Cell cycle-dependent gene networks for cell proliferation activated by nuclear CK2α complexes.

Nuclear expression of protein kinase CK2α is reportedly elevated in human carcinomas, but mechanisms underlying its variable localization in cells are poorly understood. This study demonstrates a functional connection between nuclear CK2 and gene expression in relation to cell proliferation. Growth stimulation of quiescent human normal fibroblasts and phospho-proteomic analysis identified a pool of CK2α that is highly phosphorylated at serine 7. Phosphorylated CK2α translocates into the nucleus, and this phosphorylation appears essential for nuclear localization and catalytic activity. Protein signatures associated with nuclear CK2 complexes reveal enrichment of apparently unique transcription factors and chromatin remodelers during progression through the G1 phase of the cell cycle. Chromatin immunoprecipitation-sequencing profiling demonstrated recruitment of CK2α to active gene loci, more abundantly in late G1 phase than in early G1, notably at transcriptional start sites of core histone genes, growth stimulus-associated genes, and ribosomal RNAs. Our findings reveal that nuclear CK2α complexes may be essential to facilitate progression of the cell cycle, by activating histone genes and triggering ribosomal biogenesis, specified in association with nuclear and nucleolar transcriptional regulators.

Intracellular localization of CK2α as a prognostic factor in invasive breast carcinomas.

Overexpression of the ubiquitous protein kinase, CK2α, has been reported in various human cancers. Here, we demonstrate that nuclear and nucleolar CK2α localization in invasive ductal carcinomas of the breast is a reliable predictor of poor prognosis. Cellular localization of CK2α in nuclei and nucleoli was analyzed immunohistochemically using surgical tissue blocks from 112 patients, who had undergone surgery without neoadjuvant chemotherapy. Clinical data collection and median follow-up period were for more than 5 y. In total, 93.8% of patients demonstrated elevated CK2α expression in nuclei and 36.6% of them displayed elevated expression predominantly in nucleoli. Clinicopathological malignancy was strongly correlated with elevated nuclear and nucleolar CK2α expression. Recurrence-free survival was significantly worse (P = .0002) in patients with positive nucleolar CK2α staining. The 5-y survival rate decreased to a roughly 50% in nucleolar CK2α-positive patients of triple-negative (P = .0069) and p Stage 3 (P = .0073) groups. In contrast, no patients relapsed or died in the triple-negative group who exhibited a lack of nucleolar CK2α staining. Evaluation of nucleolar CK2α staining showed a high secondary index with a hazard ratio of 6.629 (P = .001), following lymph node metastasis with a hazard ratio of 14.30 (P = .0008). Multivariate analysis demonstrated that nucleolar CK2α is an independent factor for recurrence-free survival. Therefore, we propose that histochemical evaluation of nucleolar CK2α-positive staining may be a new and robust prognostic indicator for patients who need further treatment. Functional consequences of nucleolar CK2 dysfunction may be a starting point to facilitate development of novel treatments for invasive breast carcinoma.

Maximizing the potential of scientists in Japan: promoting equal participation for women scientists through leadership development.

In order to examine the current status of gender equality in academic societies in Japan, we inquired about the number of women involved in leadership activities at society conferences and annual meetings, as these activities are critical in shaping scientific careers. Our findings show a clear bias against female scientists, and a need to raise consciousness and awareness in order to move closer to equality for future generations.

Cell cycle and activation of CK2.

Casein kinase 2 (CK2) is a highly conserved and ubiquitous eukaryotic Ser/Thr protein kinase. Genetic, biochemical, and cell biological studies have indicated the involvement of this enzyme in the control of cell proliferation and in signal transduction. The regulation of CK2 is not well defined, and it has been considered a constitutively non-regulated protein kinase. However, we show that CK2 activation occurred during the progression of cell cycle in response to FBS stimuli of G0 arrested cells. Importantly, we show that as the downstream target for CK2, the phosphorylation of eukaryotic translation-initiation factor eIF5 by CK2 may play a critical role in cell cycle progression. We find that eIF5 is associated with CK2 when the kinase activity is at the highest level in vivo, and is phosphorylated at Ser389 and Ser390 by CK2. Expression of eIF5 mutants that lack those phosphorylation sites reveals that these mutants have a dominant-negative effect on phosphorylation of endogenous eIF5, as well as a significant reduction in the formation of the mature complex, in the growth rate, and the expression of cell cycle-regulated proteins. Also, a pool of CK2 translocates into the nuclear fraction following its activation during the progression of the cell cycle. Consistent with these findings, we report that CK2 may be involved in the regulation of cell cycle progression through the phosphorylation of a key molecule for translation initiation and of nuclear substrates upon activation of CK2 by itself.

