For influenza A and B viruses to be infectious, they require eight viral RNA (vRNA) genome segments to be packaged into virions. For efficient packaging, influenza A viruses utilize cis-acting vRNA sequences, containing both non-coding and protein coding regions of each segment. Whether influenza B viruses have similar packaging signals is unknown. Here we show that coding regions at the 3' and 5' ends of the influenza B virus vRNA segment 4 are required for genome packaging, with the first 30 nt at each end essential for this process. Synonymous mutation of these regions led to virus attenuation, an increase in defective particle production and a reduction in packaging of multiple vRNAs. Overall, our data suggest that the influenza B virus vRNA gene segments likely interact with each other during the packaging process, which is driven by cis-acting packaging signals that extend into protein coding regions of the vRNA.