First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping researchers promote themselves alongside their papers. Thomas Blake is first author on ‘ Filopodial protrusion driven by density-dependent Ena–TOCA-1 interactions’, published in JCS. Thomas is a Research Associate in the lab of Jennifer Gallop at the Wellcome/Cancer Research UK Gurdon Institute, Cambridge, UK, investigating the molecular mechanisms that drive cell motility using quantitative microscopy techniques.

Thomas Blake

How would you explain the main findings of your paper in lay terms?

During the development of the nervous system, nerve cells (neurons) must navigate accurately over long distances to make connections in the brain. Neurons move by carefully controlling the shape of their cell skeleton and making dynamic ‘fingers’ called filopodia, which are made of growing and shrinking filaments of a protein called actin. Many other proteins regulate the growth of actin filaments, either by directly elongating them or by organising other components of the regulatory machinery. In this paper, we show that TOCA-1, a candidate organising protein, is found at the tips of extending filopodia in frog neurons and demonstrate, using two different statistical approaches, that TOCA-1 is directly driving this extension. We validate these findings by modifying a key region of the TOCA-1 protein, which prevents TOCA-1 from promoting filopodia extension, and by using a drug that blocks interacting proteins and prevents filopodia extension. Finally, we reveal that TOCA-1 works together with Ena, an actin filament elongator protein. The two proteins are specifically found together at early stages of the formation of filopodia – TOCA-1 arrives early at filopodia to organise other proteins including Ena, which in turn drives extension of filopodia, thereby enabling neuronal growth.

Were there any specific challenges associated with this project? If so, how did you overcome them?

I love working with frog neurons because they produce beautiful filopodia that have a physiological role during brain development. A key challenge of working with primary cells rather than a cell line is that they can be unreliable – from week to week, I could either get more cells than I could handle or none at all. Therefore, I needed to be ready to make the most of a good week. On a more technical note, establishing two-colour imaging of TOCA-1 and Ena meant finding a difficult balance between imaging speed, brightness and photostability. This was overcome by a switch to mScarlet, which is a brighter red fluorescent protein.

When doing the research, did you have a particular result or ‘eureka’ moment that has stuck with you?

I remember watching the first video of a neuronal growth cone expressing TOCA-1 with Ena where I noticed that, as expected, both proteins localised to filopodia tips. As the filopodia extended, TOCA-1 suddenly retreated from the tip moving backwards and sideways along the adjacent lamellipodium before joining another filopodium. This observation led us to form the hypothesis that we explored in the paper. Also, it reminded me of how important it is to spend time observing and analysing your images before hurrying to quantification.

Filopodia dynamics are dramatically disrupted after inhibition of Cdc42 using CASIN, a small-molecule inhibitor, in Xenopus retinal ganglion cell axonal growth cones. Actin regulators TOCA-1 and Ena are lost from filopodia tips at the same time as filopodia tips stall, demonstrating their involvement in Cdc42-driven filopodia dynamics.

Filopodia dynamics are dramatically disrupted after inhibition of Cdc42 using CASIN, a small-molecule inhibitor, in Xenopus retinal ganglion cell axonal growth cones. Actin regulators TOCA-1 and Ena are lost from filopodia tips at the same time as filopodia tips stall, demonstrating their involvement in Cdc42-driven filopodia dynamics.

Why did you choose Journal of Cell Science for your paper?

The papers I read in Journal of Cell Science show the kind of interesting science I hope to do. The community building and conferences run by Journal of Cell Science, as well as the environmental commitment shown through the Forest of Biologists, have always made it stand out for me. We support open access publishing so the Read and Publish agreement our institution has with Journal of Cell Science made it an obvious choice.

Who are your role models in science? Why?

During my PhD, while I was studying the actomyosin system of the malaria parasite, I had the privilege of working with Lawrie Bannister, a legend in the malaria electron microscopy (EM) field. Despite being ‘retired’, Lawrie demonstrated an avid curiosity for untangling the complexities of the EM images we acquired, showing his encyclopaedic knowledge of the parasite ultrastructure. Lawrie taught me the value of carefully studying your images before analysing them, as well as exemplifying kindness and courtesy in science.

What's next for you?

I am excited to keep exploring the dynamics of how actin regulators combine, and pushing forward the image analysis tools we used in the paper.

Tell us something interesting about yourself that wouldn't be on your CV

In my spare time, I am renovating my house, which is not a small project as the house had no insulation or heating when we bought it. In time for last winter, we installed a heat pump on the roof and, with some trepidation, poured a new cement floor for the underfloor heating ourselves. Doing it ourselves (as much as possible), has taught me lots of new skills. Achieving each step (while making ethical and environmental choices where we can), has given me bursts of success which helps when science is hard!

Thomas Blake’s contact details: Wellcome/Cancer Research UK Gurdon Institute, Tennis Court Road, Cambridge, CB2 1QN, UK.

E-mail: t.blake@gurdon.cam.ac.uk

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2024
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Filopodial protrusion driven by density-dependent Ena–TOCA-1 interactions
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J. Cell Sci.
137
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jcs261057
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