▪ Abstract In a wide variety of organisms, gametes develop within clusters of interconnected germline cells called cysts. Four major principles guide the construction of most cysts: synchronous division, a maximally branched pattern of interconnection between cells, specific changes in cyst geometry, and cyst polarization. The fusome is a germline-specific organelle that is associated with cyst formation in many insects and is likely to play an essential role in these processes. This review examines the cellular and molecular processes that underlie fusome formation and cyst initiation, construction, and polarization in Drosophila melanogaster. The studies described here highlight the importance of cyst formation to the subsequent development of functional gametes.

Early during Drosophila oogenesis the 16 interconnected cells of each germ-line cyst choose between two alternative fates. The single future oocyte enters meiosis, arrests, and becomes transcriptionally quiescent. The remaining 15 cells initiate a series of polyploid cell cycles to prepare for their role as nurse cells. Like many other polyploid and polytene cells, during nurse cell growth the major satellite DNAs become highly under-represented by a mechanism that has remained obscure. We implicate the cell-cycle regulator cyclin E in DNA under-representation by identifying a hypomorphic, female sterile cycE mutation, cycE01672, that increases the amount of satellite DNA propagated in nurse cells. In mutant but not wild-type endomitotic nurse cells, "late S" patterns of bromodeoxyuridine incorporation are observed similar to those in mitotic cells. CycE protein still cycles in cycE01672 germ-line cysts but at reduced levels, and it is found throughout a longer fraction of the cell cycle. Our experiments support the view that oscillating levels of CycE control the polyploid S phase. Moreover, they indicate that a checkpoint linking the presence of unreplicated DNA to the CycE oscillator is lacking, leading to incomplete replication of late-replicating sequences such as satellite DNAs. Unexpectedly, two to three of the 16 cells in cycE01672 cysts frequently differentiate as oocytes, implicating cell-cycle programming in oocyte determination.

Over-replication of two clusters of chorion genes inDrosophila ovarian follicle cells is essential for rapid eggshell biosynthesis. The relationship of this amplification to the follicle cell cycles has remained unclear. To investigate the regulation of amplification, we developed a technique to detect amplifying chorion genes in individual follicle cells using BrdU incorporation and FISH. Amplification occurs in two developmental phases. One of the gene clusters begins to amplify periodically during S phases of follicle cell endocycles. Subsequently, after endocycles have ceased, both clusters amplify continuously during the remainder of oogenesis. In contrast to the early phase, late amplification commences synchronously among follicle cells. The pattern of Cyclin E expression mirrors these two phases. We present evidence that Cyclin E is required positively for amplification. We suggest that Cyclin E also acts negatively to inhibit refiring of most origins within a cycle, and that specific factors at chorion origins allow them to escape this negative rereplication control. Our findings suggest that chorion amplification is a model for understanding metazoan replicons and the controls that restrict replication to once per cell cycle.

During metazoan development, the organization of the cell cycle is often modified in response to developmental signals. The endocycle provides a dramatic example of this phenomenon. In the endocycle, also referred to as the endoreplicative cycle, cells undergo successive rounds of DNA replication without an intervening mitosis. Often the endocycle is used to expand the genome of a group of specialized cells that are highly biosynthetically active. In these circumstances, large polyploid cells are produced in organisms that are primarily comprised of diploid cells. However, many organisms achieve growth by increasing cell size, rather than cell number. This strategy is more generally exploited in insects and plants. For instance, in the insect Drosophila melanogaster, the majority of the larval tissues, as well as many adult tissues, enter the endocycle and become polyploid. Therefore, Drosophila has been a rich source for studies on endocycle regulation. Recent work from Drosophila is beginning to reveal how developmental signals promote the transition from the mitotic cycle to the endocycle, as well as what drives endocycle progression. In addition, studies on the endocycle have provided insight into the regulatory principles underlying the once per cell cycle replication of the genome, as well as the relationship between S phase and mitosis.

The synaptonemal complex (SC) is intimately involved in the process of meiotic recombination in most organisms, but its exact role remains enigmatic. One reason for this uncertainty is that the overall structure of the SC is evolutionarily conserved, but many SC proteins are not. Two putative SC proteins have been identified in Drosophila : C(3)G and C(2)M. Mutations in either gene cause defects in SC structure and meiotic recombination. Although neither gene is well conserved at the amino acid level, the predicted secondary structure of C(3)G is similar to that of transversefilament proteins, and C(2)M is a distantly related member of the α-kleisin family that includes Rec8, a meiosis-specific cohesin protein. Here, we use immunogold labeling of SCs in Drosophila ovaries to localize C(3)G and C(2)M at the EM level. We show that both C(3)G and C(2)M are components of the SC, that the orientation of C(3)G within the SC is similar to other transverse-filament proteins, and that the N terminus of C(2)M is located in the central region adjacent to the lateral elements (LEs). Based on our data and the known phenotypes of C(2)M and C(3)G mutants, we propose a model of SC structure in which C(2)M links C(3)G to the LEs.