Research Interests and Projects:

Genetic Regulation of Drosophila Oogenesis

The formation of eggs is a central function of female biology. While a great deal is known about oocyte maturation in mammals, the earliest steps of oogenesis are less well understood. We are studying oogenesis in Drosophila, which has striking similarities to oogenesis in other animals. Our projects are centered around understanding oocyte growth that is dependent on the actions of nurse cells.

Our primary interest is in determining the mechanism of intercellular cytoplasm transport during Drosophila oogenesis. This process is required to produce mature eggs that contain maternal components sufficient to support early embryogenesis. Cytoplasm transport occurs in two phases: slow transport (several days) during which the oocyte grows at about the same rate as nurse cells and fast transport (~30 min.) during which all remaining cytoplasm is squeezed into the oocyte by nurse cell contraction.

Through our analyses of several mutants in which cytoplasm transport is defective, we have discovered that regulation of the actin cytoskeleton is crucial to carrying out transport properly. One class of genes under study is required to form the intercellular junctions called ring canals that connect the nurse cells to the oocyte. Ring canals are actin-rich structures established by arrested cytokinesis that grow to about 10 microns in diameter. Another class of genes is required late in oogenesis to carry out the actin reorganization and apoptotic death of nurse cells that are required for fast cytoplasm transport. Thus, our research uses a variety of approaches (genetics, cell biology, biochemistry) to study the regulation of nurse cell development.

Drosophila egg chambers develop in assembly lines within ovarioles

Protein Trapping Screen

In a new approach to identifying genes important for oogenesis, we are carrying out a large scale protein trapping screen. An artifical exon encoding Green Fluorescent Protein (GFP) is contained within a transposable element. Mobilization of the element can lead to insertion of the GFP exon in the intron of a gene and incorporation of the exon into the mature transcript. If the GFP coding sequence is in the same reading frame as the host gene, a GFP fusion protein can be translated from the mRNA.

Ring Canal Development and Function

The most intimate form of communication between cells is the direct sharing of cytoplasm. There are many examples where discreet, singly nucleated cells share their cytoplasm with their mitotically-derived sister cells. Organisms ranging from plants to mammals have found ways to arrest cleavage furrows during mitotic divisions of certain cell types in order to convert those cleavage furrows into stable junctions (Robinson and Cooley, 1996). In contrast to tiny gap junctions that allow passage of only small molecules and peptides (<1-2 kDa), some intercellular bridges are large enough to allow movement of organelles between the cells. Although there are a number of examples of stable intercellular bridges in somatic cells, germline cells are most remarkable for using bridges during development. Both female and male germline cells from species ranging from insects to humans spend a part of their lifetime in syncytia of mitotically related cells. In all the cases that have been characterized, the formation of intercellular bridges is closely coupled with altered cytokinesis; indeed, proteins present in cleavage furrows sometimes persist in stable bridges.

Fast cytoplasm transport

The final, rapid transport of nurse cell cytoplasm to the oocyte is driven by a contraction of the nurse cell cluster that is mediated by cortical actin and cytoplasmic myosin II. Preparations for fast cytoplasm transport involve dramatic changes in the cell biology of nurse cells. Highly organized arrays of actin filament bundles are formed in the cytoplasm of nurse cells, extending from the plasma membranes to the nuclear envelopes. These bundles of actin are necessary to keep the large nuclei from blocking ring canals during nurse cell contraction. After the bundles are established, the nuclear envelopes become permeable, releasing nuclear contents into the cytoplasm. One result of nuclear permeability is the release of free calcium into the cytoplasm, which is likely to be involved in triggering nurse cell contraction.