Meet Pavel Baranov, Professor of Biomolecular Informatics at the University College Cork, Ireland. Pavel’s research group focuses on the understanding of how proteins are synthesised and how their synthesis is regulated.
Why did you choose to become a scientist?
When I was a toddler, I wanted to be a firefighter. Within a couple of years, I decided that being an astronaut would be more fun. A few more years passed, and I began to dream of becoming a scientist. I guess at that point I stopped growing and started living my dream.
What is your research focus?
My research group studies RNA translation. Translation is at the core of biology. Cells spend most of their energy on protein synthesis and the ribosome is the most abundant molecular machine in almost all cells. Ribosomal RNAs and tRNAs are the most conserved molecules across all kingdoms of life, and it is now apparent that proteins evolved earlier than DNA. Life as we know it relies on two main type of molecules not found outside of living systems – nucleic acids and proteins. It is the process of translation that connects these two chemistries together. I could hardly think of a more fundamental, interesting and challenging cellular process than translation.
Where do you see this field heading in the future?
As translation brings two chemistries together it is far more complex than other molecular process such as transcription and replication. Because of its complexity and the lack of tools to study it, studying translation is very challenging. The tools are now being developed, e.g. variations of ribosome profiling techniques, real-time single molecule imaging, cryo-EM microscopy, etc. The main change that I foresee is that translation will draw the attention of many more biomedical researchers, for better or worse.
What is your number one tip for people looking for scientific training?
Independent practice is the key in my opinion. After taking a course you may get the impression that you can do something, but it could be a false impression – you don’t really know if you can unless you have done it.
If you weren’t a scientist, what would you be?
Science is not a job for me, it is a dream. If I were not able to make my living as a researcher, I would have to find something else to make earnings, but I would not give up on my scientific interests.
The invention of ribosome profiling is the most significant development in the field of protein synthesis since the deciphering of the ribosome 3D structure. Ribosomal profiling is a popular technique for measuring the rate of translation in addition to measuring RNA levels, but this was somewhat possible even before. The unique ability of ribosome profiling is the detection of which open reading frames are being translated in RNA. The application of ribosome profiling revealed that even in eukaryotes the same mRNA molecule is often used for making more than one polypeptide, and that our current knowledge of the human genome protein coding repertoire is still far from complete. In addition to detecting translated frames, ribosome profiling could be used to detect ribosome pauses. We recently learned that such pauses could be used to regulate gene expression and other biological processes. This course will provide trainees with everything what is needed for mastering this powerful technology, from hands-on experience in generating ribosome profiling data to bioinformatics analysis and the use of public data resources.
The field of biology owes some of its most compelling discoveries to careful visual observation. From Van Leeuwenhoek’s use of new microscopes to describe microscopic “animalcules” in the late 1600s, to Ramon y Cajal’s pioneering 19th century work illustrating beautiful and complex neuronal architecture. Images inspire us, help us generate new hypotheses, and shed light into the tiny worlds yet unexplored. Indeed, these observations uniquely help us understand the structures and dynamics of life, something that would not be achievable with approaches like biochemistry alone.
The images are only as valuable as the amount of information that we can deduce from it.
Generating meaningful images, however, is not an easy task. There have always been limits to what we can observe, due to the properties of the sample or the techniques that we can apply to it. These are the boundaries that microscopists seek to push. A successful imaging experiment requires an amenable sample, a contrast agent to reveal the structures of interest, and a microscope that is capable of capturing an image at a relevant scale. Moreover, the images are only as valuable as the amount of information that we can deduce from it. Therefore, image storage, accessibility and analysis are crucial. Each one of these steps offers opportunities for optimisation and new technologies.
The EMBO | EMBL Symposium “Seeing is Believing: Imaging the Molecular Processes of Life” (9-12 October 2019) presented us with exciting new developments in all of these fields, coupled with a drive to make new progress available as quickly as possible to the community through preprints, open-source initiatives, and resource sharing.
