Elaboration of the messenger transport organizer pathway for localization of RNA to the vegetal cortex of Xenopus oocytes

Previous studies demonstrated that there were two pathways, the messenger transport organizer (METRO) or early and the Vg1 or late, which function during stages 1 to 3 of oogenesis for the localization of RNAs at the vegetal cortex of Xenopus oocytes. In the present study we analyzed the properties of the METRO pathway, which localizes Xlsirt, Xcat2, and Xwnt11 RNAs to a specific region of the vegetal cortex during stage 1 of oogenesis. A combination of methodologies involving both fixed material and living oocytes was used to analyze RNA localization. We show that in early diplotene pre-stage 1 oocytes (25-50 μm in diameter) both endogenous and injected exogenous METRO RNAs translocated to multiple mitochondrial aggregates (pre-mitochondrial clouds) that surround the germinal vesicle (GV). However, by early stage 1 (diplotene oocytes, 50-200 μm), all three of the RNAs discriminated between the different clouds and translocated exclusively within the METRO of a single mitochondrial cloud. Therefore, in stage 1 diplotene oocytes there is a unique mechanism causing a change in the intrinsic property of the mitochondrial clouds which designates one of them as the RNA transport vehicle. During translocation through the cytoplasm Xlsirt and Xcat2 RNAs were detected associated with cytoplasmic particles of different morphologies. Additionally, we also found that the translocation of RNAs through the early or METRO pathway, unlike that of the late pathway, occurred in the absence of intact microtubule and actin microfilament cytoskeletal elements. This supports a cytoskeletal-independent model for localization of RNAs through the METRO pathway.