The Drosophila paramyosin/miniparamyosin gene expresses two products of different molecular weight transcriptionally regulated from two different promoters. Distinct muscle types also have different relative amounts of myosin, paramyosin, and miniparamyosin, reflecting differences in the organization of their thick filaments. Immunofluorescence and EM data indicate that miniparamyosin is mainly located in the M line and at both ends of the thick filaments in Drosophila indirect flight muscles, while paramyosin is present all along the thick filaments. In the tergal depressor of the trochanter muscle, both proteins are distributed all along the A band. In contrast, in the waterbug, Lethocerus, both paramyosin and miniparamyosin are distributed along the length of the indirect flight and leg muscle thick filaments. Two-dimensional and one-dimensional Western blot analyses have revealed that miniparamyosin has several isoforms, focusing over a very wide pH range, all of which are phosphorylated in vivo. The changes in isoform patterns of miniparamyosin and paramyosin indicate a direct or indirect involvement of these proteins in muscle function and flight. This wide spectrum of potential regulatory characteristics underlines the key importance of paramyosin/miniparamyosin and its complex isoform pattern in the organization of the invertebrate thick filament.