First person – Kristina Ehring Free

Kristina Ehring

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

My group investigates early developmental processes during embryogenesis. More specifically, we focus on a protein called Hedgehog (Hh), which is essential for many steps of vertebrate and invertebrate development. The main challenge for Hh proteins is their release from the cells that produce them, which is a prerequisite for Hh proteins to reach and signal to their target cells. This is because Hh proteins have two lipids attached to their N- and C-termini that tightly tether them to the plasma membrane of their producing cells, possibly within areas of high local cholesterol concentration. By investigating proteins known to be involved in the Hh release process, I found that a protein called Dispatched (Disp) induces Hh release by changing the cholesterol content or its distribution in the plasma membrane. These changes, in turn, facilitate proteolytic processing of both lipidated Hh peptides and Hh release from producing cells.

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

One specific challenge was right at the beginning of my project when I aimed to generate a CRISPR/Cas9-mediated Disp-knockout cell line. In our lab, we have based our previous studies on Bosc23 cells, which are HEK293 cell derivates, and I intended to continue to use this line. However, during the process I realized that these cells contain three Disp loci, which made it extremely difficult to generate a complete Disp-knockout cell line. Still, after sequencing and testing many independent cell clones that continue to fill our liquid nitrogen tank, I obtained and confirmed two cell lines and could finally start with the more interesting and less tedious part of my work.

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

For me, this ‘eureka’ moment happened when I first discovered accumulated cholesterol levels in my Disp-knockout cell line. At this point in time, Disp function had never been connected to free cholesterol levels within Hh-producing cells but had rather been suggested to extract lipidated Hh from the plasma membrane. But more or less at the same time, a protein with a structure closely related to the Disp structure, called Patched, was also described to reduce inner membrane leaflet cholesterol. This supported the functional Disp mechanism that I found. After the exciting observation that loss of Disp increases cellular cholesterol levels, it felt like I was on the right track, and the motivation was high to pursue this direction further. I did this by rescuing the Disp deficiency through Patched transfection and by directly proving reduced export of tritium-labelled cholesterol from my Disp-deficient cell line.

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

This is my first time publishing in JCS, but in previous years the experience of former PhD students from my lab during the submission and review process had always been very good. Like them, I considered the editorial handling extremely efficient and the choice of reviewers and their requests very fair, especially given that my findings are far from current thinking in the field. After acceptance, the production department communicated with us immediately and handled the publication process in a fast and professional manner. But of course, we also chose JCS because its ‘Cell Biology of Lipids’ special issue seemed like a perfect fit for my work.

Have you had any significant mentors who have helped you beyond supervision in the lab? How was their guidance special?

A very important mentor for me during my PhD was Professor Kay Grobe. He always supported and motivated me to work on this wonderful project. He had great ideas to improve our story and always helped with arising challenges. But everybody else in the lab was equally important for my work. Dr Dominique Manikowski gave significant input for new experiments and experimental setups, and helped with their realization. Sabine Kupich and Reiner Schulz were always great support in the lab, especially with all the western blots and cloning work. Finally, my two current PhD student colleagues Jurij Froese and Fabian Gude helped with performing various experiments and motivated me throughout the project.

What motivated you to pursue a career in science, and what have been the most interesting moments on the path that led you to where you are now?

When I started my PhD, I wanted to follow a career in science because I was fascinated by how little is known about so many areas of our lives, and I therefore wanted to play a part in discovering something completely new. Additionally, I hoped that my research could someday help to better understand a specific disease and to develop drugs to improve the lives of people suffering from it. Along the path of my PhD, the interesting moments were always meeting other people who were so passionate about their work that they inspired and motivated me to be the same. The pandemic made this part especially difficult, just like for everybody else, but fortunately did not abolish it altogether.

Who are your role models in science? Why?

I don't have a specific role model. My role models are many: all the scientists I got to know who show such great enthusiasm for their work, and that together inspire their students and make them excited about their projects, too. Additionally, it is very encouraging for me to observe other scientists who never give up and who motivate themselves even in sometimes unsatisfying times.

What's next for you?

First, I am planning on finishing my PhD soon. Because I find the topic of my thesis very exciting and I enjoyed working on it very much, maybe I will continue to work in my lab for a little longer. However, because I learned during my PhD that there are numerous options and often surprising new roads opening up for me – I never thought when I started my PhD that I would eventually do research in the field of cholesterol and plasma membrane biology! – I am pretty open to many options that might arise for me in the future.

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

Thinking back to my early childhood, I already recognized my love of nature and all living creatures. I especially remember moments in kindergarten, where I always rescued bugs and spiders with great interest and without keeping my distance. At that time, it was already fun for me to observe nature, although my fellow kids and teachers considered this behaviour somewhat funny. From today's perspective, this was a clear early indication that the study of nature would be my future destination.