Significance Atomic structures of the polymerase–endonuclease complex of the orthomyxovirus influenza and the orthobunyavirus La Crosse—two distinct segmented negative-sense (SNS) RNA viruses—demonstrate that binding of the genomic 5′ RNA rearranges the catalytic residues of the RNA-dependent RNA-polymerase (RdRP). Working with the arenavirus, Machupo, we demonstrate that 5′ RNAs from the genomic and antigenomic copies of both segments activate the RdRP in conjunction with a specific promoter. This study builds upon structural studies with two different SNS RNA viruses to reveal a previously unappreciated mechanism of RNA-guided promoter-specific polymerase regulation in SNS RNA viruses. The conservation of activating RNA elements among the polymerase–endonuclease complexes of SNS RNA viruses suggests new avenues for developing antiviral therapeutics.