The role of the sub-cellular localisation of the transcription factor FoxG1 in neuronal development of the forebrain

In order for the nervous system to function properly, different types of neurons need to be produced and the right connections need to be made between them, so that a functional circuit, not dissimilar to an electrical circuit, can be formed. For this to happen, neurons use long extensions of their cell bodies, called axons. These are dynamic processes that can grow quite long and navigate through the tissue, until they find the right cells to form connections with.
The generation of several different types of neurons relies on the activity of a set of key proteins termed Transcription Factors. These proteins are special in the sense that they drive the production of several other proteins by the cell. Transcription factors achieve this by binding to DNA, which is then translated by the cell?s machinery into making a particular protein. There are thousands of transcription factors known, each with a preference for a particular sequence of DNA, and therefore able to influence the production of different sets of proteins. The DNA of a cell is in the nucleus therefore, transcription factors are also found in the nucleus. In fact, the conventional view is that transcription factors only have a function in the nucleus.
We have been investigating the role of a transcription factor, Foxg1, which is important for the generation of neurons in the brain in all animals. Depending on when this gene malfunctions in humans, the brain may fail to form, or cancer or mental retardation can arise. We have recently made the unexpected discovery that FoxG1 is not only found in the nucleus, as expected for conventional transcription factors, but it is also found in the axons of brain neurons. In this proposal, we will ask 3 questions; in which neurons exactly is FoxG1 found in the axon, how does it get there and finally, what does it do?
From the thousands of transcription factors known, only a handful have been shown to have an ?extra-nuclear? function in axons. Their precise function there is not quite clear yet, but it seems that they help the axon grow in the right direction. Our proposed work will have two benefits; it will increase our understanding of the function of a key brain transcription factor and will further our understanding of the unconventional role that transcription factors may have in axonal development.

The development of the telencephalon is orchestrated by transcription factors (TFs), which control gene expression, often in response to signalling events. One way to regulate the activity of TF is by nucleo-cytoplamic shuttling. Since the classical view is that TFs exert their function in the nucleus it is widely believed that nuclear exclusion of a TF is only a means to prevent or terminate its transcriptional activity. However, in the study of neuronal development, a few cases have emerged where TFs can have a distinct role outside the nucleus, particularly in axons, where they may be secreted, locally modified or may modulate local translation. Axons may have a need for this unusual mode of TF regulation because they are long cytoplasmic processes navigating through a dynamic environment.

We have been investigating the function of a winged helix (forkhead) TF, FoxG1, expressed in the telencephalon in all vertebrates examined to date. We and others, collectively have shown that FoxG1 is involved in controlling proliferation, differentiation and survival in the telencepalon, and axonal guidance in the retina. All the work so far has focused, or even assumed, that FoxG1 only has a role in the nucleus. However, we have recently shown that FoxG1 is controlled by nucleo-cytoplasmic shuttling and furthermore, we now have preliminary data that FoxG1 protein is localised in axons of some telencephalic neurons and distinctly localised in the axonal terminal. The role of this localisation is unknown.

Here, we combine our expertise on axonal development and the control of FoxG1 to test the hypothesis that the non-nuclear fraction of FoxG1 plays a role in axonal development. Firstly, we will establish which cortical neurons and at what developmental stage show axonal localisation of FoxG1. Information from this aim will aid in the interpretation of functional experiments in aim 3. Secondly, we will ask whether in addition to the protein, FoxG1 mRNA is also localised to the axon and we will determine the basic mechanisms of localisation. Information from this aim, will aid in choosing the best method for depleting FoxG1 locally in the axon, in aim 3. Finally, in aim 3, we will test the function of FoxG1 in axonal development by undertaking a dual approach: we will address existing possibilities, as have been described for a handful of other TFs, but will also address function in an unbiased assay involving the localised depletion of FoxG1 in axons.