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MAPK6/ERK3 is an atypical member of the MAPKs. An essential role has been suggested by the perinatal lethal phenotype of ERK3 knockout mice carrying a lacZ insertion in exon 2 due to pulmonary disfunction and by defects in function, activation and positive selection of T cells. To study the role of ERK3 in vivo, we generated mice carrying a conditional Erk3 allele with exon3 flanked by LoxP sites. Loss of ERK3 protein was validated after deletion of Erk3 in the female germ line using zona pellucida 3 (Zp3)-cre and a clear reduction of the protein kinase MK5 is detected, providing first evidence for the existence of the ERK3/MK5 signaling complex in vivo. In contrast to the previously reported Erk3 knockout phenotype, these mice are viable and fertile, do not display pulmonary hypoplasia, acute respiratory failure, abnormal T cell development, reduction of thymocyte numbers or altered T cells selection. Hence, ERK3 is dispensable for pulmonary and T-cell functions. The perinatal lethality, lung and T-cell defects of the previous ERK3 knockout mice are likely due to ERK3-unrelated effects of the inserted lacZ-neomycin-resistance-cassette. The knockout mouse of the closely related atypical MAPK ERK4/MAPK4 is also normal suggesting redundant functions of both protein kinases.
5-iodotubercidin sensitizes cells to RIPK1-dependent necroptosis by interfering with NFκB signaling
(2023)
Receptor-interacting protein kinases (RIPK) −1 and −3 are master regulators of cell fate decisions in response to diverse stimuli and are subjected to multiple checkpoint controls. Earlier studies have established the presence of distinct IKK1/2 and p38/MK2-dependent checkpoints which suppress RIPK1 activation by directly phosphorylating it at different residues. In the present study, we investigated TNF-induced death in MAPK-activated protein kinase 2 (MK2)-deficient cells and show that MK2-deficiency or inactivation predominantly results in necroptotic cell death, even in the absence of caspase inhibition. While MK2-deficient cells can be rescued from necroptosis by RIPK1 inhibitors, RIPK3 inhibition seems to revert the process triggering apoptosis. To understand the mechanism of this necroptosis switch, we screened a 149-compound kinase inhibitor library for compounds which preferentially sensitize MK2-deficient MEFs to TNF-induced cell death. The most potent inhibitor identified was 5-Iodotubericidin, an adenosine analogue acting as adenosine kinase and protein kinase inhibitor. 5-ITu also potentiated LPS-induced necroptosis when combined with MK2 inhibition in RAW264.7 macrophages. Further mechanistic studies revealed that 5-Iodotubericidin induces RIPK1-dependent necroptosis in the absence of MK2 activity by suppressing IKK signaling. The identification of this role for the multitarget kinase inhibitor 5-ITu in TNF-, LPS- and chemotherapeutics-induced necroptosis will have potential implications in RIPK1-targeted therapies.