Casein Kinase 1 (CK1) family members are serine/threonine protein kinases that are involved in many biological processes and highly conserved in eukaryotes from protozoan to humans. Even though pathogens exploit host CK1 signaling pathways to survive, the role of CK1 in infectious diseases and host/pathogen interaction is less well characterized compared to other diseases, such as cancer or neurodegenerative diseases. Here we present the current knowledge on CK1 in protozoan parasites highlighting their essential role for parasite survival and their importance for host-pathogen interactions. We also discuss how the dual requirement of CK1 family members for parasite biological processes and host subversion could be exploited to identify novel antimicrobial interventions.

White adipose tissue has now been recognized as an important endocrine organ secreting bioactive molecules termed adipocytokines. In obesity, anti-inflammatory adipocytokines like adiponectin are decreased while pro-inflammatory factors are over-produced. These changes contribute to the development of insulin resistance and obesity-associated diseases. Since members of the casein kinase 1 (CK1) family are involved in the regulation of various signaling pathways we ask here whether they are able to modulate the functions of adiponectin. We show that CK1δ and ε are expressed in adipose tissue and that the expression of CK1 isoforms correlates with that of adiponectin. Furthermore, adiponectin co-immunoprecipitates with CK1δ and CK1ε and is phosphorylated by CK1δ at serine 174 and threonine 235, thereby influencing the formation of adiponectin oligomeric complexes. Furthermore, inhibition of CK1δ in human adipocytes by IC261 leads to an increase in basal and insulin-stimulated glucose uptake. In summary, our data indicate that site-specific phosphorylation of adiponectin, especially at sites targeted by CK1δ in vitro, provides an additional regulatory mechanism for modulating adiponectin complex formation and function.

The prognosis of lymphoid neoplasms has improved considerably during the last decades. However, treatment response for some lymphoid neoplasms is still poor, indicating the need for new therapeutic approaches. One promising new strategy is the inhibition of kinases regulating key signal transduction pathways, which are of central importance in tumorigenesis. Kinases of the CK1 family may represent an attractive drug target since CK1 expression and/or activity are associated with the pathogenesis of malignant diseases. Over the last years efforts were taken to develop highly potent and selective CK1-specific inhibitor compounds and their therapeutic potential has now to be proved in pre-clinical trials. Therefore, we analyzed expression and mutational status of CK1δ in several cell lines representing established lymphoma entities, and also measured the mRNA expression level in primary lymphoma tissue as well as in non-neoplastic blood cells. For a selection of lymphoma cell lines we furthermore determined CK1δ kinase activity and demonstrated therapeutic potential of CK1-specific inhibitors as a putative therapeutic option in the treatment of lymphoid neoplasms.

Oncogenesis in breast cancer often requires the overexpression of the nuclear receptor coactivator AIB1/SRC-3 acting in conjunction with estrogen receptor-alpha (ERalpha). Phosphorylation of both ERalpha and AIB1 has been shown to have profound effects on their functions. In addition, proteasome-mediated degradation plays a major role by regulating their stability and activity. CK1delta, a member of the ubiquitous casein kinase-1 family, is implicated in the progression of breast cancer. In this study, we show that both ERalpha and AIB1 are substrates for CK1delta in vitro, and identify a novel AIB1 phosphorylation site (S601) targeted by CK1delta, significant for the co-activator function of AIB1. CK1delta is able to interact with ERalpha and AIB1 in vivo, while overexpression of CK1delta in breast cancer cells results in an increased association of ERalpha with AIB1 as confirmed by co-immunoprecipitation assays from cell lysates. Using an siRNA-based approach, luciferase reporter assays and qRT-PCR, we observe that silencing of CK1delta leads to reduced ERalpha transcriptional activity, despite increased ERalpha levels, similarly to proteasome inhibition. We provide evidence that AIB1 protein levels are reduced by CK1delta silencing, in an estradiol-dependent manner; such destabilization can be inhibited by pre-treatment with the proteasome inhibitor MG132. We propose that differing activities adopted by ERalpha and AIB1 as a consequence of their interactions with and phosphorylation by CK1delta, particularly AIB1 stabilization, influence the transcriptional activity of ERalpha, and therefore have a role in breast cancer development.

