Progressive multifocal leukoencephalopathy (PML) is a demyelinating disorder of the human brain caused by infection with the human polyomavirus, JC. Up to 80% of humans express serum antibodies to JC virus (JCV), yet considerably fewer people develop PML-predominantly those under immunosuppressive conditions. Recent research showed JCV infection in multiple tissues throughout the body, suggesting sites for viral latency. These observations allow the proposal of pathways that JCV may use from sites of initial infection to the brain. Results from investigations into cell-surface receptors, intracellular DNA-binding proteins, and variant viral regulatory regions also suggest mechanisms that may regulate cellular susceptibility to JCV infection. Together, these data elucidate how JCV may establish infection in various cell types, persist latently or become reactivated, and ultimately reach the brain to cause PML.