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An evolutionarily conserved ribosome-rescue pathway maintains epidermal homeostasis

Nature volume 556, pages 376–380 (2018)Cite this article

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Abstract

Ribosome-associated mRNA quality control mechanisms ensure the fidelity of protein translation1,2. Although these mechanisms have been extensively studied in yeast, little is known about their role in mammalian tissues, despite emerging evidence that stem cell fate is controlled by translational mechanisms3,4. One evolutionarily conserved component of the quality control machinery, Dom34 (in higher eukaryotes known as Pelota (Pelo)), rescues stalled ribosomes5. Here we show that Pelo is required for mammalian epidermal homeostasis. Conditional deletion of Pelo in mouse epidermal stem cells that express Lrig1 results in hyperproliferation and abnormal differentiation of these cells. By contrast, deletion of Pelo in Lgr5-expressing stem cells has no effect and deletion in Lgr6-expressing stem cells induces only a mild phenotype. Loss of Pelo results in accumulation of short ribosome footprints and global upregulation of translation, rather than affecting the expression of specific genes. Translational inhibition by rapamycin-mediated downregulation of mTOR (mechanistic target of rapamycin kinase) rescues the epidermal phenotype. Our study reveals that the ribosome-rescue machinery is important for mammalian tissue homeostasis and that it has specific effects on different stem cell populations.

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Fig. 1: Differential effects of Pelo deletion.
Fig. 2: Lrig1+ stem cells account for Pelo mutant epidermal phenotype.
Fig. 3: Accumulation of short ribosome footprints and global translational changes in Pelo knockout epidermis.
Fig. 4: Inhibition of mTOR pathway attenuates Pelo phenotype progression.

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Change history

  • 16 April 2018

    The Source Data files originally published with this article were missing for Figures 1-4 and Extended Data Figures 1, 2, 4-6 and 9. This has now been corrected.

References

  1. Brandman, O. & Hegde, R. S. Ribosome-associated protein quality control. Nat. Struct. Mol. Biol. 23, 7–15 (2016).

    Article  CAS  Google Scholar 

  2. Shoemaker, C. J. & Green, R. Translation drives mRNA quality control. Nat. Struct. Mol. Biol. 19, 594–601 (2012).

    Article  CAS  Google Scholar 

  3. Blanco, S. et al. Stem cell function and stress response are controlled by protein synthesis. Nature 534, 335–340 (2016).

    Article  ADS  CAS  Google Scholar 

  4. Signer, R. A., Magee, J. A., Salic, A. & Morrison, S. J. Haematopoietic stem cells require a highly regulated protein synthesis rate. Nature 509, 49–54 (2014).

    Article  ADS  CAS  Google Scholar 

  5. Guydosh, N. R. & Green, R. Dom34 rescues ribosomes in 3′ untranslated regions. Cell 156, 950–962 (2014).

    Article  CAS  Google Scholar 

  6. Rezza, A. et al. Signaling networks among stem cell precursors, transit-amplifying progenitors, and their niche in developing hair follicles. Cell Rep. 14, 3001–3018 (2016).

    Article  CAS  Google Scholar 

  7. Becker, T. et al. Structure of the no-go mRNA decay complex Dom34–Hbs1 bound to a stalled 80S ribosome. Nat. Struct. Mol. Biol. 18, 715–720 (2011).

    Article  CAS  Google Scholar 

  8. Ishimura, R. et al. Ribosome stalling induced by mutation of a CNS-specific tRNA causes neurodegeneration. Science 345, 455–459 (2014).

    Article  ADS  CAS  Google Scholar 

  9. Kretzschmar, K., Weber, C., Driskell, R. R., Calonje, E. & Watt, F. M. Compartmentalized epidermal activation of β-catenin differentially affects lineage reprogramming and underlies tumor heterogeneity. Cell Rep. 14, 269–281 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Elkenani, M. et al. Pelota regulates epidermal differentiation by modulating BMP and PI3K/AKT signaling pathways. J. Invest. Dermatol. 136, 1664–1671 (2016).

    Article  CAS  Google Scholar 

  11. Florin, L. et al. Delayed wound healing and epidermal hyperproliferation in mice lacking JunB in the skin. J. Invest. Dermatol. 126, 902–911 (2006).

