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Chronically high stress hormone levels dysregulate sperm long noncoding RNAs and their embryonic microinjection alters development and affective behaviours

Abstract

Previous studies on paternal epigenetic inheritance have shown that sperm RNAs play a role in this type of inheritance. The microinjection of sperm small noncoding RNAs into fertilised mouse oocytes induces reprogramming of the early embryo, which is thought to be responsible for the differences observed in adult phenotype. While sperm long noncoding RNAs (lncRNAs) have also been investigated in a previous study, their microinjection into fertilised oocytes did not yield conclusive results regarding their role in modulating brain development and adult behavioural phenotypes. Therefore, in the current study we sought to investigate this further. We used our previously established paternal corticosterone (stress hormone) model to assess sperm lncRNA expression using CaptureSeq, a sequencing technique that is more sensitive than the ones used in other studies in the field. Paternal corticosterone exposure led to dysregulation of sperm long noncoding RNA expression, which encompassed lncRNAs, circular RNAs and transposable element transcripts. Although they have limited functional annotation, bioinformatic approaches indicated the potential of these lncRNAs in regulating brain development and function. We then separated and isolated the sperm lncRNAs and performed microinjections into fertilised oocytes, to generate embryos with modulated lncRNA populations. We observed that the resulting adult offspring had lower body weight and altered anxiety and affective behavioural responses, demonstrating roles for lncRNAs in modulating development and brain function. This study provides novel insights into the roles of lncRNAs in epigenetic inheritance, including impacts on brain development and behaviours of relevance to affective disorders.

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Fig. 1: Long noncoding RNA capture sequencing data analysis.
Fig. 2: Investigation of other long RNA types, and cis-target prediction.
Fig. 3: Analysis of transposable elements in the promoter regions of up and down regulated lncRNAs.
Fig. 4: Experimental design and offspring body weight.
Fig. 5: Assessment of anxiety- and depressive-like behaviours.
Fig. 6: Assessment of social behaviours.

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Data availability

The sequencing datasets generated and analysed during the current study have been deposited in the European Nucleotide Archive (ENA) repository at EMBL‐EBI under accession number PRJEB66376 (https://www.ebi.ac.uk/ena/browser/view/PRJEB66376). All other relevant data not included in the published article will be made available upon reasonable request.

References

  1. Gapp K, Soldado-Magraner S, Alvarez-Sánchez M, Bohacek J, Vernaz G, Shu H, et al. Early life stress in fathers improves behavioural flexibility in their offspring. Nat Commun. 2014b;5:5466.

    Article  PubMed  Google Scholar 

  2. Rodgers AB, Morgan CP, Bronson SL, Revello S, Bale TL. Paternal stress exposure alters sperm microRNA content and reprograms offspring HPA stress axis regulation. J. Neurosci. 2013;33:9003–12.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Short AK, Fennell KA, Perreau VM, Fox A, O’Bryan MK, Kim JH, et al. Elevated paternal glucocorticoid exposure alters the small noncoding RNA profile in sperm and modifies anxiety and depressive phenotypes in the offspring. Transl Psychiatry. 2016;6:e837.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. van Steenwyk G, Roszkowski M, Manuella F, Franklin TB, Mansuy IM. Transgenerational inheritance of behavioral and metabolic effects of paternal exposure to traumatic stress in early postnatal life: evidence in the 4th generation. Environ Epigenetics. 2018;4:1–8.

    Google Scholar 

  5. Rodgers AB, Morgan CP, Leu NA, Bale TL. Transgenerational epigenetic programming via sperm microRNA recapitulates effects of paternal stress. Proc Natl Acad Sci. 2015;112:13699–704.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Gapp K, Jawaid A, Sarkies P, Bohacek J, Pelczar P, Prados J, et al. Implication of sperm RNAs in transgenerational inheritance of the effects of early trauma in mice. Nat Neurosci. 2014a;17:667–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Sun YH, Wang A, Song C, Shankar G, Srivastava RK, Au KF, et al. Single-molecule long-read sequencing reveals a conserved intact long RNA profile in sperm. Nat Commun. 2021;12:1361.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Schertzer MD, Braceros KCA, Starmer J, Cherney RE, Lee DM, Salazar G, et al. lncRNA-induced spread of polycomb controlled by genome architecture, RNA abundance, and CpG island DNA. Mol Cell. 2019;75:523–37.e10.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Statello L, Guo C-J, Chen L-L, Huarte M. Gene regulation by long non-coding RNAs and its biological functions. Nat Rev Mol Cell Biol. 2021;22:96–118.

