Shigella flexneri type III effector OspC3 was recently found to evade host pyroptosis by targeting caspase-11 for ADP-riboxanation. The ADP-riboxanase activity was shared by its paralogues, OspC1 and OspC2, whereas the host substrates of them are still unknown. To solve this problem, we employed eAf1521 enrichment coupled with mass spectrometry (MS)-based proteomics to profile the host substrates of OspC1. In this study, we identified HERC5 as a host target of OspC1. As described previously, HERC5 functions as an E3 ISG15 ligase and catalyzes the ISGylation of a huge subset of host and pathogen proteins upon bacterial infection. Here, we show that S. flexneri hijacks host ISGylation pathway by OspC1-catalyzed ADP-riboxanation of HERC5 to promote its own survival and proliferation in host cells.