    Article  CAS  Google Scholar 

  12. Young, D. J., Guydosh, N. R., Zhang, F., Hinnebusch, A. G. & Green, R. Rli1/ABCE1 recycles terminating ribosomes and controls translation reinitiation in 3′UTRs in vivo. Cell 162, 872–884 (2015).

    Article  CAS  Google Scholar 

  13. Mills, E. W., Wangen, J., Green, R. & Ingolia, N. T. Dynamic regulation of a ribosome rescue pathway in erythroid cells and platelets. Cell Rep. 17, 1–10 (2016).

    Article  ADS  CAS  Google Scholar 

  14. Mills, E. W., Green, R. & Ingolia, N. T. Slowed decay of mRNAs enhances platelet specific translation. Blood 129, e38–e48 (2017).

    Article  CAS  Google Scholar 

  15. Ingolia, N. T., Brar, G. A., Rouskin, S., McGeachy, A. M. & Weissman, J. S. The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments. Nat. Protoc. 7, 1534–1550 (2012).

    Article  CAS  Google Scholar 

  16. Ingolia, N. T., Lareau, L. F. & Weissman, J. S. Ribosome profiling of mouse embryonic stem cells reveals the complexity and dynamics of mammalian proteomes. Cell 147, 789–802 (2011).

    Article  CAS  Google Scholar 

  17. Guydosh, N. R., Kimmig, P., Walter, P. & Green, R. Regulated Ire1-dependent mRNA decay requires no-go mRNA degradation to maintain endoplasmic reticulum homeostasis in S. pombe. eLife 6, e29216 (2017).

    Article  Google Scholar 

  18. Guydosh, N. R. & Green, R. Translation of poly(A) tails leads to precise mRNA cleavage. RNA 23, 749–761 (2017).

    Article  CAS  Google Scholar 

  19. Slevin, M. K. et al. Deep sequencing shows multiple oligouridylations are required for 3′ to 5′ degradation of histone mRNAs on polyribosomes. Mol. Cell 53, 1020–1030 (2014).

    Article  CAS  Google Scholar 

  20. Ben-Shem, A., Jenner, L., Yusupova, G. & Yusupov, M. Crystal structure of the eukaryotic ribosome. Science 330, 1203–1209 (2010).

    Article  ADS  CAS  Google Scholar 

  21. Lareau, L. F., Hite, D. H., Hogan, G. J. & Brown, P. O. Distinct stages of the translation elongation cycle revealed by sequencing ribosome-protected mRNA fragments. eLife 3, e01257 (2014).

    Article  Google Scholar 

  22. Thoreen, C. C. et al. A unifying model for mTORC1-mediated regulation of mRNA translation. Nature 485, 109–113 (2012).

    Article  ADS  CAS  Google Scholar 

  23. Kristensen, A. R., Gsponer, J. & Foster, L. J. Protein synthesis rate is the predominant regulator of protein expression during differentiation. Mol. Syst. Biol. 9, 689 (2013).

    Article  Google Scholar 

  24. Laplante, M. & Sabatini, D. M. mTOR signaling in growth control and disease. Cell 149, 274–293 (2012).

    Article  CAS  Google Scholar 

  25. Darnell, J. C. et al. FMRP stalls ribosomal translocation on mRNAs linked to synaptic function and autism. Cell 146, 247–261 (2011).

    Article  CAS  Google Scholar 

  26. Sawicka, K., Pyronneau, A., Chao, M., Bennett, M. V. & Zukin, R. S. Elevated ERK/p90 ribosomal S6 kinase activity underlies audiogenic seizure susceptibility in fragile X mice. Proc. Natl Acad. Sci. USA 113, E6290–E6297 (2016).

    Article  CAS  Google Scholar 

  27. Liu, R. et al. Impairing the production of ribosomal RNA activates mammalian target of rapamycin complex 1 signalling and downstream translation factors. Nucleic Acids Res. 42, 5083–5096 (2014).

    Article  CAS  Google Scholar 

  28. Mills, E. W. & Green, R. Ribosomopathies: There’s strength in numbers. Science 358, eaan2755 (2017).

    Article  Google Scholar 

  29. Gingold, H. et al. A dual program for translation regulation in cellular proliferation and differentiation. Cell 158, 1281–1292 (2014).