    Article  CAS  PubMed  Google Scholar 

  10. Gapp K, van Steenwyk G, Germain PL, Matsushima W, Rudolph KLMM, Manuella F, et al. Alterations in sperm long RNA contribute to the epigenetic inheritance of the effects of postnatal trauma. Mol Psychiatry. 2018;25:2162–74.

    Article  PubMed  PubMed Central  Google Scholar 

  11. Mercer TR, Clark MB, Crawford J, Brunck ME, Gerhardt DJ, Taft RJ, et al. Targeted sequencing for gene discovery and quantification using RNA CaptureSeq. Nat Protoc. 2014;9:989–1009.

    Article  CAS  PubMed  Google Scholar 

  12. Tada H, Miyazaki T, Takemoto K, Takase K, Jitsuki S, Nakajima W, et al. Neonatal isolation augments social dominance by altering actin dynamics in the medial prefrontal cortex. Proc Natl Acad Sci. 2016;113:E7097–E7105.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Zhou YY, Kaiser T, Monteiro P, Zhang X, Van der Goes MS, Wang D, et al. Mice with Shank3 mutations associated with ASD and schizophrenia display both shared and distinct defects. Neuron. 2016;89:147–62.

    Article  CAS  PubMed  Google Scholar 

  14. Hoffmann LB, Rae M, Marianno P, Pang TY, Hannan AJ, Camarini R. Preconceptual paternal environmental stimulation alters behavioural phenotypes and adaptive responses intergenerationally in Swiss mice. Physiol Behav. 2020;223:112968.

    Article  CAS  PubMed  Google Scholar 

  15. Mitra R, Sapolsky RM. Short-term enrichment makes male rats more attractive, more defensive and alters hypothalamic neurons. PLoS One. 2012;7:e36092.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Boerke A, Dieleman SJ, Gadella BM. A possible role for sperm RNA in early embryo development. Theriogenology. 2007;68:147–55.

    Article  Google Scholar 

  17. Wei W, Zhao Q, Wang Z, Liau W-S, Basic D, Ren H, et al. ADRAM is an experience-dependent long noncoding RNA that drives fear extinction through a direct interaction with the chaperone protein 14-3-3. Cell Rep. 2022;38:110546.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Martin M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet.journal. 2011;17:10.

    Article  Google Scholar 

  19. Kim D, Paggi JM, Park C, Bennett C, Salzberg SL. Graph-based genome alignment and genotyping with HISAT2 and HISAT-genotype. Nat Biotechnol. 2019;37:907–15.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, et al. The sequence alignment/map format and SAMtools. Bioinformatics. 2009;25:2078–9.

    Article  PubMed  PubMed Central  Google Scholar 

  21. Pertea M, Kim D, Pertea GM, Leek JT, Salzberg SL. Transcript-level expression analysis of RNA-seq experiments with HISAT, StringTie and Ballgown. Nat Protoc. 2016;11:1650–67.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Robinson MD, McCarthy DJ, Smyth GK. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics. 2010;26:139–40.

    Article  CAS  PubMed  Google Scholar 

  23. Chen Y, Lun ATL, Smyth GK. From reads to genes to pathways: differential expression analysis of RNA-Seq experiments using Rsubread and the edgeR quasi-likelihood pipeline. F1000Research. 2016;5:1438.

    PubMed  PubMed Central  Google Scholar 

  24. Wickham H. ggplot2: elegant graphics for data analysis. New York: Springer-Verlag; 2016.

  25. Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, et al. Gene Ontology: tool for the unification of biology. Nat Genet. 2000;25:25–29.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Mi H, Muruganujan A, Ebert D, Huang X, Thomas PD. PANTHER version 14: more genomes, a new PANTHER GO-slim and improvements in enrichment analysis tools. Nucleic Acids Res. 2019;47:D419–D426.

    Article  CAS  PubMed  Google Scholar 

  27. Carbon S, Douglass E, Good BM, Unni DR, Harris NL, Mungall CJ, et al. The Gene Ontology resource: enriching a GOld mine. Nucleic Acids Res. 2021;49:D325–D334.

