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

Abstract

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.

Protein Kinase CK2 Subunits Differentially Perturb the Adhesion and Migration of GN11 Cells: A Model of Immature Migrating Neurons

“…Figure 2E shows that there are phosphosites sensitive to the lack of one but not the other catalytic subunit (see bands marked with an asterisk in Figure 2E), denoting that these two catalytic isoforms may regulate different signal transduction pathways. Notably, the highest decrease in the phosphorylation rate was observed in KOβ cells, consistent with the concept that phosphorylation of the majority of CK2 substrates is optimally catalyzed by CK2 holoenzyme [8,31].…”

“…For example, CK2β has been proposed as a binding partner of other kinases, including A-Raf, c-Mos, and Chk1 [50]. This is in agreement with the phosphoproteomic analysis we performed in the CK2β KO C2C12 cell line, where we showed that a large number of phosphosites not conforming to the CK2 consensus are also affected [8]. Moreover, recently, it has been reported that CK2β controls the myogenic commitment of C2C12 cells by regulating MyoD expression independently from catalytic subunits [51].…”

“…Despite such a similarity, however, both catalytic subunits are active in vitro independent of their association to the β subunits [6]. Nevertheless, the phosphorylation of many typical CK2 targets, such as S129-Akt, S13-Cdc37, and S529-NF-kBp65, is substantially increased by CK2β [7,8]. This suggests that regulatory subunits control the substrate-specific targeting of catalytic subunits.…”

Effects of CK2β subunit down-regulation on Akt signalling in HK-2 renal cells

“…CRISPR-Cas9 CK2β knock out (KO) in C2C12 and GN11 cells was performed as described in [33]. Briefly, C2C12 and GN11 cells were transfected with all-in-one pD1301-AD plasmid expressing Cas9-DasherGFP and the sgRNA guide to target CK2β subunit and Lipofectamine 3000 (ThermoFisher Scientific, Waltham, MA, USA) according to manufacturing instructions.…”

Comparing Two Neurodevelopmental Disorders Linked to CK2: Okur-Chung Neurodevelopmental Syndrome and Poirier-Bienvenu Neurodevelopmental Syndrome—Two Sides of the Same Coin?

“…This may have an impact on several neural processes, such as synaptic plasticity via glutamate receptor homeostasis ( Sanz-Clemente et al, 2013 ). 5) Recently, muscle cell clones (C2C12) that are devoid of either α, α’ or β subunits were generated, and phosphoproteomic analysis revealed that lack of the β subunit affected grossly the same phosphosites than knockout of CK2α/α′, however many phosphosites that do not conform to the CK2 consensus were also altered, strengthening the argumentation for roles of CK2β that are independent of CK2 activity ( Borgo et al, 2019 ). CK2β dimers, in the absence of CK2α, were found in mouse testis and brain ( Guerra and Issinger, 1999 ) and appear to have regulatory function on several other protein kinases, such as A-Raf ( Hagemann et al, 1997 ), c-Mos ( Chen et al, 1997 ) and Chk1 ( Guerra et al, 2003 ).…”