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RNA modifications have recently emerged as critical posttranscriptional regulators of gene expression programs. They affect diverse eukaryotic biological processes, and the correct deposition of many of these modifications is required for normal development. Messenger RNA (mRNA) modifications regulate various aspects of mRNA metabolism. For example, N 6 -methyladenosine (m 6 A) affects the translation and stability of the modified transcripts, thus providing a mechanism to coordinate the regulation of groups of transcripts during cell state maintenance and transition. Similarly, some modifications in transfer RNAs are essential for RNA structure and function. Others are deposited in response to external cues and adapt global protein synthesis and gene-specific translational accordingly and thereby facilitate proper development.

Aggressive and metastatic cancers show enhanced metabolic plasticity 1 , but the precise underlying mechanisms of this remain unclear. Here we show how two NOP2/Sun RNA methyltransferase 3 (NSUN3)-dependent RNA modifications—5-methylcytosine (m 5 C) and its derivative 5-formylcytosine (f 5 C) (refs. 2 – 4 )—drive the translation of mitochondrial mRNA to power metastasis. Translation of mitochondrially encoded subunits of the oxidative phosphorylation complex depends on the formation of m 5 C at position 34 in mitochondrial tRNA Met . m 5 C-deficient human oral cancer cells exhibit increased levels of glycolysis and changes in their mitochondrial function that do not affect cell viability or primary tumour growth in vivo; however, metabolic plasticity is severely impaired as mitochondrial m 5 C-deficient tumours do not metastasize efficiently. We discovered that CD36-dependent non-dividing, metastasis-initiating tumour cells require mitochondrial m 5 C to activate invasion and dissemination. Moreover, a mitochondria-driven gene signature in patients with head and neck cancer is predictive for metastasis and disease progression. Finally, we confirm that this metabolic switch that allows the metastasis of tumour cells can be pharmacologically targeted through the inhibition of mitochondrial mRNA translation in vivo. Together, our results reveal that site-specific mitochondrial RNA modifications could be therapeutic targets to combat metastasis.

Background Adenosine-to-inosine (A-to-I) RNA editing is a process that contributes to the diversification of proteins that has been shown to be essential for neurotransmission and other neuronal functions. However, the spatiotemporal and diversification properties of RNA editing in the brain are largely unknown. Here, we applied in situ sequencing to distinguish between edited and unedited transcripts in distinct regions of the mouse brain at four developmental stages, and investigate the diversity of the RNA landscape. Results We analyzed RNA editing at codon-altering sites using in situ sequencing at single-cell resolution, in combination with the detection of individual ADAR enzymes and specific cell type marker transcripts. This approach revealed cell-type-specific regulation of RNA editing of a set of transcripts, and developmental and regional variation in editing levels for many of the targeted sites. We found increasing editing diversity throughout development, which arises through regional- and cell type-specific regulation of ADAR enzymes and target transcripts. Conclusions Our single-cell in situ sequencing method has proved useful to study the complex landscape of RNA editing and our results indicate that this complexity arises due to distinct mechanisms of regulating individual RNA editing sites, acting both regionally and in specific cell types.

The human genome is under constant invasion by retrotransposable elements. The most successful of these are the Alu elements; with a copy number of over a million, they occupy about 10 % of the entire genome. Interestingly, the vast majority of these Alu insertions are located in gene-rich regions, and one-third of all human genes contains an Alu insertion. Alu sequences are often embedded in gene sequence encoding pre-mRNAs and mature mRNAs, usually as part of their intron or UTRs. Once transcribed, they can regulate gene expression as well as increase the number of RNA isoforms expressed in a tissue or a species. They also regulate the function of other RNAs, like microRNAs, circular RNAs, and potentially long non-coding RNAs. Mechanistically, Alu elements exert their effects by influencing diverse processes, such as RNA editing, exonization, and RNA processing. In so doing, they have undoubtedly had a profound effect on human evolution.

How chronic mutational processes and punctuated bursts of DNA damage drive evolution of the cancer genome is poorly understood. Here, we demonstrate a strategy to disentangle and quantify distinct mechanisms underlying genome evolution in single cells, during single mitoses and at single-strand resolution. To distinguish between chronic (reactive oxygen species (ROS)) and acute (ultraviolet light (UV)) mutagenesis, we microfluidically separate pairs of sister cells from the first mitosis following burst UV damage. Strikingly, UV mutations manifest as sister-specific events, revealing mirror-image mutation phasing genome-wide. In contrast, ROS mutagenesis in transcribed regions is reduced strand agnostically. Successive rounds of genome replication over persisting UV damage drives multiallelic variation at CC dinucleotides. Finally, we show that mutation phasing can be resolved to single strands across the entire genome of liver tumors from F1 mice. This strategy can be broadly used to distinguish the contributions of overlapping cancer relevant mutational processes. A single-cell-based approach allows the daughters of a damaged cell to be separately tracked following single mitotic events. This technique highlights the different ways in which ultraviolet light and reactive oxygen species cause mutagenesis.

