ABSTRACT During Drosophila oogenesis, developing germline cysts are spanned by a large cytoplasmic structure called a fusome, containing α-spectrin and the adducin-like product of the hu-li tai shao (hts) gene. We found that fusomes contain two additional membrane skeletal proteins: β-spectrin and ankyrin. hts was shown previously to be required for cyst formation and oocyte differentiation; the role of the fusome itself, however, and the organization and function of its other components, remains unclear. Using the FRT/FLP recombinase system to generate clones of α-spectrindeficient cells in the ovary, we have shown that α-spectrin is also required for cyst formation and oocyte differentiation, but that its role in each process is distinct from that of Hts protein. Furthermore, α-spectrin is required for these processes in germline cells, but not in the follicle cells that surround each cyst. We have also found that the organization of membrane skeletal proteins is more dependent on α-spectrin in the fusome than at the plasma membrane in other cells. Our results suggest that the fusome and its associated membrane skeleton play a central role in regulating the divisions and differentiation of cyst cells.