proliferation, transport, metabolic processes, etc., and in some of which the catalytic subunits of CK2 do not seem to play a relevant role. On the other hand, the pool of ecto-CK2 is not apparently affected by the lack of the β-subunit. Collectively, our data corroborate the concept that the cellular functions of the β-subunit of CK2 are partially independent of CK2 holoenzyme.

Design of CK2β-Mimicking Peptides as Tools To Study the CK2α/CK2β Interaction in Cancer Cells.

The ubiquitously expressed Ser/Thr kinase CK2 is a key regulator in a variety of key processes in normal and malignant cells. Due to its distinctive anti-apoptotic and tumor-driving properties, elevated levels of CK2 have frequently been found in tumors of different origin. In recent years, development of CK2 inhibitors has largely been focused on ATP-competitive compounds; however, targeting the CK2α/CK2β interface has emerged as a further concept that might avoid selectivity issues. To address the CK2 subunit interaction site, we have synthesized halogenated CK2β-mimicking cyclic peptides modified with the cell-penetrating peptide sC18 to mediate cellular uptake. We investigated the binding of the resulting chimeric peptides to recombinant human CK2α using a recently developed fluorescence anisotropy assay. The iodinated peptide sC18-I-Pc was identified as a potent CK2α ligand (Ki =0.622 μm). It was internalized in cells to a high extent and exhibited significant cytotoxicity toward cancerous HeLa cells (IC50 =37 μm) in contrast to non-cancerous HEK-293 cells. The attractive features and functionalities of sC18-I-Pc offer the opportunity for further improvement.

Up-Regulation of the Alpha Prime Subunit of Protein Kinase CK2 as a Marker of Fast Proliferation in GL261 Cultured Cells.

Glioblastoma (GB) is the most prevalent malignant primary brain tumor in adults. The preclinical glioblastoma model GL261 is widely used for investigating new therapeutic strategies. GL261 cultured cells are used for assessing preliminary in vitro data for this model although very little is known about the molecular characteristics of this cell line. Protein Kinase CK2 is a pleiotropic serine-threonine kinase and its inhibition may be a promising therapeutic strategy for GB treatment. In our group we follow treatment response with CK2 inhibitors in vivo using the GL261 murine model. For that, it is of our interest to assess the differential expression of α, α', β CK2 subunits as well as CK2 activity in the GL261 GB model. CK2α' expression changed along the growth curve of GL261 cells, undergoing downregulation in postconfluent phase cells, whereas CK2α and CK2β expression remained essentially unchanged. Furthermore, a marked decrease in CK2 activity in slowly proliferating postconfluent phase GL261 cells was observed. Finally, CK2α' expression in orthotopic GL261 tumors was intermediate between CK2α' expression found in cultured cells in exponentially growing or postconfluent phase, reflecting the heterogeneous nature of GL261 tumours growing in vivo. The results obtained suggest that, in the GL261 cell line, CK2α' could play a specific role in highly proliferative cells. Also, the decrease in CK2 activity in slowly proliferating GL261 cells could imply a differential susceptibility to subunit-specific CK2 inhibitors in this cell line, although further studies are needed to confirm this hypothesis.

Protein kinase CK2α subunits constitutively activate ABA signaling in Arabidopsis

ABSTRACTProtein kinase CK2 (formerly known as casein kinase II), a Ser/Thr protein kinase highly conserved in eukaryotes, is essential for cell survival by regulating a wide range of plant growth, development, and stress responses. A growing body of evidence has shown a link between CK2 and abscisic acid (ABA) signaling in response to abiotic stress. However, the roles of CK2 subunits in ABA signaling remain unclear in plants. Our recent work in Arabidopsis thaliana has revealed that CK2α and CK2β subunits inversely modulate ABA signal output. Here, we examine the roles of CK2αs, by assessing how CK2αs affect ABA signaling. Together with the previous findings, our mutant and transient expression analyses demonstrate that CK2αs positively modulate ABA signaling through the core ABA signaling pathway in the presence of ABA, though the positive effect of CK2αs are much smaller than that of core ABA signaling components in ABA response. In addtion, our current and previous findings also suggest that CK2αs play a role in maintaining constitutively active ABA signaling even in the absence of ABA independently of the core ABA signaling pathway. Thus, we found that CK2αs constitutively activate ABA signaling in the presence or absence of ABA in a different manner in Arabidopsis plants.

CX-4945 Induces Methuosis in Cholangiocarcinoma Cell Lines by a CK2-Independent Mechanism.