    Article  CAS  Google Scholar 

  30. Watt, F. M., Jordan, P. W. & O’Neill, C. H. Cell shape controls terminal differentiation of human epidermal keratinocytes. Proc. Natl Acad. Sci. USA 85, 5576–5580 (1988).

    Article  ADS  CAS  Google Scholar 

  31. Page, M. E., Lombard, P., Ng, F., Göttgens, B. & Jensen, K. B. The epidermis comprises autonomous compartments maintained by distinct stem cell populations. Cell Stem Cell 13, 471–482 (2013).

    Article  CAS  Google Scholar 

  32. Nyamsuren, G. et al. Pelota regulates the development of extraembryonic endoderm through activation of bone morphogenetic protein (BMP) signaling. Stem Cell Res. 13, 61–74 (2014).

    Article  CAS  Google Scholar 

  33. Barker, N. et al. Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature 449, 1003–1007 (2007).

    Article  ADS  CAS  Google Scholar 

  34. Snippert, H. J. et al. Lgr6 marks stem cells in the hair follicle that generate all cell lineages of the skin. Science 327, 1385–1389 (2010).

    Article  ADS  CAS  Google Scholar 

  35. Madisen, L. et al. A robust and high-throughput Cre reporting and characterization system for the whole mouse brain. Nat. Neurosci. 13, 133–140 (2010).

    Article  CAS  Google Scholar 

  36. Šošić, D., Richardson, J. A., Yu, K., Ornitz, D. M. & Olson, E. N. Twist regulates cytokine gene expression through a negative feedback loop that represses NF-κB activity. Cell 112, 169–180 (2003).

    Article  Google Scholar 

  37. Lichtenberger, B. M., Mastrogiannaki, M. & Watt, F. M. Epidermal β-catenin activation remodels the dermis via paracrine signalling to distinct fibroblast lineages. Nat. Commun. 7, 10537 (2016).

    Article  ADS  CAS  Google Scholar 

  38. Skarnes, W. C. et al. A conditional knockout resource for the genome-wide study of mouse gene function. Nature 474, 337–342 (2011).

    Article  CAS  Google Scholar 

  39. Jensen, K. B., Driskell, R. R. & Watt, F. M. Assaying proliferation and differentiation capacity of stem cells using disaggregated adult mouse epidermis. Nat. Protoc. 5, 898–911 (2010).

    Article  CAS  Google Scholar 

  40. Picelli, S. et al. Full-length RNA-seq from single cells using Smart-seq2. Nat. Protoc. 9, 171–181 (2014).

    Article  CAS  Google Scholar 

  41. Afgan, E. et al. The Galaxy platform for accessible, reproducible and collaborative biomedical analyses: 2016 update. Nucleic Acids Res. 44(W1), W3–W10 (2016).

    Article  CAS  Google Scholar 

  42. Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).

    Article  CAS  Google Scholar 

  43. Love, M. I., Anders, S., Kim, V. & Huber, W. RNA-seq workflow: gene-level exploratory analysis and differential expression. F1000 Res. 4, 1070 (2015).

    Article  Google Scholar 

  44. Braun, K. M. et al. Manipulation of stem cell proliferation and lineage commitment: visualisation of label-retaining cells in wholemounts of mouse epidermis. Development 130, 5241–5255 (2003).

    Article  CAS  Google Scholar 

  45. Liakath-Ali, K. et al. Novel skin phenotypes revealed by a genome-wide mouse reverse genetic screen. Nat. Commun. 5, 3540 (2014).

    Article  ADS  Google Scholar 

  46. Lowell, S., Jones, P., Le Roux, I., Dunne, J. & Watt, F. M. Stimulation of human epidermal differentiation by delta-notch signalling at the boundaries of stem-cell clusters. Curr. Biol. 10, 491–500 (2000).

    Article  CAS  Google Scholar 

  47. Gandarillas, A. & Watt, F. M. Changes in expression of members of the fos and jun families and myc network during terminal differentiation of human keratinocytes. Oncogene 11, 1403–1407 (1995).

    CAS  PubMed  Google Scholar 

  48. Mulder, K. W. et al. Diverse epigenetic strategies interact to control epidermal differentiation. Nat. Cell Biol. 14, 753–763 (2012).

    Article  CAS  Google Scholar 

  49. Sen, G. L., Reuter, J. A., Webster, D. E., Zhu, L. & Khavari, P. A. DNMT1 maintains progenitor function in self-renewing somatic tissue. Nature 463, 563–567 (2010).

