The interplay between metabolic pathways and the epigenome is essential for proper cell differentiation. In this new study, Francisco Naya and colleagues find that the Dlk1-Dio3 noncoding RNA (ncRNA) locus regulates cell state by coordinating mitochondrial activity and histone modifications in muscle cells. To find out more about the people behind the work, we caught up with first author Amanda Pinheiro and corresponding author Francisco (Frank) Naya, Associate Professor at the Department of Biology, Boston University, USA.
Francisco Naya (left) and Amanda Pinheiro (right)
Frank, can you give us your scientific biography and the questions your lab is trying to answer?
FN: I have had a long-standing interest in cell differentiation and how specialized cells acquire their distinct structural and functional properties. Along these lines, my PhD training focused on understanding the mechanisms by which the hormone insulin is expressed in a tissue-specific manner. In my post-doctoral training, I expanded my knowledge in tissue-specific gene regulation by investigating an evolutionarily conserved and important muscle transcription factor, MEF2, in cardiac and skeletal muscle differentiation and development. As a Principal Investigator, I have continued investigating gene regulatory mechanisms in muscle differentiation not only to gain insight into the pathways that drive this process, but to better understand how this regulation breaks down in muscle diseases.
Amanda, how did you come to work in the lab and what drives your research today?
AP: My research background began with a strong focus on tissue engineering and muscle physiology, which laid the foundation for my PhD. During my doctoral studies, I wanted to explore the gene regulatory mechanisms that orchestrate muscle development and regeneration, making Frank's lab a great fit for my interests. What drives my research is a deep curiosity about the complexities of the human body and the desire to contribute to the body of knowledge that will translate fundamental scientific discoveries into innovative therapies that can improve lives.
What is the background of the field that inspired your work?
FN: Through the lab's work on the muscle transcription factor MEF2, we identified the Dlk1-Dio3 noncoding RNA cluster as a key target gene in skeletal muscle differentiation and regeneration. While individual ncRNAs from the cluster were the initial focus in our lab and others, our lab sought to understand the collective role of this complex noncoding RNA cluster in skeletal muscle differentiation. Based on previous studies suggesting a role for this ncRNA cluster in metabolic regulation, we considered that this locus could play an essential role in coordinating metabolic activity and differentiation, which is crucial for the proper execution of this process.
Can you give us the key results of the paper in a paragraph?
FN: Our research highlighted the central role of the Dlk1-Dio3 noncoding RNAs in skeletal muscle differentiation through its coordinate regulation of mitochondrial respiration and chromatin structure. We found that changes in dosage of Dlk1-Dio3 ncRNAs adversely affect muscle differentiation. In addition, these altered ncRNA levels have a profound impact on oxidative phosphorylation (respiration), chromatin accessibility, and histone modifications in both proliferating myoblasts and differentiated myotubes, resulting in altered cell states with abnormal gene expression patterns.
Amanda, when doing the research, did you have any particular result or eureka moment that has stuck with you?
AP: I find joy in celebrating the results of every experiment, as each discovery contributes a vital piece to the larger puzzle. However, one moment that stands out was seeing the results from the western blots for the mitochondrial oxidative phosphorylation complexes. It felt like a true eureka moment because the experiment was particularly time-intensive and required extensive troubleshooting, making the results all the more rewarding.
It felt like a true eureka moment because the experiment was particularly time-intensive and required extensive troubleshooting, making the results all the more rewarding
Immunofluorescence image analysis of muscle differentiation in vitro. The early stages of C2C12 differentiation reveal the fusion of myoblasts to form nascent multinucleated and elongated myotubes. Antibodies were used to detect the muscle structural proteins alpha-actinin 2 (green) and myosin heavy chain 4 (MYH4) (red), and nuclei (DAPI; blue). Individual images were merged to construct this image.
Immunofluorescence image analysis of muscle differentiation in vitro. The early stages of C2C12 differentiation reveal the fusion of myoblasts to form nascent multinucleated and elongated myotubes. Antibodies were used to detect the muscle structural proteins alpha-actinin 2 (green) and myosin heavy chain 4 (MYH4) (red), and nuclei (DAPI; blue). Individual images were merged to construct this image.
And what about the flipside: any moments of frustration or despair?
AP: The research journey inevitably comes with its share of frustrations. I recall one weekend spent working long hours on an immunocytochemistry experiment, only to discover late at night that the fluorescent signal on my slides was too weak. Knowing I'd have to start over was frustrating, but after some additional troubleshooting, I managed to get the results I needed.
Why did you choose to submit this paper to Development?
FN: Knowing that Development publishes groundbreaking findings in the areas of cell differentiation and epigenetics, we felt that our research would be a perfect fit with the research topics in the journal.
Amanda, what is next for you after this paper?
AP: Now that I have completed my PhD, I am working as an Entrepreneurial Fellow at the Chicago Biomedical Consortium. In this role, I focus on advancing biomedical therapeutics, medical devices, and other innovations originating from Chicago-based university labs, helping to bridge the gap between academic research and commercialization. My work involves evaluating the commercial potential of innovative technologies, supporting the development of business strategies, and collaborating with researchers, investors, and industry partners to drive these projects forward.
Frank, where will this story take your lab next?
FN: Based on the results of the present study, my lab is now keenly interested in identifying the specific metabolites affected in Dlk1-Dio3 ncRNA mutant muscle cells and the epigenetic factors that function downstream of this ncRNA cluster to remodel chromatin to establish gene expression patterns for proper cell differentiation.
Finally, let's move outside the lab – what do you like to do in your spare time?
AP: Outside the lab, I enjoy cooking different cuisines, traveling the world, and exercising.
FN: Outside the lab, I try to keep my mind and body active. For my mind, apart from the research, I enjoy solving challenging crossword puzzles. For my body, I am an avid cyclist.
A.P. & F.N.: Program in Molecular Biology, Cell Biology, and Molecular Biology, Boston University, Boston, MA 02215, USA.
F.N.: Department of Biology, Program in Cell and Molecular Biology, Boston University, Boston, MA 02215, USA.