    Article  CAS  Google Scholar 

  28. Jin Y, Tam OH, Paniagua E, Hammell M. TEtranscripts: a package for including transposable elements in differential expression analysis of RNA-seq datasets. Bioinformatics. 2015;31:3593–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Gao Y, Wang J, Zhao F. CIRI: an efficient and unbiased algorithm for de novo circular RNA identification. Genome Biol. 2015;16:4.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Gao Y, Zhang J, Zhao F. Circular RNA identification based on multiple seed matching. Brief Bioinform. 2018;19:803–10.

    Article  CAS  PubMed  Google Scholar 

  31. Mo C-F, Wu F-C, Tai K-Y, Chang W-C, Chang K-W, Kuo H-C, et al. Loss of non-coding RNA expression from the DLK1-DIO3 imprinted locus correlates with reduced neural differentiation potential in human embryonic stem cell lines. Stem Cell Res Ther. 2015;6:1.

    Article  PubMed  PubMed Central  Google Scholar 

  32. Kopp F, Mendell JT. Functional classification and experimental dissection of long noncoding RNAs. Cell. 2018;172:393–407.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Sakashita A, Maezawa S, Takahashi K, Alavattam KG, Yukawa M, Hu Y, et al. Endogenous retroviruses drive species-specific germline transcriptomes in mammals. Nat Struct Mol Biol. 2020;27:967–77.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Hoffmann LB, McVicar EA, Harris RV, Collar-Fernández C, Clark MB, Hannan AJ, et al. Increased paternal corticosterone exposure influences offspring behaviour and expression of urinary pheromones. BMC Biol. 2023;21:186.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Thivisol, UMCC, Ho, P, Li, B, Trompke, M, Hoffmann, LB, Hannan, AJ, et al. Paternal early life stress exerts intergenerational effects on male C57Bl/6J offspring risk-taking behaviors and predator scent-induced c-Fos expression. Neuronal Signal. 2023;7:NS20220097.

  36. Deng S, Zhang J, Su J, Zuo Z, Zeng L, Liu K, et al. RNA m6A regulates transcription via DNA demethylation and chromatin accessibility. Nat Genet. 2022;54:1427–37.

    Article  CAS  PubMed  Google Scholar 

  37. Gokool A, Loy CT, Halliday GM, Voineagu I. Circular RNAs: the brain transcriptome comes full circle. Trends Neurosci. 2020;43:752–66.

    Article  CAS  PubMed  Google Scholar 

  38. Xu C, Zhang J. Mammalian circular RNAs result largely from splicing errors. Cell Rep. 2021;36:109439.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Kapusta A, Kronenberg Z, Lynch VJ, Zhuo X, Ramsay L, Bourque G, et al. Transposable elements are major contributors to the origin, diversification, and regulation of vertebrate long noncoding RNAs. PLoS Genet. 2013;9:e1003470.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Bourque G. Transposable elements in gene regulation and in the evolution of vertebrate genomes. Curr Opin Genet Dev. 2009;19:607–12.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank Brett Purcell and Travis Featherby for their assistance with behavioural data collection, Craig Thomson and Maria Bastias for their support with animal housing, Fiona Waters for performing the microinjections, the staff at the WEHI animal facility, and the Spartan High Performance Computing resource offered by the University of Melbourne. LBH is supported by the Melbourne Research Scholarship. AJH, TWB and TYP were supported by NHMRC Project Grant funding. AJH also received support from a NHMRC Principal Research Fellowship and a DHB Foundation (Equity Trustees) Grant. The Florey Institute of Neuroscience and Mental Health acknowledges the support from the Victorian Government’s Operational Infrastructure Support Grant.

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Contributions

LBH contributed to planning experiments, conducted all experiments and analyses, and wrote the manuscript. BL assisted in conducting animal behavioural studies. QZ and WW performed the CaptureSeq gene expression analysis. LJL conducted the capture sequencing. TWB supervised the CaptureSeq and gave critical feedback to its analysis. TYP envisioned the study, supervised study design, data collection and statistical analyses, and reviewed the manuscript. AJH envisioned and funded the study, contributed to planning experiments, provided critical feedback throughout the experiments, and reviewed and edited the manuscript.

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Correspondence to A. J. Hannan.

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Hoffmann, L.B., Li, B., Zhao, Q. et al. Chronically high stress hormone levels dysregulate sperm long noncoding RNAs and their embryonic microinjection alters development and affective behaviours. Mol Psychiatry 29, 590–601 (2024). https://doi.org/10.1038/s41380-023-02350-2

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