Magdalena Winklhofer

• Winklhofer, Magdalena; Nilsson, Göran Erik & Lefevre, Sjannie (2026). Long non-coding RNA transcripts in crucian carp brain – annotation and expression patterns in anoxia and reoxygenation. RNA Biology. ISSN 1547-6286. 23(1), p. 1–21. doi: 10.1080/15476286.2026.2643366. Full text in Research Archive Show summary

  Long non-coding RNA transcripts (lncRNAs) are important regulators of various cellular processes, including gene expression. However, whether they are involved in the regulation of anoxia-induced transcriptomic changes in crucian carp ( Carassius carassius ), a species that can survive without oxygen for months at low temperatures, has remained unexplored. The existing genome annotation includes mainly protein-coding genes, which are more straightforward to annotate than the lower-expressed lncRNAs. Therefore, this study aimed to annotate lncRNAs in the crucian carp genome, investigate their potential involvement in regulating mRNA abundance in response to anoxia and reoxygenation, and further characterize any differentially regulated lncRNAs and their potential interaction partners (nearby genes). Using next-generation RNA sequencing of brain samples from crucian carp across normoxia, anoxia, and reoxygenation conditions (n=10 per group), 145,264 transcripts were assembled with a reference-guided approach. Using the bioinformatic tools CPC2, CNCI, CPAT, and FEELnc, we identified 6,072 lncRNAs among the assembled transcripts. We observed a substantial transcriptomic response to anoxia, with a total of 56,440 differentially expressed (DE) transcripts (adjusted p-value < 0.05; | log2(fold change)| > 0.38), including 1,321 lncRNAs. The FEELnc classification module identified multiple proximal RNA transcripts that may help to predict lncRNA functions, and due to their proximity to potential interaction partners (IPs), were likely cis -acting. Most (60%) of the interaction sites between DElncRNAs and their targets were located between genes (intergenic), predominantly on the same strand in intergenic subtypes, and primarily nested in the genic context. The log2-fold changes in RNA abundance of DE predicted IPs exhibited a positive correlation with the log2-fold changes of their corresponding DElncRNAs. These findings indicate that lncRNAs are at least co-expressed with and may regulate transcription of specific genes in crucian carp during anoxia and reoxygenation.

• Winklhofer, Magdalena; Andersen, Øivind & Lefevre, Sjannie (2025). Changes in DNA Methylation During Anoxia and Reoxygenation in Crucian Carp Brain. BioRxiv. ISSN 0362-4331. doi: 10.1101/2025.07.30.667361. Full text in Research Archive Show summary

  Oxygen is essential for cellular metabolism, and its absence poses a severe risk to vertebrates. However, the crucian carp ( Carassius carassius ) can survive without oxygen for months due to physiological adaptations. This study investigates the role of epigenomic regulation, specifically DNA methylation, in response to anoxia and subsequent reoxygenation. Brain samples from crucian carp subjected to normoxia, anoxia, and reoxygenation (n = 4 per condition) were analyzed through transcriptome (mRNA) and whole-genome bisulfite sequencing. Results revealed a notable transcriptional response to anoxia, with 1,265 differentially expressed genes (DEGs) identified compared to normoxia. Using the machine-learning-based MethylScore, we found 14 genes with differentially methylated regions (DMRs) during anoxia. These DMR-associated genes did not meet the strict DEGs criteria (adjusted p-value < 0.05; absolute log2fold-change > 0.38), but 17 genes exhibited differences in mRNA abundance when focusing solely on DMRs. DMRs were primarily located within transcriptional start sites, and analysis of protein products revealed that genes involved in transcription regulation, protein trafficking, and signal transduction were differentially methylated and expressed under varying conditions. This study advances our understanding of epigenomic regulation in oxygen deprivation and highlights the potential in exploring alternative epigenomic mechanisms.

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• Winklhofer, Magdalena & Lefevre, Sjannie (2024). DNA Methylation Alterations During Anoxia and Reoxygenation in Crucian Carp Brain. Full text in Research Archive
• Winklhofer, Magdalena & Lefevre, Sjannie (2024). Crucian carp’s fin-tastic tactics for anoxia survival - DNA methylation as an epigenetic switch to protect crucian carp during anoxia and reoxygenation. Full text in Research Archive
• Winklhofer, Magdalena (2022). Poster presentation at the Epigenetic Symposium in Oslo. Full text in Research Archive Show summary

  Epigenetic mechanisms during anoxia and re-oxygenation in crucian carp brain Crucian carp (Carassius carassius) overwinter in ice-covered lakes and can survive hypoxia and anoxia for several months, depending on the temperature. The physiological adaptations allowing this survival are well understood at the whole-organism level, but we know less about the molecular machinery coordinating the various elements of adaptation to anoxia and re-oxygenation, and the possible role of epigenetic mechanisms. Posttranslational modifications such as methylation, on either the histone tail or the globular domain, or both, can cause chromatin remodeling. Consequently, gene expression can be activated, altered or repressed solely depending on the epigenetic environment. Previous experiments analyzing whole brain transcriptome (RNAseq) data have shown that a large proportion of the transcriptome is regulated differentially in response to anoxia, and we suspect that at least some of these changes in gene transcription during anoxia are regulated by epigenetic mechanisms. By whole genome DNA sequencing before and after bisulfite conversion, we want to determine if DNA methylation loci are altered. Furthermore, the resulting data from methylation sequencing can be correlated with RNAseq data to investigate which specific genes are under epigenetic regulation. The obtained data will help to uncover the role of epigenetic mechanisms involved in response to anoxia and further guide us towards uncovering potentially novel molecular players in anoxia tolerance.

• Winklhofer, Magdalena (2022). Poster at the conference in Montpellier (05.07.2022 - 08.07.2022). Full text in Research Archive Show summary

  Epigenetic mechanisms during anoxia and re-oxygenation in crucian carp brain Crucian carp (Carassius carassius) overwinter in ice-covered lakes and can survive hypoxia and anoxia for several months, depending on the temperature. The physiological adaptations allowing this survival are well understood at the whole-organism level, but we know less about the molecular machinery coordinating the various elements of adaptation to anoxia and re-oxygenation, and the possible role of epigenetic mechanisms. Posttranslational modifications such as methylation, on either the histone tail or the globular domain, or both, can cause chromatin remodeling. Consequently, gene expression can be activated, altered or repressed solely depending on the epigenetic environment. Previous experiments analyzing whole brain transcriptome (RNAseq) data have shown that a large proportion of the transcriptome is regulated differentially in response to anoxia, and we suspect that at least some of these changes in gene transcription during anoxia are regulated by epigenetic mechanisms. By whole genome DNA sequencing before and after bisulfite conversion, we want to determine if DNA methylation loci are altered. Furthermore, the resulting data from methylation sequencing can be correlated with RNAseq data to investigate which specific genes are under epigenetic regulation. The obtained data will help to uncover the role of epigenetic mechanisms involved in response to anoxia and further guide us towards uncovering potentially novel molecular players in anoxia tolerance.

View all works in NVA