Perspectives

Ribosomopathies and the paradox of cellular hypo- to hyperproliferationKim De Keersmaecker,1,2 Sergey O. Sulima,1,2 and Jonathan D. Dinman3

1KU Leuven Department of Oncology, Leuven, Belgium; 2VIB Center for the Biology of Disease, Leuven, Belgium; and 3Department of Cell Biology and

Molecular Genetics, University of Maryland, College Park, MD

Ribosomopathies are largely congenital

diseases linked to defects in ribosomal

proteins or biogenesis factors. Some of

these disorders are characterized by hypo-

proliferative phenotypes such as bonemar-

row failure and anemia early in life, followed

by elevated cancer risks later in life. This

transition fromhypo- to hyperproliferation

presents an intriguingparadox in the field of

hematology knownas “Dameshek’s riddle.”

Recent cancer sequencing studies also

revealed somatically acquired mutations

and deletions in ribosomal proteins in

T-cell acute lymphoblastic leukemia and

solid tumors, further extending the list

of ribosomopathies and strengthening

the association between ribosomal defects

and oncogenesis. In this perspective, we

summarizeandcommenton recent findings

in the field of ribosomopathies. We explain

how ribosomopathies may provide clues to

help explain Dameshek’s paradox and high-

light some of the open questions and chal-

lenges in the field. (Blood. 2015;125(9):

1377-1382)

Introduction

Ribosomopathies are a collection of disorders of ribosome dysfunc-tion characterized by a defect in ribosome biogenesis, typically dueto a mutation in a ribosomal protein or biogenesis factor. They arerare diseases, with the most frequent having incidences in the rangeof 1 in 50 000 to 1 in 200 000 live births. These include Diamond-Blackfan Anemia (DBA) and 5q- syndrome that are associatedwithmutations in ribosomal proteins, as well as diseases linked to defectsin ribosome biogenesis factors, such as Schwachman-Diamondsyndrome and Treacher Collins syndrome. An example of a more rareribosomopathy is X-linked dyskeratosis congenita. All of thesediseases are characterized by distinct clinical phenotypes and patientspresent with one or several of a wide variety of developmental ab-normalities including hematopoietic defects (eg, anemia and othercytopenias), craniofacial malformations, short stature, and mentaland motor retardation. Although diverse, these phenotypes can allbe categorized as cellular hypoproliferative defects. Intriguingly,some ribosomopathy patients are also at increased risk of devel-oping leukemias and solid tumors—hyperproliferative cellulardefects.1-3 For example, DBA is characterized by hypoproliferationphenotypes including macrocytic anemia, short stature, craniofacialdefects, and thumb abnormalities beginning early in life. In addition,these patients have been reported to have a fivefold higher incidenceof cancers, with the highest predisposition to colon cancer (36-foldhigher incidence than in the general population), osteosarcoma(33-fold higher), and acute myeloid leukemia (28-fold higher).1

Although these associations with elevated cancer risks have beendescribed inDBA, it is important to note that this is a rare disease andthat the number of cancer cases studied remains limited. Furtherstudies demonstrating a causal link between ribosomal defects andelevated cancer risks inDBAare needed.Additionally, it is importantto note that elevated cancer risks have not been demonstrated in allribosomopathies (eg, Treacher Collins syndrome has not been linkedto any higher cancer predisposition).

It is not our aim here to give an extensive overview of ri-bosomopathies or an in depth description of the exact clinical mani-

festations of the different diseases. Therefore, we refer to excellentreviews written by others.4-6 It is important to point out that ri-bosomopathies represent only a subset of congenital bone marrowfailure syndromes: marrow failure and myelodysplasia with associ-ated risk for transformation can also be caused by other classes ofcongenital and noncongenital defects, such as mutations in genesinvolved in DNA repair (mutations in Fanconi anemia pathway),7

telomere regulation,8 and epigenetics and splicing.9 However, theseare not the subjects of this review.

In this review,we summarize and comment on recentfindings in thefield of ribosomopathies. To begin, we highlight the rapid expansion ofthe ribosomopathy field due to the discovery of somatic mutations inribosomal proteins in T-cell acute lymphoblastic leukemia (T-ALL)and other cancer samples. Next, we summarize the state-of-the-art ofthe molecular biology underlying the hypo- and hyperproliferativephenotypes in ribosomopathies and provide a model that may partiallyexplain the transition fromone to theother (Dameshek’s riddle). Finally,we discuss some of the open questions and future perspectives in thefield.

