Most life forms harbor multiple, diverse mobile genetic elements (MGE) that widely differ in their rates and mechanisms of mobility. Recent findings on two classes of MGE in prokaryotes revealed a novel mechanism, RNA-guided transposition, where a transposon-encoded guide RNA directs the transposase to a unique site in the host genome. Tn7-like transposons, on multiple occasions, recruited CRISPR systems that lost the capacity to cleave target DNA and instead mediate RNA-guided transposition via CRISPR RNA. Conversely, the abundant transposon-associated, RNA-guided nucleases IscB and TnpB that appear to promote proliferation of IS/IS and IS transposons were the likely evolutionary ancestors of type II and type V CRISPR systems, respectively. Thus, RNA-guided target recognition is a major biological phenomenon that connects MGE with host defense mechanisms. More RNA-guided defensive and MGE-associated functionalities are likely to be discovered.