The kinase MK2 in DNA replication upon genotoxic stress and chemotherapy

Abstract

DNA damage constitutes a constant threat to genomic integrity. Cells evolved programs controlled by a complex signaling network to cope with these lesions in order to avoid tumorigenesis. These cellular processes and the signaling cascades that regulate them form the DNA damage response (DDR). Whereas many aspects of the DDR have been investigated in great detail, comparably little is known about how cells respond to genotoxic stress during DNA replication. Here, we identify a hitherto unknown function for the kinase MK2 in the control of replication upon genotoxic stress in S-phase. Originally described as a mediator of general stress signaling in the p38/MK2 pathway, recent studies reported a role of MK2 in checkpoint signaling. In our lab MK2 was previously found to be required for efficient phosphorylation of the histone variant H2AX (yielding γH2AX), a hallmark of the DDR, upon DNA damage induced by ultraviolet (UV) irradiation. This suggests a more general function of the kinase in the DDR than anticipated. We now report that depletion or inhibition of MK2 protects cells from the consequences of UV-induced DNA damage, and mice with genetic ablation of MK2 and its relative MK3 display strongly reduced apoptosis in the skin after UV irradiation. As UV-induced DNA damage mainly affects cells during replication but also in other phases of the cell cycle, we tested whether DNA damage induced by the S-phase-specific drug gemcitabine elicited an MK2-dependent DDR, as well. We found that inhibition or depletion of MK2 indeed reduces the accumulation of γH2AX and increases cell viability following gemcitabine treatment, and this effect cannot be attributed to cell cycle modulation by MK2. MK2 inhibition also rescues slow replication fork progression and increased origin firing caused by gemcitabine, demonstrating that the kinase affects replication in response to DNA damage in S-phase. We furthermore observed that MK2 is required for the genotoxic effects caused by inhibition or depletion of the essential checkpoint kinase Chk1 and that MK2 inhibition also alleviates deregulated replication caused by inhibition of Chk1. Such antagonistic activity between the two kinases comes as a surprise as both share the same target phosphorylation motif. We speculate that MK2 and Chk1 target different proteins, mediated by specific interaction partners not shared between the two. In search for the mechanism underlying the effect of MK2 on replication, we did not find any influence on regulators of origin firing, arguing that MK2 acts directly at the replication fork. The rescue of gemcitabine-induced slow fork speed by MK2 inhibition suggests that the kinase controls lesion bypass mechanisms. Accordingly, we observed that the rescue of deregulated replication by MK2 inhibition depends on translesion synthesis (TLS). We speculate that MK2 directly targets components of the TLS machinery, thereby repressing TLS. In conclusion, our data for the first time reveal an activity of MK2 in replication upon DNA damage. MK2 is required for slow fork speed and increased origin firing upon replicative stress, and this activity depends on TLS. We propose that MK2 balances the DDR by repressing TLS to limit the mutagenic effects of this lesion bypass mechanism, promoting DNA repair or cell death. These findings also identify the p38/MK2 pathway as a potential drug target as enhanced MK2 activation might sensitize cells to chemotherapy.