“To be” or “not to be” a ribosomopathy:toward a definition

The birth of the field of ribosomopathies dates back to 1999, whenDraptchinskaia et al described mutations in the ribosomal proteincoding gene RPS19/eS19 (note that the second nomenclature refersto the universal ribosomal protein nomenclature, which was recentlyintroduced to conform to atomic resolution ribosome structures fromall3 domains of life)10 in DBA.11 Since then, the list of ribosomopathieshas continued to grow. One example is the recent discovery ofcongenital mutations in ribosomal protein SA (RPSA/uS2) in patientswith isolated congenital asplenia.12Another important expansion of thelist of ribosomopathies comes from the recent application of unbiased

Submitted October 1, 2014; accepted January 7, 2015. Prepublished online as

Blood First Edition paper, January 9, 2015; DOI 10.1182/blood-2014-10-

569616.

© 2015 by The American Society of Hematology

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next generation sequencing-based mutation screening to cancersamples. Although previously all ribosomopathies (with the exceptionof the 5q- syndrome, linked to somatic loss of the RPS14/uS14 gene)have been associated with congenital genetic defects, genome-widesequencing efforts of cancer samples revealed that some of the genesassociated with such congenital defects in ribosomopathies could alsobe targets of somatic mutation in cancers. For example, we recentlyidentified inactivating mutations in the gene encoding ribosomalprotein L5 (RPL5/uL18) in patients with T-ALL, some of which wereidentical to mutations described in DBA patients.13 Rare somaticmutations in T-ALL patients were also identified in RPL11/uL5,another gene that is affected inDBA.14 Interestingly, somatic defectsin ribosomal proteins that have not been implicated in congenitalribosomopathies have now also been identified in T-ALL, gastric,and ovarian cancer. These include somatic mutations in the RPL10/uL16 andRPL22/eL22 genes.13,15-17 Consistentwith the observationthat patients with congenital ribosomopathies are predisposed todevelop various tumor types, the presence of somatic ribosomedefects is emerging as a common occurrence in cancers.18 In the nearfuture, the completion of ongoing cancer sequencing projects willdeepen our understanding of the spectrum of ribosomal mutationsin cancer and may also help identify ribosome biogenesis factorsimplicated in carcinogenesis.

The discovery of somatic ribosome defects in cancer samples callsinto question the current definition of “ribosomopathy.” Should thedefinition be limited to diseases and syndromes involving congenitalmutations in ribosomal protein or biogenesis factor genes only, orshould it be broadened to include somatically acquired mutations aswell? Precedent for the latter definitionmay be found in 5q- syndrome.The observations that not all diseases typically accepted as ribosomo-pathies are associated with elevated cancer risks and that clinicalhypoproliferative phenotypes have not been noted in cancers withsomatic ribosome mutations suggests that the “ribosomopathies”should be broadly defined. However, to err on the side of caution, wedo think that a causal link between the ribosome defect and associateddisease should have been demonstrated before calling a disease aribosomopathy. Therefore, we propose a conservative definitionof ribosomopathies as “any disease associatedwith amutation in aribosomal protein or biogenesis factor impairing ribosome biogenesisin which a defect in ribosome biogenesis or function can be clearlylinked to disease causality.” It is not clear at this point if a disease suchas cartilage hair hypoplasia falls under this definition. So far, therehas been no clear demonstration of a role for RNA component ofmitochondrial RNA processing endoribonuclease (RMRP) (the geneaffected in cartilage hair hypoplasia) in ribosomebiogenesis in humans.Moreover, it is currently unclear whether ribosome dysfunction is thecellular basis for the disorder. This definition also leaves ribosomemutant cancers in a gray zone; although the loss of an RPL22/eL22allele is able to accelerate tumor formation in amousemodel of T-cellleukemia,15 the causal role of other somatic ribosome mutations de-scribed in cancer remains to be shown.

It is worth noting that somatic mutations in ribosomal proteins andbiogenesis factors have only been described in a limited set of cancersamples to date. However, several cancer types are characterized byrecurrent large deletions in which no tumor suppressor gene has yetbeen identified and which contain a ribosomal protein gene that isconsidered intrinsically “uninteresting.” Our recent findings suggestthat defects in ribosomal proteins and other synthesis factors should notbe overlooked in cancer genotypes. Additionally, it has been clearlyestablished by many research groups that protein synthesis appearsderegulated in the majority, if not all, cancer samples. This is notsurprising as this process plays such a central role in the cell that the

signaling cascades and transcriptional regulators that are prominentlyaltered in cancer also (de)regulate translation. Indeed, aberrationsaffecting the PI3K/AKT/mTORpathway, the tumor suppressor proteinp53 (TP53) pathway, and theMYC transcription factor in cancer alsoimpact cellular protein translation by deregulating expression ofcomponents of the translation machinery and by influencing the setofmessenger RNAs (mRNA)which aremost efficiently translated inthe cell.19,20 However, whether altered translation in these instancesis the cause of or consequence of cellular transformation is not clear.Therefore, we think it is not appropriate at this point to label suchtumors ribosomopathies.