CK1δ, a member of the casein kinase 1 family, is involved in the regulation of various cellular processes and has been associated with the pathophysiology of neurodegenerative diseases and cancer. Therefore recently, interest in generating highly specific inhibitors for personalized therapy has increased enormously. However, the efficacy of newly developed inhibitors is affected by the phosphorylation state of CK1δ. Cellular kinases phosphorylating CK1δ within its C-terminal domain have been identified but still more information regarding the role of site-specific phosphorylation in modulating the activity of CK1δ is required. Here we show that Chk1 phosphorylates rat CK1δ at serine residues 328, 331, 370, and threonine residue 397 as well as the human CK1δ transcription variants 1 and 2. CK1δ mutant proteins bearing one, two or three mutations at these identified phosphorylation sites exhibited significant differences in their kinetic properties compared to wild-type CK1δ. Additionally, CK1δ co-precipitates with Chk1 from HT1080 cell extracts and activation of cellular Chk1 resulted in a significant decrease in cellular CK1δ kinase activity. Taken together, these data point towards a possible regulatory relationship between Chk1 and CK1δ.

CK2 is a pleiotropic S/T protein kinase (formerly known as casein kinase 2) which is attracting increasing interest as therapeutic target, and the identification of its substrates is a crucial step in determining its involvement in different pathological conditions. We recently found that S131 of Akt2 (homologous to the well established CK2 target S129 of Akt1) is not phosphorylated by CK2 either in vitro or in vivo, although the consensus sequence recognized by CK2 (S/T-x-x-E/D/pS/pT) is conserved in it. Here, by exploiting synthetic peptides, in cell transfection experiments, and computational analysis, we show that a single sequence element, a T at position n+1, hampers phosphorylation, causing an α-helix structure organization which prevents the recognition of its own consensus by CK2. Our results highlight the role of negative determinants as crucial modulators of CK2 targeting and corroborate the concept that Akt1 and Akt2 display isoform specific features. Experiments with synthetic peptides suggest that Akt2 S131 could be phosphorylated by kinases of the Plk (Polo-like kinase) family, which are insensitive to the presence of the n+1 T. The low phylogenetic conservation of the Akt2 sequence around S131, as opposed to the extremely well-conserved Akt1 homologous sequence, would indicate a dominant positive role in the selective pressure only for the Akt1 phosphoacceptor site committed to undergo phosphorylation by CK2. By contrast, Akt2 S131 may mediate the response to specific physio/pathological conditions, being consequently shielded against basal CK2 targeting.

CK2 (an acronym derived from the misnomer "casein kinase 2") denotes a ubiquitous, highly pleiotropic protein kinase which has been implicated in global human pathologies, with special reference to cancer. A large spectrum of fairly selective, cell permeable CK2 inhibitors are available, one of which, CX4945 is already in clinical trials for the treatment of neoplasia. Another recently developed CK2 inhibitor, GO289, displays in vitro potency and selectivity comparable to CX4945. Here the cellular efficiency of these two inhibitors has been evaluated by treating C2C12 myoblasts for 5 h with each of them at 4 μM concentration and running a quantitative phosphoproteomics analysis of phosphosites affected by the two compounds. A small but significant proportion of the quantified phosphosites is decreased by treatment with CX4945 and, even more with GO289. This figure substantially increases if a subset of quantified phosphosites conforming to the CK2 consensus (pS/pT-x-x-D/E/pS/pT) is considered. Also in this case GO289 is more effective than CX4945. By adopting stringent criteria two shortlists of 70 and 35 sites whose phosphorylation is decreased >50% by GO289 and CX4945, respectively, have been generated. All these phosphosites conform to the consensus of CK2 with just sporadic exceptions. Their WebLogos are indistinguishable from that of bona fide CK2 phosphosites and their Two-Sample Logos rule out any significant contribution of Pro-directed and basophilic protein kinases to their generation. To sum up, we can conclude that by treating C2C12 cells for 5 h with either CX4945 or GO289 off-target effects are negligible since almost all the phosphosites undergoing a substantial reduction are attributable to CK2, with a higher inhibitory efficacy displayed by GO289. CX4945 and GO289 provide highly selective tools to control the CK2-dependent phosphoproteome compared with previously developed CK2 inhibitors.