Citation: Terzian T, Box N (2013) Genetics of Ribosomal Proteins: “Curiouser and Curiouser”. PLoS Genet 9(1): e1003300. https://doi.org/10.1371/journal.pgen.1003300
Editor: Gregory S. Barsh, Stanford University School of Medicine, United States of America
Published: January 31, 2013
Copyright: © 2013 Terzian, Box. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: TT is supported by the US National Institutes of Health-NIAMS (K01-AR063203, P30-AR057212). NB is funded by NIAMS (P30-AR057212) and NCI (P30-CA046934). The funders had no role in the preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
The Mystery of Minutes
Lewis Carroll wrote about Alice, but he might just as well have been referring to Calvin Bridges. As a student in T.H. Morgan's lab, Bridges described some of the earliest Drosophila mutations, including so-called Minutes, in which heterozygotes exhibited small body size and developmental abnormalities in tissues undergoing rapid cell division, and homozygotes were lethal . At the time, it was curious how dozens of different loci could yield the same phenotype, and even curiouser how flies multiply heterozygous at different Minute loci were no more severely affected than a single Minute mutant. This mystery—how dozens of genes could encode similar but separate proliferative functions in all cells—was solved more than 50 years later with the realization that mutations of ribosomal protein genes occur in almost all Minute loci .
In this issue of PLOS Genetics, Watkins-Chow et al.  add to a more recent curiosity: even though ribosomes (and ribosomal protein [RP] genes) have remained nearly identical across more than a billion years of evolution, mutations of RP genes in mice and in humans give rise to a surprising diversity of phenotypes. This work adds a new piece to a very old puzzle, and suggests the possibility that RP genes do more than just contribute to ribosomes.
RP Mutations in Mammals: More than Minutes
To date, 11 different RP mutant mice have now been reported. These mice carry deletions, missense, or splicing mutations that have arisen spontaneously, from N-Ethyl-N-Nitrosurea (ENU) mutagenesis screens, or from targeted gene deletion – (Table 1). Overall, RP mutant mice exhibit an unexpected array and diversity of phenotypes. For example, the spontaneous mutant Belly spot and tail is caused by a splicing abnormality in Rpl24 ; heterozygous mutants (Rpl24Bst/+) are small with white hind feet, a midline belly spot (Bst), abnormal retinal development, and skeletal abnormalities that include a curly tail . Heterozygosity for a targeted mutation of Rps6 causes embryonic lethality , while heterozygosity for targeted mutations of Rpl22 or of Rpl29 have no effect. (Homozygosity for targeted mutations of Rpl22 and of Rpl29 causes a T-cell–specific developmental defect  and generalized reduced growth , respectively.) Mice heterozygous for mutations in Rps19, Rps20, or Rpl27a exhibit epidermal hyperpigmentation, anemia, and reduced body size while the Rpl27a heterozygotes also exhibit cerebellar ataxia. Rps19-null mice are embryonic lethal prior to implantation . CD74-Nid67+/− mice with a heterozygous deletion of eight genes including Rps14 developed macrocytic anemia and other hematopoietic defects . Finally, heterozygous Rpl38 mutants had abnormalities of skeletal patterning .
Watkins-Chow et al.  present two different missense alleles of Rps7, Montu (Mtu) and Zuma (Zma), that were generated from an ENU mutagenesis screen. Most Rps7Mtu/+ mice die in utero (74% on a C3H/HeJ background), but the survivors show pleiotropic phenotypes including reduced body size, abnormal skeletal morphology, and mid-ventral white spotting. This phenotype cluster is reproduced in Rps7Zma/+ mice and is similar to that in Rpl24 mutant mice . These observations support the existence of distinctive spatial and temporal characteristics for RPs : Rps7, Rps19, Rps20, and Rpl24 may be necessary for melanocyte development , , , , while Rps6, Rps19, Rps20, and Rpl27a are important for keratinocytes , . Additionally, Rps7, Rpl24, and Rpl38 are crucial for skeletal and retinal development , ,  and Rps14, Rps19, Rps20, and Rpl27a are necessary for hematopoeisis , , . RP genotype–phenotype correlations are also found in humans where mutations cause diseases with similarly complex clinical manifestations such as Diamond Blackfan anemia (DBA). DBA is characterized by diverse abnormalities including anemia, congenital craniofacial malformations, and defects in kidney development .
