Soon after the 16-cell cyst is formed (in germarium region 1), one of the cells with four ring canals is selected to become the oocyte (in germarium region 2). How this choice is made is still unclear. Whatever mechanisms are involved, oocyte specification requires polarized transport and localization of cytoplasmic proteins and mRNAs, restriction of meiosis, and migration of cen-trioles into the oocyte (3-7). A polarized microtubule network, organized by the fusome, facilitates the transport of cytoplasmic factors from the nurse cells to the oocyte (40-42). Centriole migration is guided by the fusome (43), and meiotic entry and progressive restriction of synaptonemal complexes to one cell within the 16-cell cyst are controlled by the BicD and Egl proteins (44,45).
Further progression of oogenesis requires inactivation of the meiotic checkpoint that detects unrepaired double-strand DNA breaks (46), and the maintenance and determination of oocyte fate regulated by PAR-1 (6,47-51). Determination of oocyte fate occurs when the 16-cell cyst, encapsulated by follicle cells, reaches the end of the germarium (region 3) and is accompanied by the anterior-posterior translocation of Bic-D, Orb, centrioles, and the microtubule organizing center (MTOC) within the oocyte. This posterior shift of oocyte markers represents the first sign of polarity within the oocyte itself.
Immunocytological analysis of oocyte specification and determination involves studying the localization of proteins like Bic-D, Orb, PAR-1, PAR-6, Bazooka, y-tubulin, and Inscuteable (a component of the synaptonemal complex). The sources for antibodies against these proteins are listed in refs. 45 and 51 and in Appendix B. The immunostaining methods detailed in Subheadings 3.1.-3.4. can be used to visualize these proteins.
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