Advances in sample preparation
At the heart of every imaging approach is the sample. Even the best microscope is ineffective with dim or improperly prepared samples. At Seeing is Believing, we saw an emphasis on using expansion of samples to help overcome the resolution limits of microscopy and solve some traditionally difficult problems. In particular, we were impressed with expansion-based approaches to study centriole structure (Paul Guichard, Ultrastructural Expansion Microscopy) and resolve microtubules tightly packed within axons (Lukas C. Kapitein). By far, the biggest emphasis in sample improvement was on the development of new fluorescent probes and biosensors. Kai Johnsson presented design strategies for the improvement of live cell dyes, and introduced new MaP dyes that are SNAP and HALO compatible, and importantly require no wash to clear unbound probe. Periklis Pantazis presented a mechanosensor based on the Piezo1 stretch activated ion channel, allowing users to visualise mechanical stress within a live cell. Atsushi Miyawaki wowed the audience by meeting the challenge to “be better than a firefly” with a new variant of luciferase named AkaBLI, which his lab generated through targeted evolution. This improved luciferase allowed them to visualise neuronal activity within freely behaving mice and marmosets.
Advances in microscopy
The features of our microscopes directly determine which questions we can address. Seeing is Believing highlighted exciting new development in building cutting-edge microscopy tools. Reto Fiolka presented a novel single-objective light-sheet microscope enabling imaging of live cells in microfluidics devices or 3D environments with 200 nm lateral resolution. Kevin Dean complemented novel light-sheet development by presenting an axially swept light-sheet microscope ideally suited for all clearing techniques that provides an unprecedented field of view enabling whole tissue imaging with sub-micron resolution. With her imaging approach, Alexandra Pacureanu surprised the audience with how X-ray holographic nano-tomography is capable of resolving the fine, dense and complex neuronal circuitry in large tissues or even organism providing a new route to understand how the nervous system processes information.
All acquired data is meaningless if we cannot extract information from it. At Seeing is Believing, it became obvious how artificial neuronal networks have become important for image analysis. Applications range from segmentation to denoising an image (BGnet, W.E. Moerner and Noise2Void, A. Krull/Florian Jug). Particularly, the convolutional network architecture U-Net has become an important tool. To provide a user-friendly environment to apply those state-of-the art image analysis tools, Anna Kreshuk presented the iLastik platform as an easy to use tool. A new fundamental approach to handle, visualise and process the large amount of data coming from the microscopes was presented by Ivo Sbalzarini. Instead of using pixels to save an image, adaptive particles approximate the image content. Furthermore, Gaudenz Danuser gave a thought-provoking talk on how current perturbation-based approaches in cell biology can mislead us in our analysis. Danuser emphasised that the observed phenotype from a perturbation of a system (e.g. loss of a protein’s function) is not equal to the real function of the gene. For example, cutting a wire from the battery to the electronic board of radio would lead to the “phenotype” loss of sound. However, the function of the wire was simply to provide power to the radio, not to produce sound! As a better perturbation-free alternative, Danuser introduced a concept used in econometrics known as Granger causality.
Advances in biology
All of these new developments culminated in impressive new insights into biological processes. There were many talks on mitochondria and endoplasmic reticulum dynamics revealed by novel live-cell super-resolution techniques. Suliana Manley gave one of the most intriguing of those talks, on modes of asymmetric and symmetric mitochondrial division.
Jennifer Lippincott-Schwartz also gave a stunning presentation on how RNA granules can hitch a ride through an ANXA-11 mediated connection to lysosomes, and how ALS associated mutations in ANXA-11 break this connection. Furthermore, an intriguing new mRNA reading frame sensor (Moon and Sun tags) was presented by Sanne Boersma of the Tanenbaum lab to understand stochasticity of mRNA translation.
To conclude, the field of microscopy has grown so much that some may feel we have solved all the theoretical problems, and only engineering challenges are left – hardware improvements, new materials, new engineering solutions. At the closing dinner of the conference, however, Atsushi Miyawaki from RIKEN beautifully summarised how he felt about the future of microscopy, and of Seeing is Believing. Standing in the banquet hall of the Heidelberg Castle, he told us that castles in Japan remain unfinished. This state of incompletion is not due to any fault of the architects, but a feature of beauty, as it was believed that things that were incomplete had room to grow, and that growth is valuable. No matter how high our achievements are in the field of microscopy and image analysis, there will always be unforeseen avenues of growth. Attending Seeing is Believing has hopefully prepared us to follow those avenues, and to share what we find so we may all grow together.