    Article  ADS  CAS  Google Scholar 

  50. Lattouf, R. et al. Picrosirius red staining: a useful tool to appraise collagen networks in normal and pathological tissues. J. Histochem. Cytochem. 62, 751–758 (2014).

    Article  Google Scholar 

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Acknowledgements

K. L.-A. thanks W. Engel for inspiration. F.M.W. acknowledges funding from the Wellcome Trust, Cancer Research UK and the UK Medical Research Council. We are also grateful for funding from the Department of Health via the National Institute for Health Research comprehensive Biomedical Research Centre award to Guy’s & St Thomas’ National Health Service Foundation Trust in partnership with King’s College London and King’s College Hospital NHS Foundation Trust. R.G. acknowledges funding from Howard Hughes Medical Institute. K.H.S. is the recipient of a Finnish Cultural Foundation Fellowship and T.L. is supported by a Sir Henry Dale Fellowship (206211/Z/17/Z). We thank C. Lelliott and V. Vancollie for Hbs1l knockout samples, and A. Ahmed, G. Donati, R. Driskell, S. Habib, M. Lynch, B. Oulès, D. V. K. Pantakani, I. Sancho-Martinez, G. Walko and all members of the Watt laboratory for discussions. Technical help from M. Battilocchi, C. Li and staff at the Nikon Imaging Centre and the Hodgkin Biological Services Facility, King’s College London, is gratefully  acknowledged. We thank the High-Throughput Genomics Group at the Wellcome Trust Centre for Human Genetics, Oxford, UK (funded by Wellcome Trust: 203141/Z/16/Z) for generation of the sequencing data.

Reviewer information

Nature thanks X. Dai, V. Horsley and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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Author notes

  1. Kifayathullah Liakath-Ali

    Present address: Department of Molecular and Cellular Physiology and Howard Hughes Medical Institute, Stanford University Medical School, Stanford, CA, USA

Authors and Affiliations

  1. Centre for Stem Cells and Regenerative Medicine, King’s College London, London, UK

    Kifayathullah Liakath-Ali, Inês Sequeira, Beate M. Lichtenberger, Angela Oliveira Pisco, Kalle H. Sipilä, Ajay Mishra & Fiona M. Watt

  2. Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins School of Medicine, Baltimore, MD, USA

    Eric W. Mills, Colin Chih-Chien Wu & Rachel Green

  3. Skin and Endothelium Research Division, Department of Dermatology, Medical University of Vienna, Vienna, Austria

    Beate M. Lichtenberger

  4. Cambridge Infinitus Research Centre, University of Cambridge, Cambridge, UK

    Ajay Mishra

  5. Centre for Gene Regulation and Expression, School of Life Sciences, University of Dundee, Dundee, UK

    Harunori Yoshikawa, Tony Ly & Angus I. Lamond

  6. Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, UK

    Tony Ly

  7. Institute of Human Genetics, University Medical Centre of Göttingen, Göttingen, Germany

    Ibrahim M. Adham

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Contributions

K.L.-A. and F.M.W. conceptualized and led the study. K.L.-A., I.S. and B.M.L. performed and analysed mouse experiments. K.L.-A., K.H.S. and A.M. performed and analysed cell culture experiments. A.O.P. analysed data from ribosome profile and RNA-seq experiments. I.M.A. generated the Pelo conditional knockout mouse. E.W.M., C.C.-C.W. and R.G. generated and analysed ribosome-profiling data. H.Y., T.L. and A.I.L. generated and analysed polysome data. K.L.-A. and F.M.W. wrote the manuscript with input from all authors.

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Correspondence to Fiona M. Watt.

