First person – Edismauro Freitas Filho

Edismauro Freitas Filho

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

RACK1 is a scaffold protein that acts as a platform for the binding of other proteins, thus facilitating or inhibiting their function. RACK1 acts as a central hub in diverse intracellular signaling pathways, and its function has been described in some cells of the immune system, such as lymphocytes and dendritic cells, but not in mast cells. Mast cells are specialized immune cells involved in allergic and inflammatory reactions. The physiological action of mast cells is a result of the release of their biologically active mediators. The factors controlling mediator release from mast cells are not well understood at a molecular level. In our study, we demonstrated that RACK1 is a novel modulator of mast cell mediator release due to its critical function in controlling actin cytoskeleton dynamics, which affects the mobilization of Ca²⁺. This is the first report to demonstrate that RACK1 integrates these crucial cellular events during antigen stimulation in mast cells. Our results likely represent common RACK1 regulatory themes that might also apply to other secretory cells. Moreover, our discovery of new RACK1 binding partners and the elucidation of their related functions and mechanisms may contribute to an improved understanding of the impact of RACK1 in various essential biological processes.

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

Trying to modulate the levels of RACK1 expression in our model, RBL-2H3 cells, was technically challenging. We made several adjustments to the RACK1-knockdown protocol, which is based on lentiviral transduction, before developing our final protocol. Based on our preliminary findings and on results described in the literature, we were concerned that a high level of reduction or complete ablation of RACK1 expression would result in cell death. Finding the ideal level of RACK1 knockdown that was sufficient to produce profound phenotypic effects in the knockdown cells while retaining viability was tricky. Furthermore, to induce reliable RACK1 interactions during mast cell antigen activation that could be used in co-immunoprecipitation experiments with endogenous RACK1 was laborious. We had to verify that the results of these experiments were reproducible and that the controls, sample acquisition and analysis were adequate.

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

While working on the initial reduction of RACK1 expression in RBL-2H3 cells, I observed that the overall morphological changes of mast cells were associated with a higher release of β-hexosaminidase, a pre-formed mediator stored in mast cell secretory granules. This was definitely a moment that convinced me that we were on the right track with our experimental hypothesis. The scaffold protein RACK1 had appeared during our functional proteomic analysis of raft-associated proteins in mast cells. The proteomic analysis showed that RACK1 was upregulated in raft microdomains that were disorganized owing to the aggregation of specific rodent mast cell gangliosides, a condition where the cells had their antigen-induced degranulation inhibited. This intriguing finding had stayed with me, and it gave me the incentive to investigate whether RACK1 could be involved in mast cell signal transduction and regulated secretion.

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

Since starting my postgraduate studies in cellular and molecular biology, I have considered Journal of Cell Science to be an outstanding journal due to its commitment to publishing significant advances across various areas of cell science. Our results encompass diverse fields of cell biology, which consequently makes our article an attractive paper for JCS readers.

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

Both Dr Maria Célia Jamur and Dr Constance Oliver fully supported me while I was a graduate student. They offered me the opportunity to learn from their expertise and perspectives, giving indispensable guidance for my scientific growth and as a person. Moreover, Dr Jamur and Dr Oliver taught me how to become an independent researcher and how to think outside the box, providing tools to test hypotheses.

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?

I grew up in close contact with nature on my grandparents' farm, and I have always been fascinated by trying to understand how biological systems operate. The most significant moments were those that challenged me further, such as presenting my data orally at international conferences, and a fellowship abroad that allowed me to work with diverse techniques and outstanding investigators, and receive awards. The sum of these moments has brought me to where I am today and given me the incentive to continue.

Who are your role models in science? Why?

All the LGBTQI+, women and minority researchers working hard to be heard by the scientific community.

What's next for you?

For the next few years, I intend to stay in academia in order to improve my research experience and develop my professional skills as an independent researcher, looking for qualified positions abroad as a scientist.

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

I am a determined person, so I try to use this trait in my creative side. I enjoy planning things that will positively surprise my relatives and friends.