The Role of p53 in RP Haploinsufficiency Phenotypes
While disturbances in ribosome biogenesis have been linked to many human diseases , there is also increasing evidence that RP mutations may be associated with cancer susceptibility –. In keeping with these observations, a sensitive connection between RPs and the tumor suppressor p53 has been identified (reviewed in ). p53 is a major cellular stress sensor that is best known for its ability to induce apoptosis, cell cycle arrest, and senescence in response to a variety of insults including DNA damage, oncogene activation, hypoxia, and more recently, ribosomal stress . Thus, the p53 pathway provides a surveillance mechanism for the preservation of genomic and ribosomal integrity. Perturbation of ribosomal integrity induces the release of several RPs from the nucleolus that interact with and suppress the activity of the main negative regulator of p53, Mdm2, leading to the stabilization and activation of p53 –. This is observed in a number of RP mutant mice that exhibit cell cycle arrest and apoptosis in the affected cell types and in which the resulting pathologies are ameliorated or suppressed by p53 deletion (Table 1). In Rps19Dsk3/+, Rps20Dsk4/+, Rpl24Bst/+, Rpl27aSfa/+, and Rps7Zma/+ mice, the removal of one p53 copy is sufficient to alleviate all phenotypic abnormalities –, . In the case of Rps6+/− mice, fetal mortality is delayed by only a couple of days in the absence of p53 . Amazingly, the embryonic lethality of Rps7Zma/+ mice was completely suppressed by the loss of one p53 allele, and Rps7Zma/+:p53+/− mice are for the most part identical to their wild-type littermates . Taken together, these experiments establish p53 as a true sensor of nucleolar stress and highlight the extraribosomal activity of RPs as modulators of the Mdm2–p53 pathway.
RP Functions in Mutant Mice
Mutant RP phenotypes in mice appear to be the result of three distinct mechanisms: 1) global suppression of protein synthesis; 2) specific suppression of protein synthesis; and 3) extra-ribosomal functions. Diminished global protein synthesis was identified in Rpl24Bst/+ (∼30% reduced) and Rpl29−/− mouse neural tube and somites (∼45% reduced), although no concurrent phenotype was described . In Rps7Mtu/+ mice, a novel defect in 18S rRNA preprocessing was identified in brain and liver, without a reduction in protein synthesis. We speculate that an accompanying reduction in protein synthesis could explain the homozygous embryonic lethality in these mice. On the other hand, the abnormal homeotic transformations observed in Rpl38Ts/+ mice were attributed to the unique role of Rpl38 in translation of specific Hox mRNAs rather than to a global effect . RPL38 appeared to facilitate 80S complex formation during the earliest steps of translation initiation on selective Hox mRNAs. In addition to these global and gene-specific translational defects, extra-ribosomal functions of numerous RPs have been described in prokaryotes and lower eukaryotes such as yeast and flies, and in cultured human cells , , . This expanding list of functions includes cellular apoptosis, transcription and mRNA processing, DNA repair, development, and tumorigenesis. To date, the ability of RPs to activate p53 is the only described extra-ribosomal function in mice. In Rps7Mtu/+ mice, impaired rRNA preprocessing and p53 activation occur simultaneously, illustrating the complex roles of RPs in mammalian tissues. With this in mind, mouse RP phenotypes (heterozygous and homozygous) need to be analyzed on a p53-null as well as on a p53 wild-type genetic background. Since so little is known about how changes in ribosomal protein levels may impact cellular function in vivo, a full repertoire of mouse RP models and improved characterization are sorely needed.
Due to the essential nature of RPs, it is apparent that even small perturbations to their functions can result in an increasing array of diseases, and it is clear now more than ever that the mystery of the underlying mechanistic basis for RP mutant phenotypes needs to be solved before state-of-the art translational approaches can bring effective treatments. Moreover, additional mouse models for RP disorders will provide an important preclinical resource for developing new treatments of ribosomopathies.
We apologize for the omission of many important references due to space restraints. We are grateful to Dr. Greg Barsh for providing valuable input and constructive comments, and Ros Attenborough for helpful suggestions. We also thank Drs. Enrique Torchia and Tyler Vukmer for critical reading of the manuscript.
- 1. Watkins-Chow DE, Cooke J, Pidsley R, Edwards A, Slotkin R, et al. (2013) Mutation of the Diamond-Blackfan anemia gene Rps7 in mouse results in morphological and neuroanatomical phenotypes. PLoS Genet 9: e1003094 .
- 2. Barkic M, Crnomarkovic S, Grabusic K, Bogetic I, Panic L, et al. (2009) The p53 tumor suppressor causes congenital malformations in Rpl24-deficient mice and promotes their survival. Mol Cell Biol 29: 2489–2504.
- 3. Terzian T, Dumble M, Arbab F, Thaller C, Donehower LA, et al. (2011) Rpl27a mutation in the sooty foot ataxia mouse phenocopies high p53 mouse models. J Pathol 224: 540–552.
- 4. Oliver ER, Saunders TL, Tarle SA, Glaser T (2004) Ribosomal protein L24 defect in belly spot and tail (Bst), a mouse Minute. Development 131: 3907–3920.
- 5. Volarevic S, Stewart MJ, Ledermann B, Zilberman F, Terracciano L, et al. (2000) Proliferation, but not growth, blocked by conditional deletion of 40S ribosomal protein S6. Science 288: 2045–2047.
