Excimer emission usually has a significant Stokes shift, making it an ideal candidate for constructing artificial light-harvesting systems (LHSs). However, the emission of excimers is altered by intermolecular interactions. Precise modulation of the spatial distance and interaction between luminescent units is the key to achieving enhanced emission of excimers. In the current study, electrostatic assembly of polyanionic PSS with cationic bolaamphiphile (PRB) containing pyrene was used to achieve close stacking of pyrene, resulting in significantly enhanced excimer emission. Furthermore, the morphology of the assemblies changed from nanodisks to nanospheres. In particular, the quantum yield of the PRB with PSS increased from 0.5% to 23.1%. In contrast, the amphiphilic molecule containing pyrene with a single hydrophilic head also showed enhanced fluorescence after being co-assembled with PSS. Nevertheless, the fluorescence quantum yield of excimers was much lower than that of the co-assemblies formed with PRB. The assembly formed by PSS and PRB acted as an energy donor. It underwent Förster resonance energy transfer with Nile red, achieving high energy conversion efficiency (Φ_ET = 49.6%) and antenna effect (9.8) at a donor/acceptor molar ratio of 12.5:1.