Current Project(s) :

Systems biology approach to identify putative networks involved in neurite outgrowth.

Neurons, with their long axons and elaborate dendritic arbor, establish the complex circuitry that is essential for the proper functioning of the nervous system. While a catalogue of structural, molecular and functional differences between axon and dendrite is accumulating, the mechanisms involved in very early events of neuronal differentiation (neurite initiation and elongation) are not well understood, mainly because the precise knowledge of key molecules involved is still missing. Understanding how the neurite initiation site forms and what makes it privileged on the molecular level is a major challenge not only because it is an important event during nervous system development but also because the establishment of subcellular domains with distinct molecular components and properties is a fundamental problem in cell biology. Furthermore, how is the neurite elongation controlled?

The aim of this project is to identify new proteins/networks regulating neurite outgrowth. For this, a microscopy-based systematic screen designed to identify known and novel proteins involved in neurite initiation and elongation was established and applied (figure 1). 21 new proteins that affected the number of neurites per cell or neurite length when ectopically expressed in cells were identified. Among those, 3 were previously linked to various human neurological disorders. Localisation experiments with GFP-tagged proteins in fixed and living cells revealed a further 14 proteins that associated with neurite tips either early or late during neurite outgrowth. Complementing the functional, real time qRT-PCR and time-lapse data with those available in the literature, a putative molecular network that participates in the regulation of neurite outgrowth was proposed (figure 2.). The prediction potential of the network was tested by examining the role of Gsk3ß and PI3K in early events during neurite outgrowth. Furthermore, co-expression of the identified effector proteins with well known regulators of neurite outgrowth provided a first glimpse on a possible functional relationship of these proteins during neurite outgrowth. A web page which contains all the screen results (http://neurite.embl.de) was constructed and it is available to the scientific community.

Currently, I'm focusing on further characterisation of several Golgi localised proteins that after overexpression affected the neurite length, therefore potentially involved in the regulation of the neuronal vesicle trafficking.

Laketa V, Simpson JC, Bechtel S, Wiemann S, Pepperkok R: High-content microscopy identifies new neurite outgrowth regulators. Mol Biol Cell. 2007 Jan;18(1):242-52. Epub 2006 Nov 8.

Patent #US60/849,032: "New molecules regulating neurite outgrowth"

Figure 1. Strategy to identify proteins involved in neurite outgrowth.

Figure 2. Hypothetical network involved in early events in neurite outgrowth in PC12 cells.

Phosphoinositides regulation of tyrosine kinase receptor endocytosis and vesicle trafficking

Phosphoinositides have emerged as crucial signalling components in a variety of cellular processes during the past two decades. In collaboration with Sirus Zarbakhsh (Carsten Schultz laboratory, EMBL, Heidelberg) I'm testing the role of different phosphoinositides in processes such as receptor tyrosine kinase endocytosis, vesicle trafficking and neurite outgrowth. Recently synthesised membrane permeable phosphoinositide analogues (Carsten Schultz laboratory, EMBL, Heidelberg) provide an excellent and vital tool in examining the roles of different phosphoinositide species in biological processes.

Currently, I'm conducting a large scale high content siRNA screen to identify phosphoinositide targets during epidermal growth factor receptor (EGFR) endocytosis.