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Extended data figures and tables

Extended Data Fig. 1 Pelo is expressed in all skin cell subpopulations and knockout of Hbs1l leads to mild dermal phenotype.

a, b, Pelo and Hbs1l are ubiquitously expressed in all cell populations of embryonic and neonatal skin. mRNA expression data obtained from hair and skin gene expression library (Hair-GEL; http://www.hair-gel.net). c, Schematic of Hbs1l knockout first allele. d, Immunolabelling of tail epidermal wholemounts with antibodies against Krt14, Krt15, Lrig1 and FASN. e, Tail skin sections immunolabelled for Ki67, showing no significant change in the distribution of Ki67+ cells in Hbs1l−/− epidermis. f, H&E staining of adult control and Hbs1l−/− tail skin. g, Herovici’s polychrome staining to visualize immature (blue) and mature (pink) dermal collagen. h, Picrosirius staining of tail skin showing the birefringence of collagen fibres against a black background. i, Immunostaining of tail skin sections with pan-keratin (PanKrt) and vimentin (Vim) antibodies. j–m, Quantification of dermal thickness (j), dermal cell density (k), dermal cellularity (l) and total collagen deposition (m). Dashed lines mark epidermal–dermal boundary. Scale bars, 100 μm. *P = 0.0286 (j, m). n = 12 sections analysed over four mice per group.

Source data

Extended Data Fig. 2 Delayed wound closure in Pelo null epidermis.

a, Histology of skin 10 days post wound (dpw) shows delayed wound closure in PeloepiKO mice. b, c, EdU staining of 10 dpw skin shows reduced proliferation in the wound bed. Itgα6 staining demarcates dermal–epidermal boundary. Box indicates the wound bed. d, Histology of 5 dpw wound shows altered epidermal architecture. e, f, EdU labelling of 5 dpw skin shows reduced proliferation at wound edge. g, h, Immunostaining of Krt14 and Krt10 in 10 and 5 dpw skin shows abnormal differentiation in PeloepiKO mice (arrows). i, TdTomato genetic labelling shows the contribution of Lrig1, Lgr5 and Lgr6 progeny in tail wound healing. Note altered migration of Lrig1 cells in Pelofl/flLrig1CreERT tdTomato mice when compared to Lgr5 and Lgr6 on Pelo deletion. *P = 0.0123 (c), *P = 0.0330 (e); n = 9 sections analysed over 3 mice per group. Scale bars, 100 μm.

Source data

Extended Data Fig. 3 Pelo deletion leads to progressive hair follicle and sebaceous gland abnormalities.

a–c, Confocal images of tail epidermal wholemounts immunostained for Krt14, hair follicle bulge markers CD34 and Krt15, sebocyte maturation marker fatty acid synthase (FASN) and junctional zone stem cell marker Lrig1 show progressive changes in hair follicle and sebaceous gland structure from P16 to P120 in PeloepiKO mice. Note that the FASN staining in P84 and P120 PeloepiKO epidermis is non-specific owing to highly keratinized hair follicles. Asterisks in b indicate non-specific staining of sebaceous glands. Scale bars, 100 μm.

Extended Data Fig. 4 Postnatal epidermal Pelo deletion impairs barrier function and wound healing.

a, b, Breeding scheme and topical 4-OHT treatment regimen. c, Representative Pelofl/flKrt14CreERT mouse showing skin lesions (dashed area) in 4-OHT-treated dorsal skin. d, TEWL is increased in 4-OHT-treated skin of Pelofl/flKrt14CreERT mice. e, Rate of wound closure. f, Tail epidermal wholemounts immunostained with Krt14 and Krt15 antibodies showing altered sebaceous gland architecture (arrows) in 4-OHT-treated Pelofl/flKrt14CreERT mice. g, Tail epidermal wholemounts from TdTomato (red) genetically labelled Pelofl/flKrt14CreERT mice show keratinized cysts in hair follicles (arrows). h, Cumulative mean values of gene expression obtained from ribosome profiling show downregulation of markers of sebaceous gland differentiation and increase in Myc. i, Tail epidermal wholemounts showing altered expression of FASN, Scd1 and Lrig1 (arrows) in sebaceous glands of 4-OHT-treated Pelofl/flKrt14CreERT mice (middle and right). Dashed lines indicate pilosebaceous units. Scale bars, 100 μm. **P = 0.0072, *P = 0.0650, ns, not significant. n = 3 in treated and untreated control groups.