Cholangiocarcinoma is a disease with a poor prognosis and increasing incidence and hence there is a pressing unmet clinical need for new adjuvant treatments. Protein kinase CK2 (previously casein kinase II) is a ubiquitously expressed protein kinase that is up-regulated in multiple cancer cell types. The inhibition of CK2 activity using CX-4945 (Silmitasertib) has been proposed as a novel treatment in multiple disease settings including cholangiocarcinoma. Here, we show that CX-4945 inhibited the proliferation of cholangiocarcinoma cell lines in vitro. Moreover, CX-4945 treatment induced the formation of cytosolic vacuoles in cholangiocarcinoma cell lines and other cancer cell lines. The vacuoles contained extracellular fluid and had neutral pH, features characteristic of methuosis. In contrast, simultaneous knockdown of both the α and α′ catalytic subunits of protein kinase CK2 using small interfering RNA (siRNA) had little or no effect on the proliferation of cholangiocarcinoma cell lines and failed to induce the vacuole formation. Surprisingly, low doses of CX-4945 increased the invasive properties of cholangiocarcinoma cells due to an upregulation of matrix metallopeptidase 7 (MMP-7), while the knockdown of CK2 inhibited cell invasion. Our data suggest that CX-4945 inhibits cell proliferation and induces cell death via CK2-independent pathways. Moreover, the increase in cell invasion brought about by CX-4945 treatment suggests that this drug might increase tumor invasion in clinical settings.

CK2 Controls Th17 and Regulatory T Cell Differentiation Through Inhibition of FoxO1.

Growing evidence demonstrates that the highly conserved serine/threonine kinase CK2 promotes Th17 cell differentiation while suppressing the generation of Foxp3+ regulatory T cells (Tregs); however, the exact mechanism by which CK2 regulates the Th17/Treg axis remains unclear. CK2 can be composed of three distinct subunits: two catalytic subunits, CK2α and CK2α', and the regulatory subunit CK2β. We generated mice that lack the major catalytic subunit of CK2, CK2α, specifically in mature T cells using the distal Lck-Cre (CK2α-/-). Importantly, CK2α deficiency resulted in a significant decrease in the overall kinase activity of CK2. Further, CK2α deficiency resulted in a significant defect in Th17 cell polarization and a reciprocal increase in Tregs both in vitro and in vivo in the context of autoimmune neuroinflammation. The transcription factor forkhead box protein O1 (FoxO1) directly inhibits Th17 cell differentiation and is essential for the generation of Tregs. CK2α-/- CD4+ T cells exhibit less phosphorylated FoxO1 and a corresponding increase in the transcription of FoxO1-regulated genes. Treatment of CK2α-/- CD4+ T cells with the FoxO1 inhibitor AS1842856 or short hairpin RNA knockdown of FoxO1 is sufficient to rescue Th17 cell polarization. Through use of a genetic approach to target CK2 kinase activity, the current study provides evidence of a major mechanism by which CK2 regulates the Th17/Treg axis through the inhibition of FoxO1.

Abstract C44: Epigenetic regulation of CRLF2 oncogene expression by Casein Kinase II (CK2) signaling in B-cell acute lymphoblastic leukemia that occurs at high frequency in Hispanic children