How ribosomopathies can offer new insightsinto Dameshek’s riddle

In a 1967 editorial, the founding editor-in-chief of Blood, WilliamDameshek, posed the following riddle: “What do aplastic anemia,paroxysmal nocturnal hemoglobinuria (PNH), and hypoplastic leuke-mia have in common?”21 Here, Dameshek described the intriguingparadox in which patients who initially develop a hypoproliferativedisease, such as aplastic anemia, tend to be at higher risk of hy-perproliferative diseases, such as PNH (clonal expansion of red cells)or acute leukemia (clonal expansion of white cells) later in life.Although not explicitly described by Dameshek, this observation ofa class of diseases characterized in the early stages by a hypo-proliferative disorder, followed by the propensity to develop into ahyperproliferative disease, fits the clinical presentation of certainribosomopathies. As such, Dameshek’s riddlemay bemore broadlyapplicable than initially postulated.

The challenge is to explain this paradox. Dameshek proposed thata sufficiently severe insult to the marrow, whether chemical, ionizingradiation, viral, or by a congenital defect,may result in a variable degreeof injury with a variable degree of hypoplasia. In some cases, abnormalclones of abnormal leukocytes (hypoplastic leukemia) or red cells(PNH) could conceivably arise during the process of repair.21 In otherwords, Dameshek proposed that an initial insult first results inhypoproliferative disease, and that something involved in the processof its repair provides a subset of cells with the capacity to proliferateexcessively. This idea was conceived in 1967, before the moleculargenetics era. Remarkably, his model remains consistent with themodels that are currently being proposed to explainDameshek’s riddle.The differences lie in the depth of our understanding of the molecularnature of the “initial insult,” the identities of the signaling cascades thatlead to impaired proliferation, and the nature of the molecular “repairprocesses” that may underlie the transition to hyperproliferation.

As discussed above, the initial insult occurring in ribosomopathiesare defects in ribosomal protein genes and biogenesis factors.Ribosome biogenesis is a complex multistage process.22 Given thecentral role of the ribosome in the genetic program, there is strongpressure in cells to identify and remove malfunctioning ribosomesbefore they are released into the cytoplasmic pool of actively translatingribosomes. It is becoming clear that this process involves quality controlmechanisms that operate at each step of the process.23,24 This has 2consequences. The first is that cells may not produce enough functionalribosomes to support their metabolic demands. This is particularlygermane to hematopoietic cells, whose normally high proliferative ratesrequire large amounts of ribosomes. This alone may account forcellular hypoproliferation. In addition, too few functional ribosomesdueto insufficient quantities of one or more ribosomal proteins may alsoresult in the release of excess free ribosomal proteins into the cytoplasm.

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This is amolecular stress signal. Themost well-documented research inthis area focuseson cytoplasmic releaseofRPL5/uL18andRPL11/uL5.These have been shown to bind to and inhibit the murine doubleminute 2 protein (MDM2, or HDM2 in humans), an ubiquitin ligasethat directs degradation of the TP53 protein. HDM2 inhibitionresults in TP53 stabilization, initiating a series of cellular signal-ing cascades leading to cell-cycle arrest and apoptosis, adding tocellular hypoproliferation.25-30 This research only represents the tipof the iceberg: TP53 independent cell-cycle arrest and apoptosismechanisms have been described that may also contribute to theribosomopathic-stress-induced hypoproliferation.31-34