Source data

Extended Data Fig. 5 Knockdown of PELO in human keratinocytes phenocopies mouse epidermal phenotype and proliferation difference in mice lacking Pelo in Lrig1+, Lgr5+ and Lgr6+ stem cells.

a–d, PELO knockdown validation. a, qRT–PCR for individual siRNAs transfected in human primary keratinocytes. b, Clonal growth. c, d, Colony number and average size of individual colonies. e–g, Clonal growth of keratinocytes, comparing pooled PELO siRNA knockdown (PELOsiRNA) and scrambled (Scr) control. h–l, Effect of PELO knockdown in human epidermal reconstitution assay on decellularized dermis. h, i, Epidermal thickness of de-epidermized dermis (DED) cultures is significantly increased on PELO knockdown. j–l, Immunolabelling for Krt14 (K14), Ki67, p63 and differentiation markers Krt10 (K10) and involucrin (IVL) shows increased number of differentiated cell layers (j) and increased number of cells expressing Ki67 and p63 (k, l) in PELOsiRNA reconstituted epidermis. Dashed lines indicated dermal–epidermal boundary. m, Assessing proliferation by Ki67 and p63 labelling in dorsal IFE sections of mice lacking Pelo in Lrig1, Lgr5 and Lgr6 stem cells. Scale bars, 100 μm. ***P = 0.0009 (a, for siRNA#10), ***P = 0.0004 (a, for siRNA#11), **P = 0031 (a, for siRNA#12); *P = 0.0286 (c); *P = 0.0286 (d); **P = 0.0022 (f); **P = 0.0087 (g); ****P ≤ 0.0001 (i); *P = 0.0229 for Ki67 and *P = 0.0107 for p63 (l). n = 2 independent transfections; n = 3 dishes (a–g) and n = 2 sections of reconstituted epidermis (h, l).

Source data

Extended Data Fig. 6 Lrig1+ stem cells account for Pelo mutant epidermal phenotype.

a, Tail epidermal wholemounts labelled with Krt14 and Ki67 antibodies, showing increased proliferation and alterations to the junctional zone (asterisks) and sebaceous glands (arrow) in Pelofl/flLrig1CreERT2 mice. b, Cross-section of dorsal skin stained for EdU shows increased proliferation and alterations in hair follicle infundibulum structure (arrow) in Pelofl/flLrig1CreERT2 mice. c–e, Confocal images of tail epidermal wholemounts (c, e) and dorsal skin sections (d) of tdTomato labelled Pelofl/flLrig1CreERT2, Pelofl/flLgr5CreERT2 and Pelofl/flLgr6CreERT2 mice. c, d, Expansion of tdTomato-labelled Lrig1 (arrows) but not Lgr5 or Lgr6 progeny upon Pelo deletion. e, f, Increase in proliferation (Ki67 labelling) of Lrig1+ (arrows) but not Lgr5+ and Lgr6+ populations. Scale bars, 100 μm. *P = 0.0047 (f). n = 9 wholemounts analysed over 3 mice per group. All mice were in telogen of the hair cycle (2–3 months old) when treated with 4-OHT. Treatment regime and harvest of tissue were as indicated in Fig. 2c. Dashed lines mark epidermal–dermal boundary.

Source data

Extended Data Fig. 7 Pelo knockout epidermal cells do not accumulate 3′ UTR footprints.

a, b, Empiric cumulative distribution plots of relative 3′ UTR ribosome occupancy for all transcripts (a) or those with at least one read mapped to the 3′ UTR (b). c–e, Gene Ontology of genes differentially expressed in Pelo-null epidermis. Functional, component and process categories of genes enriched in PeloepiKO mice.

Extended Data Fig. 8 Computational analysis of differentially regulated pathways between control and PeloepiKO and comparison of molecular signatures in PeloepiKO and Gtpbp2-deficient brain.

a, Number of genes that were differentially expressed in PeloepiKO and control epidermis and their associated functions. b, c, Ingenuity Pathway Analysis showing changes in mTOR pathway genes in PeloepiKO versus control epidermis (b) and their predicted molecular activities (c). d, Venn diagram shows common differentially expressed genes in our study and that of Ishimura et al.8 when comparing control and mutant mice. The 314 overlapping genes are enriched in top canonical pathways that are highly related to translation.

