First person – Bina Prajapati Free

Bina Prajapati

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

The nucleus is the regulatory hub of the cell where DNA is stored and processes, such as DNA replication, regulation of gene expression and transcription, occur. Various proteins, including RNA polymerase II, transcription factors, cofactors and other enzymes, are involved in transcription, a process in which information from DNA is copied into mRNA. In this study, we were interested in understanding the function of myocardin-related transcription factors (MRTFs), which are cofactors of the serum response factor (SRF), in a cellular stress response mechanism known as the heat shock response (HSR). Dysregulation of MRTF and SRF is linked to cardiovascular diseases and invasive cancers. We found that MAL-D (the MRTF protein found in Drosophila melanogaster fruit flies) is necessary for activating heat shock protein (hsp) genes in fruit fly ovaries. However, in mammalian cells, MRTF is not required for hsp gene expression, and it does not bind hsp genes during heat shock. Additionally, we found that the well-known and frequently utilized MRTF-SRF inhibitor drug CCG-1423 and its derivative CCG-203971 actually have broader targets besides MRTF. These inhibitors reduced induction of hsp gene expression upon heat shock in both flies and mammalian cells, even in cells lacking the MRTF. Next, we found that CCG-1423-derived compounds inhibited the activity of SRF and an exogenous gene promoter system as well as reduced the expression of the ecdysone responsive gene, an important gene in a different response mechanism. The drugs also decreased overall RNA production while causing RNA polymerase II to accumulate at hsp gene sites under normal temperatures and interfered with the overall transcriptional machinery involved in the heat shock response, preventing the repression of genes other than hsp genes. Thus, we show that the CCG compounds inhibit general transcription and that in the future, these compounds can be used for studying transcription beyond the MRTF-SRF pathway.

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

Staining the total nascent RNA in small Drosophila cells with a large nucleolus was challenging. I performed several optimizations and time points for the 5-ethynyl uridine (5-EU) RNA labelling experiments. Imaging the cells required multiple trials due to the very bright nucleolus and nonspecific signals from the cytoplasm. To tackle these issues, I used positive and negative controls and optimized imaging parameters with excellent technical support from the Viikki Light Microscopy Core Unit personnel at the University of Helsinki. Performing precision run-on sequencing (PRO-seq) to map RNA polymerase II locations was lengthy and daunting, but I received clear instructions and extensive on-site and online help from our collaborator and mentor Anniina Vihervaara, even during the challenging COVID times.

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

The first eureka moment for me was when I found that CCG-1423 increases the basal level of hsp gene expression after 24 h of treatment. My second eureka moment was seeing the general effect of the drug manifested as an obvious decreased 5-EU signal in Drosophila cells under the microscope after we had speculated that CCG-1423 might not only be targeting MRTF or actin.

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

We chose Journal of Cell Science because it publishes good quality research articles and most of the actin-related articles that I have enjoyed reading are published in this journal. The review process is clear, and I also like that JCS supports open science for researchers from developing nations through the Electronic Information for Libraries (EIFL) initiative. I hope JCS will continue to participate in such novel causes.

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

My bench supervisor Maria Fülöp, who is also a shared co-first author of the paper (listed as Maria Sokolova), gets the most credit for her support and guidance. I have learnt almost everything from her. Next are all the lab members who have helped bring this work together, made it easier to communicate both in science and in life in general, and created a fun environment at work, making everyday life in the lab much easier. Our collaborator Anniina's support in establishing PRO-seq in the lab has also been crucial.

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?

My motivation to pursue a career in science began during secondary school when I learned about germ theory. I was fascinated by how something as minute as bacteria or viruses could have such detrimental effects on health and the world economy. This curiosity led me to pursue a bachelor's and a master's degree in microbiology along with my minor studies in genetics. Learning about the repertoire of cellular machinery and the mechanisms of gene regulation that keep us alive made me realize that I must understand more about cellular processes. A special moment early in my scientific career was attending my first international conference, the VIII International Congress on Stress Responses in Biology and Medicine, organized by Prof. Lea Sistonen and other esteemed scientists in Turku, Finland. This experience escalated my interest in studying heat shock and transcription machinery even more.

Who are your role models in science? Why?

My role models in science are Kary Mullis, the mind behind the globally used PCR technique, and Jennifer Doudna and Emmanuelle Charpentier, the scientists behind the discovery of CRISPR. I am fascinated by their genius and believe that their remarkable discoveries have significantly accelerated the field of science, diagnostics and therapies, especially for COVID-19 intervention. Additionally, Professor Dennis Bamford, my master's thesis supervisor Hanna Oksanen and my PhD supervisor Maria Vartiainen have greatly influenced my scientific journey. They have allowed me to grow as a scientist at my own pace and taught me the importance of maintaining work–life balance.

What's next for you?

I am finishing my PhD and have started working as a laboratory coordinator in the Finnish Genome Editing Centre at the University of Helsinki. I am open to learning newer techniques in genome editing that could be translated into clinical therapies. At some point in life, I wish to have my own research lab in my home country, Nepal.

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

I am enthusiastic about plants, bonsai culture and self-sufficient gardening. I enjoy meditating, watching thought-provoking movies, strolling through nature and engaging in pottery-making and upcycling. Plants are a source of life lessons. Waiting for my orchids and peace lilies to bloom and battling bug infestations has taught me patience and resilience. As the saying goes, “As the seed buried in the earth cannot imagine itself as an orchid or hyacinth, neither can a heart packed with hurt imagine itself loved or at peace. The courage of the seed is that once cracking, it cracks all the way” (Mark Nepo). This verse resonates with our paper. We demonstrated that the inhibitors in our study have broader implications than previously shown and we dared to pursue what initially seemed like a bizarre result.