Characterisation of the c10orf76-PI4KB complex, and its necessity for Golgi PI4P levels and enterovirus replication

The lipid kinase PI4KB, which generates phosphatidylinositol 4-phosphate(PI4P), is a key enzyme in regulating membrane transportand is also hijacked by multiple picornaviruses to mediate viral replication. PI4KBcan interact with multiple protein binding partners, which are differentially manipulated by picornaviruses to facilitate replication. The protein c10orf76 is a PI4KB-associated protein that increases PI4P levels at the Golgi, and is essential for the viral replicationof specific enteroviruses. We used hydrogen deuterium exchangemass spectrometry to characterize the c10orf76- PI4KBcomplex and reveal that binding is mediated by the kinase linker of PI4KB, with formation of the heterodimeric complex modulated by PKA-dependent phosphorylation. Complex-disrupting mutations demonstrate that PI4KBis required for membrane recruitment of c10orf76 to the Golgi, and that an intact c10orf76- PI4KBcomplex is required for the replication of c10orf76-dependent enteroviruses. Intriguingly, c10orf76 was also required for proper Arf1 activation at the Golgi, providing a putative mechanism for the c10orf76-dependent increase in PI4P levels at the Golgi.