Extended Data Fig. 9 Pelo epidermal deletion results in increased protein synthesis and basal stem cell size and rapamycin treatment reduces proliferation of Pelo-null epidermis.

a, Gating strategy for measurement of OP-P incorporation into cell populations. b, c, Confocal images of tail and ear epidermal wholemounts immunolabelled for Krt14 and P-cadherin (P-Cad), showing IFE basal cells. d, e, Representative flow cytometric dot plot showing increased cell size (FSC-A) of Itgα6high cells and altered S and G2/M cell cycle phases in PeloepiKO epidermis. f, g, Breeding scheme and rapamycin treatment regimen. h, Immunolabelling of tail epidermal wholemounts with Krt14 and Ki67 antibodies shows reduced proliferation in rapamycin-treated mice compared to vehicle-treated group. Note that there was no significant change in epidermal proliferation of control mice treated with rapamycin when compared to vehicle-treated mice. i, j, Cross-sections of IFE from PeloepiKO and control back skin immunolabelled with Krt14 and Ki67 antibodies, showing significant reduction in Ki67+ and suprabasal Krt14+ cells in rapamycin-treated compared to vehicle-treated mice. k, Cross-sections of IFE from control and PeloepiKO back skin immunolabelled with Krt14 and pS6K antibodies showing marked increase in pS6K labelling, indicating mTOR hyperactivation in vehicle-treated PeloepiKO skin. l, Cross-sections of IFE of control and Pelofl/flKrt14CreERT mice (with simultaneous 4-OHT and rapamycin treatment) immunolabelled for Krt14 and EdU showing significant reduction in EdU+ and suprabasal Krt14+ cells in rapamycin-treated compared to vehicle-treated mice. m, Cross-sections of IFE of control and Pelofl/flKrt14CreERT mice (with simultaneous 4-OHT and rapamycin treatment) immunolabelled for Krt14 and p4EBP antibodies. Note reduced pS6K labelling (k) and p4EBP1 (m) in rapamycin-treated epidermis. Greyscale images for pS6K are shown below merged images. Scale bars, 100 μm. *P = 0.0132 (j), n. s., non significant. n = 9 sections analysed over 3 mice per group. Dashed lines mark epidermal–dermal boundary.

Source data

Extended Data Fig. 10 RNA-seq of Lrig1+, Lgr5+ and Lgr6+ cells reveals Lgr5 as a transcriptionally unique subpopulation and subtle changes in transcription in all subpopulations when Pelo is deleted.

a, Schematic illustration of the EGFPhigh sorting and RNA-seq strategy for control and Pelo-deleted subpopulations using Pelofl/flLrig1EGFP-CreERT2, Pelofl/flLgr5EGFP-CreERT2 and Pelofl/flLgr6EGFP-CreERT2 mice. b, Principal component analysis of RNA-seq data shows that the Lgr5 subpopulation is remarkably different from the other two. Note that there is no major change in the clusters when Pelo is deleted in any of the subpopulations. c, Hierarchical clustering of the subpopulations corroborates minimal transcriptional changes between control and Mut mice, revealing two major clusters, one for Lgr5 and another for Lrig1 and Lgr6. d, Venn diagram illustrating the differentially expressed genes in common among the three subpopulations when comparing control and mutant cells. e–g, Top differentially regulated transcription factors between control and Pelofl/flLrig1EGFP-CreERT2 (e), Pelofl/flLgr5EGFP-CreERT2 (f) and Pelofl/flLgr6EGFP-CreERT2(g) subpopulations. h–j, Top differentially regulated canonical pathways between control and Pelofl/flLrig1EGFP-CreERT2 (h), Pelofl/flLgr5EGFP-CreERT2 (i) and Pelofl/flLgr6EGFP-CreERT2 (j) subpopulations. k, Schematic of skin showing location of marker expression and the various transgenic mice used in this study.

Supplementary information

Reporting Summary

Supplementary Table 1

A list of specific keratin genes enriched for short footprints

Supplementary Table 2

GO analysis of transcripts enriched in short ribosomal footprints. Genes enriched for each GO category are presented in separate tabs.

Supplementary Table 3

GO analysis of differentially expressed transcripts. Genes enriched for each GO category are presented in separate tabs.

Supplementary Table 4

A list of differentially expressed genes among Lrig1+, Lgr5+, Lgr6+ control and Pelo-mutant populations, intersections between Lrig1+, Lgr5+, Lgr6+ control and Pelo-mutant populations.

Supplementary Table 5

A list of upstream regulators and canonical pathways activated in among and between control Lrig1+, Lgr5+ and Lgr6+ subpopulations.

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Liakath-Ali, K., Mills, E.W., Sequeira, I. et al. An evolutionarily conserved ribosome-rescue pathway maintains epidermal homeostasis. Nature 556, 376–380 (2018). https://doi.org/10.1038/s41586-018-0032-3

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