Figure 1: Confocal (upper half) and super-resolution (STED, lower half) micrograph of localising oskar mRNPs in close proximity to the posterior pole of a developing Drosophila oocyte. oskar mRNPs are labeled with a an array of 37 short single stranded DNA oligonucleotides complementary to oskar mRNA, each labeled with a single Atto-Rho14 fluorescent dye (images courtesy of Imre Gaspar).
Figure 2: An mRNA biosensor (TRICK) for oskar mRNA visualises the first round of oskar-TRICK translation in Drosophila oocytes. In early-stage oocytes oskar-TRICK mRNA is labeled by both NLS-PCP-GFP (green) and NLS-MCP-RFP (red), fluorescent RNA-binding proteins, indicating translational repression. In later stages, the NLS-PCP-GFP fluorescent signal is reduced at the posterior pole and Oskar protein (blue) is detected by immunofluorescence, consistent with translation of a portion of the transcripts (see Halstead*, Lionnet*, Wilbertz*, Wippich* et al., Science 2015).
The Ephrussi group dissects the mechanisms underlying intracellular RNA transport and localised translation – fundamental processes mediating the functional polarisation of cells during development and in the nervous system.
Previous and current research
Post-transcriptional RNA regulation is central to organismal development and function. The combination of intracellular mRNA localisation and localised translation is a powerful strategy that allows quick and local deployment of protein activities in cells in response to extrinsic signals. mRNA localisation is widespread and conserved from yeast to man. It is involved in the establishment of cell asymmetry, is particularly evident in large cells, such as eggs and neurons, and has key roles in cell fate decisions, cell migration, cell morphology, and polarised cell functions. Asymmetric RNA localisation can be achieved by different mechanisms, such as active transport of RNAs by motor proteins moving on cytoskeletal elements, local protection of RNAs from degradation, facilitated diffusion and trapping.
In the large Drosophila melanogaster oocyte, asymmetrically localised cell fate determinants specify the body axes and pattern the future embryo and fly, making it an ideal model for the study of RNA localisation. During oogenesis, the embryonic axis determinant-encoding oskar, bicoid and gurken mRNAs are transported to specific sites within the oocyte, where they are anchored and locally translated, thus ensuring spatial restriction of their protein products. A polarised cytoskeleton and specific motor proteins mediate mRNA transport and anchoring within the oocyte. Our research combines live-imaging, super-resolution microscopy, genetics and biochemistry to understand how mRNAs are transported, anchored and locally translated.
Of particular interest is oskar mRNA, which encodes Oskar protein, which is endowed with the unique ability to induce the formation of germ cells, the cells that ensure perpetuation of the species. oskar mRNA is transported to the posterior pole of the oocyte, where it is translated. Oskar protein, which contains LOTUS and SGNH-hydolase-like domains, nucleates formation of germ granules, RNP complexes containing RNAs and proteins essential for germ cell formation and function. Taking a structure-function approach that combines structural biology, genetics, proteomics and transcriptomics, we are addressing how germ granules form and function.
Other classes of RNAs also have important developmental functions. We are investigating the roles of long non-coding RNAs, piRNAs, and of non-canonical RNA binding proteins in Drosophila development.
With its exceptional genetic tools, Drosophila is ideally suited for investigation of the fundamental mechanisms that govern animal development.
Future projects and goals
We combine genetics, biochemistry and a broad spectrum of cell biological and imaging approaches to study:
- How RNA targeting signals and proteins associated to form RNPs competent for mRNA transport and translational control.
- Spatial and temporal control of translation.
- Polarisation of the cytoskeleton.
- The roles and regulation of cytoskeletal motors in RNA localisation.
- Developmental roles of non-canonical RNA binding proteins.
- Germ plasm assembly and function.