Abstract B-cell acute lymphoblastic leukemia (B-ALL) occurs more frequently in Hispanic children as compared to non-Hispanic whites. The specific subtype of B-ALL that is caused by overexpression of CRLF2 (CRLF2 B-ALL), occurs 5 times more frequently in Hispanic children as compared to the others. Since this type of B-ALL is associated with poor prognosis, the death rate of B-ALL is 39% higher in Hispanic children than in non-Hispanic whites. Thus, understanding the molecular mechanisms that regulate CRLF2 expression in CRLF2 B-ALL is essential for the development of targeted therapy for this disease. Our previous work determined that transcription of CRLF2 is negatively regulated by a tumor-suppressor protein, Ikaros. Ikaros deregulation is a feature of over 80% of CRLF2 B-ALL. Here we present evidence that Ikaros-mediated repression of CRLF2 transcription in B-ALL in Hispanic children is regulated by Casein Kinase II (CK2). CK2 is an oncogenic kinase that is overexpressed in B-ALL. We have previously shown that CK2 can directly phosphorylate Ikaros and that phosphorylation by CK2 can impair Ikaros function as transcriptional regulator. We tested whether inhibition of CK2 affects the ability of Ikaros to regulate CRLF2 transcription. Molecular inhibition by shRNA that targets the catalytic subunit of CK2, as well as pharmacologic inhibition of CK2 by the specific inhibitor, CX-4945, resulted in reduced expression of CRLF2 as measured by qRT-PCR. This was associated with increased Ikaros binding to the CRLF2 promoter as measured by quantitative chromatin immunoprecipitation (qChIP). To determine whether Ikaros function is essential for CRLF2 repression following CK2 inhibition, we compared expression of CRLF2 in cells that have Ikaros knocked-down by shRNA vs. cells with control shRNA, following treatment with CX-4945. Results showed that Ikaros knockdown abolished the ability of CK2 inhibitors to repress transcription of CRLF2 in B-ALL. These results demonstrate that Ikaros is an essential component of CK2 signaling that regulates CRLF2 expression. Analysis of the epigenetic signature at the CRLF2 promoter performed by serial qChIP assays showed that increased Ikaros binding to the CRLF2 promoter, following CK2 inhibition, is associated with enrichment for the H3K9me3 histone modification, which is a marker of repressive chromatin. In conclusion, we demonstrate that expression of the CRLF2 oncogene in acute leukemia that disproportionally occurs in Hispanic children is epigenetically regulated by the CK2-Ikaros axis. In CRLF2 B-ALL, Ikaros-mediated repression of CRLF2 is impaired due to overexpression of CK2. Treatment of CRLF2 B-ALL with CK2 inhibitors restores Ikaros tumor suppressor function, resulting in CRLF2 repression. Results identified a signaling network that regulates CRLF2 expression and suggest that restoration of Ikaros activity with CK2 inhibitors can be a therapeutic approach for CRLF2 B-ALL to reduce the health disparity for Hispanic children with ALL. Supported by 1R01CA209829. Citation Format: Chunhua Song, Zheng Ge, Kimberly J. Payne, Sinisa Dovat. Epigenetic regulation of CRLF2 oncogene expression by Casein Kinase II (CK2) signaling in B-cell acute lymphoblastic leukemia that occurs at high frequency in Hispanic children [abstract]. In: Proceedings of the Tenth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2017 Sep 25-28; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2018;27(7 Suppl):Abstract nr C44.

New insights into donor directionality of mating-type switching in Schizosaccharomyces pombe.

Mating-type switching in Schizosaccharomyces pombe entails programmed gene conversion events regulated by DNA replication, heterochromatin, and the HP1-like chromodomain protein Swi6. The whole mechanism remains to be fully understood. Using a gene deletion library, we screened ~ 3400 mutants for defects in the donor selection step where a heterochromatic locus, mat2-P or mat3-M, is chosen to convert the expressed mat1 locus. By measuring the biases in mat1 content that result from faulty directionality, we identified in total 20 factors required for donor selection. Unexpectedly, these included the histone H3 lysine 4 (H3K4) methyltransferase complex subunits Set1, Swd1, Swd2, Swd3, Spf1 and Ash2, the BRE1-like ubiquitin ligase Brl2 and the Elongator complex subunit Elp6. The mutant defects were investigated in strains with reversed donor loci (mat2-M mat3-P) or when the SRE2 and SRE3 recombination enhancers, adjacent to the donors, were deleted or transposed. Mutants in Set1C, Brl2 or Elp6 altered balanced donor usage away from mat2 and the SRE2 enhancer, towards mat3 and the SRE3 enhancer. The defects in these mutants were qualitatively similar to heterochromatin mutants lacking Swi6, the NAD+-dependent histone deacetylase Sir2, or the Clr4, Raf1 or Rik1 subunits of the histone H3 lysine 9 (H3K9) methyltransferase complex, albeit not as extreme. Other mutants showed clonal biases in switching. This was the case for mutants in the NAD+-independent deacetylase complex subunits Clr1, Clr2 and Clr3, the casein kinase CK2 subunit Ckb1, the ubiquitin ligase component Pof3, and the CENP-B homologue Cbp1, as well as for double mutants lacking Swi6 and Brl2, Pof3, or Cbp1. Thus, we propose that Set1C cooperates with Swi6 and heterochromatin to direct donor choice to mat2-P in M cells, perhaps by inhibiting the SRE3 recombination enhancer, and that in the absence of Swi6 other factors are still capable of imposing biases to donor choice.