Although the human ribosome consists of 81 ribosomal proteins,only a fraction of these have been described as mutated or deleted inribosomopathies. Similarly, only a restricted set of biogenesis factorshave been implicated in ribosomopathies to date.4,5 This is most likelyexplained by the fact that mutation or deletion of many ribosomalproteins and biogenesis factors would be incompatible with cellsurvival, and affected cells would thus not live long enough toproduce the ribosomopathic phenotype. Indeed, the ribosomalproteins currently implicated in ribosomopathies encode proteinslocated on the cytoplasmic/solvent-accessible surfaces of theribosome.35 None of these proteins stabilize interdomain ribosomalRNA (rRNA) interactions nor are they located in regions of theribosome that directly mediate mRNA decoding or peptidyl transfer,leaving ribosomopathic ribosomes functional enough to supportviability. Two typical trans-acting ribosomopathogenic factors,Shwachman–Bodian–Diamond syndrome protein (SBDS) anddyskerin (DKC1) are also consistent with this idea: SBDS is in-volved in the late, cytoplasmic quality control of pre-60S subunits,24

and DKC1 catalyzes pseudouridylation of rRNA. Although rRNAmodifications occur early during rRNA transcription, these are thoughtto merely “fine-tune” ribosome structure rather than playing centralroles in domain folding.36-38

The next issue concerns the nature of the “repair process” inDameshek’s model. Not surprisingly, this is less well understoodbecause it is more complex. To approach this, we have turned tomolecular evolutionary theory. As described above, the originalmutation results in too fewcells available to perform their physiologicalfunction, eg, the production of red blood cells. From an evolutionarystandpoint, this places pressure on the dwindling population ofhematopoietic stem cells, selecting for mutations that enable cells toproduce enough ribosomes to meet their protein synthetic require-ments. One such class of mutants is those that repair the original defect(revertants). A second class, referred to as second site suppressors, ismutations that either inactivate or bypass the ribosome biogenesisquality control apparatus. Cells harboring this class ofmutationswouldproduce enough ribosomes to satisfy their translational requirements,enabling them to out-compete those having only the original mutation.Although this solves the hypoproliferation problem, the tradeoff is thatthese cells must use defective ribosomes. Emerging research indicatesthat these ribosomes have specific defects in translational fidelity(ie, their ability to faithfully translate the genetic code). Indeed,our laboratories have begun to identify a causal chain of eventsdetailing how ribosomopathy associated defects, such as RPL10/uL16R98S or Cbf5p-D95A (a catalytically impaired mutant of Cbf5p, theyeast homolog of DKC1) cause subtle changes in ribosome structurethat alter the basic biochemistry of mutant ribosomes, and how this inturn, affects specific aspects of translational fidelity, altering theexpression of specific sets of genes.35,38 For RPL10/uL16 R98S,we also showed that these changes in gene expression confer newstresses on this population of cells, hastening the selection for ad-ditional compensatory mutations that eventually result in cellular

immortalization (ie, hyperproliferation).35 This is a classic “deal withthe devil,” a process that initially enables cells to fulfill theirphysiological function but eventually leads to their uncontrolled growthand cancer.

The current challenge is to identify the critical second sitesuppressors. Once identified, rational strategies can be designed toreverse their effects. The ideal samples would be tumors collectedfrom patients who developed a ribosome defect-associatedmalignancy.Such patient samples are however scarce, because fortunately, not allpatients develop malignancies. Moreover, cancer samples are oftencharacterized by genetic instability and accumulation of defects,making it difficult to pinpoint the exact nature of the originalhypoproliferation suppressor mutation. This is where the moleculargenetics of model organisms becomes a powerful tool. Recent workfrom our laboratories provides insight into the molecular nature ofthe “repair process.” This began with the identification of a recurrentarginine to serine mutation (R98S) in ribosomal protein RPL10/uL16in T-ALL patient cells. Intriguingly, while T-ALL represents a hyper-proliferative disease, the introduction of this mutation into yeast andmammalian cells impairs ribosome biogenesis and cellular prolif-eration, causing the hypoproliferative phenotype.13 In-depth analysis ofthe RPL10/uL16 R98S mutant using the yeast genetic model revealedthat thismutation does not completely inhibit ribosomebiogenesis; thisis why they are alive, albeit barely, and importantly, allowing cells togrow over many generations selected for a population of cells withnormal growth rates. Genetic analysis revealed that this was due to theacquisition of a compensatorymutation in a ribosome biogenesis factorthat bypassed a critical quality control checkpoint. Importantly, whilethis second sitemutation rescued the hypoproliferative defect, it did notcorrect the underlying structural, biochemical, and translational fidelitydefects, and altered gene expression profiles conferred by the originalRPL10/uL16 R98Smutation (Figure 1A-B).35 Although we have notbeen able to identify analogous mutations in biogenesis factors inRPL10/uL16 R98S positive T-ALL samples, this may be due to thepaucity of knowledge pertaining to human biogenesis factors (ie,mammalian ribosome biogenesis factors are still poorly defined).Additionally, yeast and human cells are sufficiently different so thatmutations in alternative classes of genes may be required to rescuehuman cell proliferation defects. For example, human cells may mu-tate components of the TP53 pathway or of the other pathways thatinitiate hypoproliferation upon ribosome biogenesis-induced stress(Figure 1C). It would be highly interesting to perform polysomeprofiling on noncancerous vs cancerous tissue from patients witha congenital ribosome defect to determine whether a compensatorymutation was acquired that corrects the biogenesis defect (Figure 1B)or whether the biogenesis defect is still present, as predicted by themodel shown in Figure 1C.

