Archaeal promoters contain a TATA-box, an adjacent upstream TFB-recognition element (BRE), and a downstream initiator (INR) region from which transcription originates. While the contribution of A-box and BRE to promoter strength is well established, the role of DNA sequences within the INR region and its vicinity on transcription efficiency and start site selection remains unclear. Here, I demonstrate using the strong Sulfolobus shibatae viral T6 promoter that either substitution of its natural sequence from –17 and beyond with plasmid DNA or introduction of point transversion mutations at +3, –2, –4, and –5 positions reduce promoter strength dramatically, whereas +1, –1, and –2 mutations influence the transcription start site. These data therefore reveal that the INR region plays a role as important as the BRE and the A-box in T6 gene transcription. Key words: Archaea, transcription, initiator (INR), Sulfolobus shibatae, core promoter.