Abstract Viruses utilize a plethora of strategies to manipulate the host pathways and hijack its machineries for efficient replication. Several DNA as well as handful of RNA viruses are reported to interact with proteins involved in DNA damage responses (DDR). As the DDR pathways have never been explored in Alphaviruses, this investigation intended to determine the importance of the DDR pathways in CHIKV infection through in vitro, in vivo and ex vivo models. The study reveals that CHIKV infection activates the Chk2 and Chk1 proteins associated with DDR signaling pathways and increases DNA damage by 95%. Inhibition of both ATM-ATR kinases by ATM/ATR kinase inhibitor (AAKi) shows drastic reduction in viral particle formation in vitro. Next, the treatment of mice with this drug has been shown to reduce the disease score substantially in CHIKV-infected C57BL/6 mice with 71% decrement in the viral copy and the same has been established in hPBMC-derived monocyte-macrophage populations. Additionally, gene silencing of Chk2 and Chk1 reduces viral progeny formation around 73.7% and 78% respectively. Moreover, it has been demonstrated that CHIKV-nsP2 interacts with Chk2 and Chk1 during CHIKV infection and docking analysis depicts the specific amino acids responsible for these interactions. Further, the data suggests that CHIKV infection induces cell cycle arrest in G1 and G2 phases. In conclusion, this work demonstrated for the first time the mechanistic insight of the induction of DDR pathways by CHIKV that might contribute to the designing of effective therapeutics for the control of this virus infection in future. IMPORTANCE Viruses being intra-cellular parasite, need several host cell machineries so as to achieve effective replication of their own genome, along with virus-encoded enzymes. One of the strategies is to hijack the DDR pathways. Several DNA as well as handful of RNA viruses interact with the cellular proteins involved in DDR pathways, however, reports with respect to the association of Chk2 and Chk1 in alphavirus infection are scanty. Hence, this study is amongst the first to report that modulation of DDR pathways is crucial for effective CHIKV infection. This work also shows that there is interaction of CHIKV-nsP2 with two crucial host factors, Chk2 and Chk1 for efficient viral infection. Interestingly, CHIKV infection was found to cause DNA damage and arrest cell cycle in G1 and G2 phases to facilitate viral infection. This information might facilitate to develop effective therapeutics for the control of the CHIKV infection in future.

DNA damage response signaling is crucial for effective Chikungunya virus replication