Investigating the in vivo regulation of Dynein activity

The establishment and maintenance of polarity is a fundamental feature of eukaryotic cells. Molecular motors within cells transport a variety of cargo including RNAs, proteins, vesicles, and organelles. The asymmetric distribution of these cargos by motor proteins is the underlying basis of cell polarity. The focus of this application is on the microtubule motor, cytoplasmic Dynein. Dynein is responsible for the transport of most cellular cargo towards the microtubule minus-end. In the absence of cargo, Dynein has limited activity. Fully activating this motor requires the multi-subunit Dynactin complex, and one of several activating cargo adaptors. As suggested by their name, cargo adaptors link Dynein with cargo and are also required for stimulating the activity of the motor. The majority of studies regarding the mechanism of Dynein activation have been performed using in vitro single molecule experiments. By contrast, much less is known regarding how this motor is activated in vivo and which cargo adaptors are critical for its activity. Addressing this gap is knowledge is the focus of this application. The model that I propose to use for these studies is the well-characterized Drosophila melanogaster egg chamber, the reproductive tissue of the female fly. Dynein is essential for the proper maturation of the Drosophila egg chamber and oocyte, and numerous cargoes are known to be transported by Dynein in this tissue. Thus, the egg chamber is an ideal model for examining the mechanism of cargo adaptor mediated Dynein activation. Based on my preliminary results, I will test two hypotheses. In Aim 1, I will test the hypothesis that Bicaudal-D (BicD) is the primary activation cargo adaptor for Dynein in the female germline. In Aim 2, I will test the hypothesis that in the absence of the cargo adaptors Hook or Ninein, that BicD is able to functionally compensate for their loss.