Dengue virus (DENV) is a member of the Flavivirus genus of positive-sense RNA viruses. DENV RNA replication requires cyclization of the viral genome mediated by two pairs of complementary sequences in the 5' and 3' ends, designated 5' and 3' cyclization sequences (5'-3' CS) and the 5' and 3' upstream of AUG region (5'-3' UAR). Here, we demonstrate that another stretch of six nucleotides in the 5' end is involved in DENV replication and possibly genome cyclization. This new sequence is located downstream of the AUG, designated the 5' downstream AUG region (5' DAR); the motif predicted to be complementary in the 3' end is termed the 3' DAR. In addition to the UAR, CS and DAR motifs, two other RNA elements are located at the 5' end of the viral RNA: the 5' stem-loop A (5' SLA) interacts with the viral RNA-dependent RNA polymerase and promotes RNA synthesis, and a stem-loop in the coding region named cHP is involved in translation start site selection as well as RNA replication. We analyzed the interplay of these 5' RNA elements in relation to RNA replication, and our data indicate that two separate functional units are formed; one consists of the SLA, and the other includes the UAR, DAR, cHP, and CS elements. The SLA must be located at the 5' end of the genome, whereas the position of the second unit is more flexible. We also show that the UAR, DAR, cHP, and CS must act in concert and therefore likely function together to form the tertiary RNA structure of the circularized DENV genome.