Gene expression control by RNA pseudouridylation in response to metabolic stress

Popular Abstract

PUS7 role in cellular adaptation to metabolic stress

When we hear the word metabolism, our minds often jump to calories and diets. Cellular metabolism however refers to all essential mechanisms that keep our body up and running. Unsurprisingly, complex networks navigate this sensitive system, with many pathways that allow adaptation to changing and stressful environments. If this delicate balance is disturbed, diseases like cancer and diabetes ensue. But, what, or who, regulates these pathways?

While the notion that DNA carries all genetic information in the cell is well established, little emphasis has been put on its temporal successor: RNA. RNA can exist in various forms, and external signals often trigger chemical changes that alter its structure and function. Dynamic regulation of RNA modification acts as a translator between the cell and its environment to ensure adaptation to harsh conditions, with dysregulation of these mechanisms often resulting in disease. RNA modifications have become prominent suspects in the hunt for understanding metabolic regulation. An example of RNA modification is Pseudouridine (ψ), a chemical variant of Uridine. Enzymes like PUS7 are responsible for these modifications. While PUS7 has been linked to regulating gene expression as a response to stress stimuli, its function in adaptation to metabolic stress remains largely unknown. So, in this thesis, we wanted to investigate whether PUS7 plays a role in withstanding cellular starvation.

To answer this question, fibroblast cells harvested from mouse embryos were put on a diet. After depriving both normal and PUS7-deficient cells of essential nutrients for three days, three aspects were studied: cell death, protein synthesis and activation of a stress response pathway. Interestingly, we found that when the ψ-writer is not present, cells show increased cell death and decreased ability to activate the cell’s integrated stress response, a pathway allowing our cells to switch from growth to survival. Hence, these findings seem to point towards a decreased ability to withstand starvation when PUS7 is depleted, supporting the notion that PUS7 may be a regulator in the cell’s response to nutrient deprivation.

Studying RNA modifications. To understand how PUS7 impacts cellular metabolism by ψ, reliable detection methods are needed. But how do we study RNA modifications? So far, methods requiring either costly machinery or complex analysis methods are set as the gold standard for ψ detection. Therefore, we wanted to find a simple and fast alternative. For this, we experimented with one of the most basic methods in molecular biology: quantitative PCR - a method that, at its very basis, allows ‘counting’ of DNA and cDNA (RNA converted to DNA) by use of fluorescent markers interacting with double stranded, but not single stranded DNA. First, a previously established chemical treatment that makes gaps at ψ sites was used. Then, short DNA sequences were designed that end directly at this gap, thereby disturbing binding when modification is present and in turn, less fluorescent signal is obtained. Put simply, decreased fluorescence signals presence of modification. This idea was tested, and results showed consistent 40-60% reduction of signal in modified samples, thereby providing proof that this system works. While more work is needed to make this system fully applicable, this novel principle could potentially be the basis of a simple and cost-efficient alternative to current ψ detection methods that can be employed across systems and labs.

More work will be necessary to understand what role PUS7-mediated ψ plays in regulating cellular metabolism. Studying these pathways will give insight in how dysregulation of these processes contributes to human disease and potentially inspire novel therapeutic targets to fight them.

Master’s Degree Project in Molecular Biology 60 credits 2025
Department of Biology, Lund University
Advisor: Cristian Bellodi, Division of Molecular Hematology, BMC, Faculty of Medicine, Lund University

• author: Colazzo, Anita
• supervisor: Cristian Bellodi
• organization: Degree Projects in Molecular Biology
• course: MOBN03 20242
• year: 2025
• type: H2 - Master's Degree (Two Years)
• subject: Biology and Life Sciences
• language: English
• id: 9213482
• date added to LUP: 2025-10-02 16:00:19
• date last changed: 2025-10-02 16:00:19