To examine the regions of the male-specific filamentous bacteriophage f1 genome that include signals for mRNA processing, the 5′ endpoints of the major in VIVO phage mRNAs have been located in the f1 DNA sequence by S1 nuclease mapping. The 5′ ends of the purified mRNAs and additional phage-specific RNAs transiently visible early after infection occur in clusters of T-rich residues within genes that code for three phage proteins. When a 270-nucleotide region encompassing the 5′ endpoints of three processed RHAs is transcribed as part of the bacteriophage λ N. mRNA in uninfected female cells, RNA 5′ ends identical to ends of the three f1 RNAs are generated from the λ-f1 precursor. This finding indicates that the mRNA processing activity is encoded by the bacterial host, and that its recognition sites are present in the local regions near the 5′ ends which result from RNA cleavage. Several characteristics of f1 mRNA processing events have implications for the differential regulation of adjacent phage genes constrained in the same transcription unit, and may be representative of similar processing events occurring in the bacterial cell.