Stably bound adaptor proteins modulate directionality of RNP transport

Abstract

AbstractKinesin-1 and cytoplasmic dynein are molecular motors that mediate long range transport of cargoes along the microtubule cytoskeleton. oskar RNA has been documented to switch between the motors during its localization in the Drosophila germline syncytium. oskar RNA undergoes dynein-mediated transport from the transcriptionally active nurse cells into the oocyte, following which the RNA translocates via kinesin to the posterior pole. Adaptor proteins link the RNA to its motors: the Egalitarian-Bicaudal-D complex links dynein to oskar RNA for the initial phase of transport, whereas atypical Tropomyosin 1 (aTm1) links kinesin-1 to oskar RNA for the latter phase. Components of the Exon Junction Complex (EJC) as well as the SOLE, a stem loop formed upon splicing of oskar RNA, have also been found to be necessary for kinesin-mediated transport of oskar RNA. In this study, to dissect the minimal elements required for kinesin-based transport, we tethered aTm1 or kinesin-1 to oskar RNA constructs lacking the SOLE. Our results suggest that stably bound aTm1 can indeed bypass the SOLE and EJC to mediate kinesin-1 activity, but the effects of tethered aTm1 are less potent than that of tethered kinesin-1. We also tethered Bicaudal-D to oskar RNA, to test whether this would affect kinesin-directed transport of oskar RNA, and found that tethered Bicaudal-D directs dynein mediated localization. Our results show that activated Bicaudal-D, along with the recruited dynein, is sufficient for dynein activity. We also show that stable binding of kinesin-1 to the RNA cargo is sufficient for strong kinesin-1 activity. Stably bound aTm1, meanwhile, can only mediate mild kinesin activity, suggesting that other factors may be required to stabilize the binding of kinesin-1 to the RNA cargo.