Casein kinase 2 \u03b1 and \u03b2 subunits inversely modulate ABA signal output in Arabidopsis protoplasts

Main conclusionOur transient gene expression analyses in Arabidopsis protoplasts support the view that CK2αs and CK2βs positively and negatively modulate ABRE-dependent gene expression, respectively.The phytohormone abscisic acid (ABA) regulates the expression of thousands of genes via ABA-responsive elements (ABREs), and has a crucial role in abiotic stress response. Casein kinase II (CK2), a conserved Ser/Thr protein kinase in eukaryotes, is essential for plant viability. Although the CK2 has been known as a tetrameric holoenzyme comprised of two catalytic α and two regulatory β subunits, each of the two types of subunits has been proposed to have independent functions. The Arabidopsis genome encodes four α subunits (CK2α1, CK2α2, CK2α3, CK2α4) and four β subunits (CK2β1, CK2β2, CK2β3, CK2β4). There is a growing body of evidence linking CK2 to ABA signaling and abiotic stress responses. However, the roles of each CK2 subunit in ABA signaling remain largely elusive. Using the transient expression system with the core ABA signaling components in Arabidopsis leaf mesophyll protoplasts, we show here that CK2α1 and CK2α2 (CK2α1/2) positively modulate ABRE-dependent gene expression as ABA signal output in ABA signaling, whereas all four CK2βs negatively modulate the ABRE-dependent gene expression mediated by subclass III SnRK2–AREB/ABF pathway and by CK2α1/2. These data indicate that CK2α1/2 and CK2βs positively and negatively modulate ABA signal output, respectively, suggesting that the quantitative balance of CK2 subunits determines the ABA signal output in plants. Given that CK2s act as pleiotropic enzymes involved in multiple developmental and stress–responsive processes, our findings suggest that CK2 subunits may be involved in integration and coordination of ABA-dependent and -independent signaling.

Casein kinase II phosphorylates the C-terminal region of Lif1 to promote the Lif1-Xrs2 interaction needed for non-homologous end joining

A DNA double strand break (DSB) is one of the most cytotoxic DNA lesions, but it can be repaired by non-homologous end joining (NHEJ) or by homologous recombination. The choice between these two repair pathways depends on the cell cycle stage. Although NHEJ constitutes a simple re-ligation reaction, the regulatory mechanism(s) controlling its activity has not been fully characterized. Lif1 is a regulatory subunit of the NHEJ-specific DNA ligase IV and interacts with Xrs2 of the MRX complex which is a key factor in DSB repair. Specifically, the C-terminal region of Lif1, which contains a CK2-specific phosphorylation motif, interacts with the FHA domain of Xrs2 during canonical- NHEJ (C-NHEJ). Herein, we show that Lif1 and Cka2, a catalytic subunit of yeast CK2, interact and that the C-terminal phosphorylation consensus motif in Lif1 is phosphorylated by recombinant CK2. These observations suggest that phosphorylation of Lif1 by CK2 at a DSB site promotes the Lif1-Xrs2 interaction and facilitates C-NHEJ.

Ablation of beta subunit of protein kinase CK2 in mouse oocytes causes follicle atresia and premature ovarian failure

Premature ovarian failure (POF), a major cause of female infertility, is a complex disorder, but the molecular mechanisms underlying the disorder are only poorly understood. Here we report that protein kinase CK2 contributes to maintaining follicular survival through PI3K/AKT pathway and DNA damage response pathway. Targeted deletion of CK2β in mouse oocytes from the primordial follicle stage resulted in female infertility, which was attributed to POF incurring by massive follicle atresia. Downregulated PI3K/AKT signaling was found after CK2β deletion, indicated by reduced level of phosphorylated AKT (S473, T308, and S129) and altered AKT targets related to cell survival. Further studies discovered that CK2β-deficient oocytes showed enhanced γH2AX signals, indicative of accumulative unrepaired DSBs, which activated CHK2-dependant p53 and p63 signaling. The suppressed PI3K/AKT signaling and failed DNA damage response signaling probably contribute to large-scale oocyte loss and eventually POF. Our findings provide important new clues for elucidating the mechanisms underlying follicle atresia and POF.

Crystal structures of human CK2α2 in new crystal forms arising from a subtle difference in salt concentration.

The catalytic subunits of protein kinase CK2 are classified into two subtypes: CK2α1 and CK2α2. CK2α1 is an attractive drug-discovery target for various diseases such as cancers and nephritis. CK2α2 is defined as an off-target of CK2α1 and is a potential target in the development of male contraceptive drugs. High-resolution crystal structures of both isozymes are likely to provide crucial clues for the design of selective inhibitors of CK2α1 and/or CK2α2. To date, several crystal structures of CK2α1 have been solved at high resolutions of beyond 1.5 Å. However, crystal structures of CK2α2 have barely achieved a low resolution of around 3 Å because of the formation of needle-shaped crystals. In this study, new crystal forms were exploited and one provided a crystal structure of CK2α2 at 1.89 Å resolution. This result, together with the structure of CK2α1, will assist in the development of highly selective inhibitors for both isozymes.