- 6. Anderson SJ, Lauritsen JP, Hartman MG, Foushee AM, Lefebvre JM, et al. (2007) Ablation of ribosomal protein L22 selectively impairs alphabeta T cell development by activation of a p53-dependent checkpoint. Immunity 26: 759–772.
- 7. McGowan KA, Li JZ, Park CY, Beaudry V, Tabor HK, et al. (2008) Ribosomal mutations cause p53-mediated dark skin and pleiotropic effects. Nat Genet 40: 963–970.
- 8. Kirn-Safran CB, Oristian DS, Focht RJ, Parker SG, Vivian JL, et al. (2007) Global growth deficiencies in mice lacking the ribosomal protein HIP/RPL29. Dev Dyn 236: 447–460.
- 9. Kondrashov N, Pusic A, Stumpf CR, Shimizu K, Hsieh AC, et al. (2011) Ribosome-mediated specificity in Hox mRNA translation and vertebrate tissue patterning. Cell 145: 383–397.
- 10. Matsson H, Davey EJ, Draptchinskaia N, Hamaguchi I, Ooka A, et al. (2004) Targeted disruption of the ribosomal protein S19 gene is lethal prior to implantation. Mol Cell Biol 24: 4032–4037.
- 11. Barlow JL, Drynan LF, Hewett DR, Holmes LR, Lorenzo-Abalde S, et al. (2010) A p53-dependent mechanism underlies macrocytic anemia in a mouse model of human 5q- syndrome. Nat Med 16: 59–66.
- 12. Panic L, Tamarut S, Sticker-Jantscheff M, Barkic M, Solter D, et al. (2006) Ribosomal protein S6 gene haploinsufficiency is associated with activation of a p53-dependent checkpoint during gastrulation. Mol Cell Biol 26: 8880–8891.
- 13. Warner JR, McIntosh KB (2009) How common are extraribosomal functions of ribosomal proteins? Mol Cell 34: 3–11.
- 14. Boria I, Garelli E, Gazda HT, Aspesi A, Quarello P, et al. The ribosomal basis of Diamond-Blackfan Anemia: mutation and database update. Hum Mutat 31: 1269–1279.
- 15. Narla A, Ebert BL Ribosomopathies: human disorders of ribosome dysfunction. Blood 115: 3196–3205.
- 16. Montanaro L, Trere D, Derenzini M (2008) Nucleolus, ribosomes, and cancer. Am J Pathol 173: 301–310.
- 17. Rao S, Lee SY, Gutierrez A, Perrigoue J, Thapa RJ, et al. Inactivation of ribosomal protein L22 promotes transformation by induction of the stemness factor, Lin28B. Blood 120: 3764–3773.
- 18. Chakraborty A, Uechi T, Kenmochi N Guarding the ‘translation apparatus’: defective ribosome biogenesis and the p53 signaling pathway. Wiley Interdiscip Rev RNA 2: 507–522.
- 19. Kruse JP, Gu W (2009) Modes of p53 regulation. Cell 137: 609–622.
- 20. Dai MS, Zeng SX, Jin Y, Sun XX, David L, et al. (2004) Ribosomal protein L23 activates p53 by inhibiting MDM2 function in response to ribosomal perturbation but not to translation inhibition. Mol Cell Biol 24: 7654–7668.
- 21. Dai MS, Lu H (2004) Inhibition of MDM2-mediated p53 ubiquitination and degradation by ribosomal protein L5. J Biol Chem 279: 44475–44482.
- 22. Lohrum MA, Ludwig RL, Kubbutat MH, Hanlon M, Vousden KH (2003) Regulation of HDM2 activity by the ribosomal protein L11. Cancer Cell 3: 577–587.
- 23. Zhang Y, Wolf GW, Bhat K, Jin A, Allio T, et al. (2003) Ribosomal protein L11 negatively regulates oncoprotein MDM2 and mediates a p53-dependent ribosomal-stress checkpoint pathway. Mol Cell Biol 23: 8902–8912.
- 24. Chen D, Zhang Z, Li M, Wang W, Li Y, et al. (2007) Ribosomal protein S7 as a novel modulator of p53-MDM2 interaction: binding to MDM2, stabilization of p53 protein, and activation of p53 function. Oncogene 26: 5029–5037.
- 25. Zhu Y, Poyurovsky MV, Li Y, Biderman L, Stahl J, et al. (2009) Ribosomal protein S7 is both a regulator and a substrate of MDM2. Mol Cell 35: 316–326.
- 26. Wool IG (1996) Extraribosomal functions of ribosomal proteins. Trends Biochem Sci 21: 164–165.
- 27. Lindstrom MS (2009) Emerging functions of ribosomal proteins in gene-specific transcription and translation. Biochem Biophys Res Commun 379: 167–170.
- 28. Fitch KR, McGowan KA, van Raamsdonk CD, Fuchs H, Lee D, et al. (2003) Genetics of dark skin in mice. Genes Dev 17: 214–228.