Membrane trafficking is a highly regulated process, crucial for cellular homeostasis. Defects in the regulation of intracellular transport often result in diseases such as cancer or neurodegenerative disorders. Neuronal functions indeed are particularly dependent on a healthy membrane trafficking, as they require neuron-specific membrane transport between distant axonal and dendritic extensions for neurite outgrowth, neuronal maintenance, for synaptic transmission, and for correct distribution of cell surface receptors. Therefore, neurons have specialized mechanisms that regulate the transport of proteins, organelles, and receptors over long distances, in addition to exocytosis and endocytosis of synaptic vesicles at synapses.
One important class of membrane trafficking regulators are the Rab GTPases. In this project, the master student will investigate the role of a Rab mutant responsible of the Warburg Micro syndrome, a rare inherited neurodevelopmental disorder. Although the genetic causes of Warburg Micro syndrome are known, it is still unclear how the identified mutations in a specific Rab protein, Rab18, or in its regulators affect the cell physiology.
Recent work from our group showed that Rab18 plays a role together with the cytoskeleton in transporting endoplasmic reticulum (ER) tubules to the plasma membrane to mediate the formation of cell protrusions (Guadagno et al., 2020). In addition, Rab18 is involved in a pathway inhibiting neurite growth in vitro (Wu et al., 2016). Therefore, in this project, it will be investigated if Warburg Micro mutants affect the ER by preventing its interaction with the cytoskeleton. It will be further explored whether the defects in neurite outgrowth are the consequence of this altered interaction.
For this, co-immunoprecipitation and pull-down experiments will be used to demonstrate whether the interaction of the mutants with the cytoskeleton is altered, and state-of-the art microscopy to study how these mutants affect the ER morphology and dynamics, neurite outgrowth, and the underlying molecular mechanisms in neurons.
Methods:
This project will use techniques in biochemistry, cellular and molecular biology, including:
- recombinant DNA technology
- protein expression and purification
- immunoprecipitation
- Western blotting
- cell culture
- cell transfection
- RNA interference
- immunofluorescence and state-of-the-art microscopy techniques, using microscopes available at the NorMic imaging platform (http://www.mn.uio.no/ibv/english/research/about/infrastructure/imaging/)
- quantitative image analysis using software such as ImageJ, Imaris, etc.
Work place and environment:
The project will be performed at the Department of Biosciences in the group of Cinzia Progida.
The group consists of researchers at different stages of their careers (Master, PhD students, post-docs and an experienced technician).
If any questions, contact: c.a.m.progida@ibv.uio.no
Phone: 22854441
Room: 3626
References:
Guadagno, N.A., Margiotta, A., Bjrnestad, S.A., Haugen, L.H., Kjos, I., Xu, X.C., Hu, X., Bakke, O., Margadant, F., and Progida, C. (2020). Rab18 regulates focal adhesion dynamics by interacting with kinectin-1 at the endoplasmic reticulum. J Cell Biol 219.
Wu, Q., Sun, X., Yue, W., Lu, T., Ruan, Y., Chen, T., and Zhang, D. (2016). RAB18, a protein associated with Warburg Micro syndrome, controls neuronal migration in the developing cerebral cortex. Mol Brain 9, 19.