RNA I, a negative controller of ColE1-type plasmid replication, is metabolized by several RNases in Escherichia coli. Two small derivatives of RNA I are accumulated at nonpermissive temperatures in an E. coli strain carrying the rnpA49 mutation, a thermosensitive mutation in the rnpA gene encoding the protein component of RNase P. A primer extension analysis was carried out to compare 5` processing of RNA I in the E. coli rnpA49 cells at both permissive and nonpermissive temperatures. Derivatives of RNA I having different 5` ends were observed in the cells grown at permissive and nonpermissive temperatures. Some of the derivatives may be generated by the cleavage of RNase P.