Ansell's mole-rat (Fukomys anselli) is an African rodent known for its subterranean lifestyle and unique phenotypic traits, including extreme longevity, magnetoreception, and a cooperative breeding social structure. Efforts to dissect the genetic architecture of these traits and to decipher their phylogenetic relationships within the broader African mole-rat family would greatly benefit from a reference-grade genome. Here, we report a first genome assembly of a male Ansell's mole-rat. By combining Oxford Nanopore Technologies long reads and Illumina short reads with Hi-C data, we generated a chromosome level assembly with a total length of 2.27 Gb, 412 scaffolds, and a scaffold N50 of 72.4 Mb. We identified 99.54% of expected genes and annotated 29,094 transcripts using RNA sequencing data. This high-quality de novo genome of F. anselli lays the foundation for dissecting the genetic and evolutionary basis of its extraordinary traits and resolving African mole-rat phylogeny.

Ansell’s mole-rat (Fukomys anselli) is an African rodent known for its subterranean lifestyle and unique phenotypic traits, including extreme longevity, magnetoreception, and a cooperative breeding social structure. Efforts to dissect the genetic architecture of these traits and to decipher their phylogenetic relationships within the broader African mole-rat family would greatly benefit from a reference-grade genome. Here, we report a first genome assembly of a male Ansell’s mole-rat. By combining Oxford Nanopore Technologies (ONT) long-reads and Illumina short-reads with Hi-C data, we generated a chromosome level assembly with a total length of 2.27 Gb, 412 scaffolds and a scaffold N50 of 72.4 Mb. We identified 99.54% of expected genes and annotated 29,094 transcripts using RNA sequencing data. This high-quality de novo genome of Fukomys anselli lays the foundation for dissecting the genetic and evolutionary basis of its extraordinary traits and resolving African mole-rat phylogeny. Bekavac, Coimbra, and Busa et al. assemble a de novo genome of a male Fukomys anselli individual. They use short-read and long-read whole genome sequencing and Hi-C sequencing to achieve chromosome-level contiguity, and annotate the genome using RNA-seq from nine diverse tissues. This high-quality genome enables exploration of the genetic basis of Ansell’s mole-rat’s remarkable traits, including longevity, hypoxia tolerance, and magnetoreception. It also allows for comprehensive analysis of the currently-contested phylogenetic relationships within the mole-rat family.

Long-lived species suppress cancer despite accumulating somatic mutations throughout life, but how this is achieved in renewing tissues remains unclear. Here, we show that naked molerat skin uncouples high cellular turnover from cancer risk through coordinated epithelial and stromal mechanisms that constrain clonal outgrowth and suppress tumor-promoting inflammation. Despite elevated epidermal proliferation and mutational burden, naked mole-rat skin maintains tissue integrity through an expanded pool of early committed progenitor (hybrid) cells and replication-coupled genome maintenance pathways. Under chronic carcinogenic stress, undifferentiated basal cells replenish the hybrid progenitor pool. This dilutes initiated clones and permits only limited selective expansion of rare cancer gene-mutant clones. Depletion of the hybrid compartment shifts this protective state toward clonal expansion and inflammatory activation. These expanding clones are further constrained by a highly tumor-suppressive stromal microenvironment, driven by fibroblasts that adopt a metabolically restricted, non-inflammatory program. Together, our data uncover a multilayered tumor-suppressive strategy that couples turnover-driven regeneration with an anti-permissive stromal niche to prevent malignant progression under mutagenic stress.Competing Interest StatementThe authors have declared no competing interest.Footnotes* This version of the manuscript has been revised to incorporate additional experiments and analyses that strengthen the conclusions. The authorship and acknowledgments have been updated.

Human cancers are heterogeneous. Their genomes evolve from genetically diverse germlines in complex and dynamic environments, including exposure to potential carcinogens. This heterogeneity of humans, our environmental exposures, and subsequent tumours makes it challenging to understand the extent to which cancer evolution is predictable. Addressing this limitation, we re-ran early tumour evolution hundreds of times in diverse, inbred mouse strains, capturing genetic variation comparable to and beyond that found in human populations. The sex, environment, and carcinogenic exposures were all controlled and tumours comprehensively profiled with whole genome and transcriptome sequencing. Within a strain, there was a high degree of consistency in the mutational landscape, a limited range of driver mutations, and all strains converged on the acquisition of a MAPK activating mutation with similar transcriptional disruption of that pathway. Despite these similarities in the phenotypic state of tumours, different strains took markedly divergent paths to reach that state. This included pronounced biases in the precise driver mutations, the strain specific occurrence of whole genome duplication, and differences in subclonal selection that reflected both cancer susceptibility and tumour growth rate. These results show that interactions between the germline genome and the environment are highly deterministic for the trajectory of tumour genome evolution, and even modest genetic divergence can substantially alter selection pressures during cancer development, influencing both cancer risk and the biology of the tumour that develops.Competing Interest StatementS.J.A. receives funding from AstraZeneca for a PhD studentship. J.C. has received an honorarium from Roche Diagnostics. P.F. is a member of the Scientific Advisory Board of Fabric Genomics, Inc..