Regulatory role of CK2 during the progression of cell cycle.

The protein kinase casein kinase 2 (CK2) is a ubiquitous eukaryotic serine/threonine protein kinase that plays an important role in cell cycle progression. We find that (1) CK2 interacts with a tumor suppressor protein, adenomatous polyposis coli (APC) that occurs at the highest level in G2/M, and (2) the C-terminal region of APC, between amino acid residues 2086-2394, has the strongest activity to suppress CK2. Over-expression of this fragment in HEK293 cells or colorectal carcinoma cells that have truncated mutant APC proteins down-regulates cell proliferation rates as well as colony formation on soft agar. These results indicate that the complex formation between CK2 and full-length APC regulates CK2 activity that, in turn, regulates cell cycle progression, whereas truncated APC in colorectal carcinomas are unable to regulate the cell cycle. In the process to look for the downstream target for CK2, we found that eukaryotic translation initiation factor 5 (eIF5) is phosphorylated by CK2 in vivo as well as in vitro. These results suggest an important role of CK2 on promotion of cell growth through eIF5.

CK2 phosphorylation of eukaryotic translation initiation factor 5 potentiates cell cycle progression.

Casein kinase 2 (CK2) is a ubiquitous eukaryotic Ser/Thr protein kinase that plays an important role in cell cycle progression. Although its function in this process remains unclear, it is known to be required for the G(1) and G(2)/M phase transitions in yeast. Here, we show that CK2 activity changes notably during cell cycle progression and is increased within 3 h of serum stimulation of quiescent cells. During the time period in which it exhibits high enzymatic activity, CK2 associates with and phosphorylates a key molecule for translation initiation, eukaryotic translation initiation factor (eIF) 5. Using MS, we show that Ser-389 and -390 of eIF5 are major sites of phosphorylation by CK2. This is confirmed using eIF5 mutants that lack CK2 sites; the phosphorylation levels of mutant eIF5 proteins are significantly reduced, relative to WT eIF5, both in vitro and in vivo. Expression of these mutants reveals that they have a dominant-negative effect on phosphorylation of endogenous eIF5, and that they perturb synchronous progression of cells through S to M phase, resulting in a significant reduction in growth rate. Furthermore, the formation of mature eIF5/eIF2/eIF3 complex is reduced in these cells, and, in fact, restricted diffusional motion of WT eIF5 was almost abolished in a GFP-tagged eIF5 mutant lacking CK2 phosphorylation sites, as measured by fluorescence correlation spectroscopy. These results suggest that CK2 may be involved in the regulation of cell cycle progression by associating with and phosphorylating a key molecule for translation initiation.

Association and regulation of casein kinase 2 activity by adenomatous polyposis coli protein.

Mutations in the adenomatous polyposis coli (APC) gene are responsible for familial adenomatous polyposis coli and also sporadic colorectal cancer development. By using antibodies raised against the N-terminal region of APC protein, we have detected the variable masses of endogenous APC proteins in individual cell lines established from human colorectal carcinomas caused by nonsense mutations of the gene. Phosphorylation of immunoprecipitates of full-length and truncated APC were observed in in vitro kinase reaction, indicating association of APC with protein kinase activity. The kinase activity complexed with APC was sensitive to heparin and used GTP as phosphoryl donor, suggesting an involvement of casein kinase 2 (CK2). Both CK2alpha- and beta-subunits were found to associate with APC in immunoprecipitates as well as in pull-down assays, with preferential interaction of APC with tetrameric CK2 holoenzyme. In synchronized cell populations, the association of APC with CK2 was cell cycle dependent, with the highest association in G(2)/M. Unexpectedly, APC immunoprecipitates containing full-length APC protein inhibited CK2 in vitro, whereas immunoprecipitates of truncated APC had little effect. This was confirmed by using recombinant APC, and the inhibitory region was localized to the C terminus of APC between residues 2086 and 2394. Overexpression of this fragment in SW480 cells suppressed cell proliferation rates as well as tumorigenesis. These results demonstrate a previously uncharacterized functional interaction between the tumor suppressor protein APC and CK2 and suggest that growth-inhibitory effects of APC may be regulated by inhibition of CK2.