Ecto-protein kinase CK2, the neglected form of CK2.

Ecto-protein kinases, including protein kinase CK2 (former name, casein kinase 2), have been the focus of research for more than 30 years. At the beginning of the ecto-kinase research their identification was performed with substrates and inhibitors whose specificity under the current knowledge was rather limited. Since all currently known ecto-kinases, including ecto-CK2, have intracellular counterparts, one has to exclude that an ecto-localization originates from intracellular counterparts after cell damage. Protein kinase CK2 is involved in cellular key processes such as cell cycle progression, inhibition of apoptosis, DNA damage repair, differentiation and many other processes. CK2 is composed of two catalytic CK2α or CK2α' subunits and two non-catalytic CK2β subunits. Progress in the ecto-kinase and in particular ecto-CK2 studies was made with the use of transfected tagged CK2 subunits, which allowed to follow their individual transport and localization on the cell surface after transfection. Furthermore, immunofluorescence studies with antibodies against CK2 subunits as well as affinity chromatography with a binding partner of CK2 subunits have improved ecto-kinase research. The use of new and more specific inhibitors as well as of substrates, which do not cross the plasma membrane, have further improved the specificity for ecto-CK2. From the various substrates of ecto-CK2, it can be concluded that ecto-CK2 plays a role in Alzheimer disease, cell adhesion, platelet aggregation, immune response and cellular signalling. New tools and techniques, to study ecto-CK2 activity, are required to identify new substrates and thereby new functional implications for ecto-CK2.

Under-expression of CK2β subunit in ccRCC represents a complementary biomarker of p-STAT3 Ser727 that correlates with patient survival.

Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive subtype of renal cancer. STAT3 pathway is altered in these tumors and p-STAT3 Ser727 is an independent prognostic factor for ccRCC. Protein kinase CK2 is altered in different types of tumors and overexpression of CK2α is considered predictive of bad prognosis and metastatic risk. CK2 subunits analyses in ccRCC samples showed increased CK2α/α’ nuclear content in all cases, but decreased cytosolic CK2β (CK2βcyt) levels in the more advanced tumors. Stable downregulation of CK2β in renal proximal tubular (HK-2) and clear cell adenocarcinoma (786-O) cells triggered changes in E-cadherin, vimentin and Snail1 protein levels indicative of epithelial-to-mesenchymal transition (EMT), and increased HIF-α. Moreover, CK2β was required in order to observe STAT3 Ser727 phosphorylation in HK-2 but not in 786-O cells.We also observed that CK2β improved the prognostic value of p-STAT3 Ser727, as CK2βcyt>41 (median value) discriminates patients free of disease for a period of 10 years upon surgery, from those with CK2βcyt<41, when p-STAT3 Ser727levels are low.We conclude that CK2β down-regulation might represent a mechanism to support EMT and angiogenesis and that CK2βcyt levels are instrumental to refine prognosis of ccRCC patients with low p-STAT3 Ser727 levels.

Association of protein kinase CK2 inhibition with cellular radiosensitivity of non-small cell lung cancer

Protein kinase CK2 is a highly conserved protein Ser/Thr protein kinase and plays important roles in cell proliferation, protein translation and cell survival. This study investigated the possibility of using CK2 inhibition as a new approach for increasing the efficacy of radiotherapy in non-small cell lung cancer (NSCLC) and its underlying mechanisms. Kinase inhibition of CK2 was attempted either by using the specific CK2 inhibitor, Quinalizarin or by applying siRNA interference technology to silence the expression of the catalytic subunit of CK2 in A549 and H460 cells. The results showed that CK2α knockdown or Quinalizarin significantly enhanced the radiosensitivity of various NSCLC cells. The notable findings we observed after exposure to both CK2 inhibition and ionizing radiation (IR) were a prolonged delay in radiation-induced DNA double-strand breaks (DSB) repair, robust G2/M checkpoint arrest and increased apoptosis. In vivo studies further demonstrated that compared with each treatment alone, CK2 inhibition combined with IR reduced tumor growth in the H460 cell xenograft model. In conclusion, CK2 is a promising target for the enhancement of radiosensitivity in NSCLC.