Zebrafish are also becoming an attractive model to study ribosomemutation-associated cancers. Seventeen zebrafish lines have beendescribed harboring heterozygous inactivation of different ribosomalprotein genes that are predisposed to develop malignancies. Thesetumors have much in common with those observed in Tp53 nullzebrafish, and tumors inducedby ribosomal proteinhaploinsufficiencyshowed a specific translation defect impairing Tp53 protein pro-duction.39 Unfortunately, a hypoproliferative phenotype has not beendescribed in these fish, and therefore it remains unclear whether or notdefective Tp53 translation can actually cause a switch from a hypo- toa hyperproliferative phenotype in this model system. A mouse modelon RPL22/eL22 haploinsufficiency supports the idea that a ribosomalprotein defect may contribute to a hyperproliferative phenotypeprovided that additionalmutations are present.RPL22/eL22 has beendescribed as a target for heterozygous somaticmutations and deletions

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in T-ALL, and in microsatellite instability-positive gastric andendometrial cancer.15,17,40 Although Rpl22/eL221/2 mice do notdevelop any malignancies within 25 weeks, crossing of these miceto a strain with T-cell–specific expression of myristoylated Akt2(MyrAkt2) accelerates the latency of T-cell lymphoma developmentfrom 19weeks (MyrAkt22Rpl22/eL221/1) to 11weeks (MyrAkt22

Rpl22/eL221/2).15 Although very interesting, these data still do notidentify the nature of the extra mutations that human cancer cellsacquire to be transformed in the context of a ribosomal defect. It isalso worth noting that although the loss of 1 copy of Rpl22/eL22 issufficient to accelerate T-cell lymphoma development in aMyrAkt2background, the loss of both Rpl22/eL22 copies are needed to

Figure 1. Model explaining transition from hypo- to

hyperproliferation phenotypes in ribosomopathies.

(A) In the initial phase of the disease, a mutation in a

ribosomal protein or ribosome biogenesis factor (sym-

bolized by the red star on the 60S subunit) impairs

proper ribosome biogenesis, resulting in lower concen-

trations of assembled ribosomes. This ribosome bio-

genesis defect impairs proper proliferation of the cells by

activating the TP53 pathway and/or by TP53 indepen-

dent mechanisms. At this stage, functional ribosomes

are still assembled to a certain extent. These ribosomes

are however intrinsically defective, and may induce

translational changes/shifts in the cells. (B-C) The

impaired proliferation imposes strong pressure on cell

populations, selecting for cells that acquire a compen-

satory mutation rescuing the impaired proliferation.

The nature of this compensatory mutation is currently

unclear. Cells may acquire a mutation in a biogenesis

factor rescuing the biogenesis defect (B). Alternatively,

the signaling pathways inducing the proliferation im-

pairment upon a biogenesis defect (TP53 or other)

may be crippled by a compensatory mutation (C). In

both scenarios, after acquiring the compensatory mu-

tation, defective ribosomes would still be formed that

alter the translational capacity/fidelity leading to the

cell obtaining a clonal advantage over other cells. How-

ever, these ribosomes are intrinsically defective, leading

to altered gene expression programs. Dashed arrows

indicate the more speculative parts.

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observe a hyperproliferative phenotype of arrested development ofalphabeta-lineage T cells at the b-selection checkpoint.15,41

Future perspectives and conclusions

A long list of genes associated with ribosomopathies has beengenerated over the past 3 decades.However, it is clear that we are stillfar from the goal of a complete understanding of the pathogenesis ofribosomopathies. Essential information regarding the cellular changesassociated with ribosomal mutations remains to be discovered. WhilemRNA expression studies may provide certain clues,42-45 given thatprotein translation is the central function of ribosomes, it may beessential to complement these studies with a detailed analysis of theeffects on protein production. This necessity is illustrated by the recentfinding that in a substantial fraction of DBA cases, one consequenceof the ribosomal protein mutations is the failure of translation of thetranscription factor GATA-1, an essential factor for the developmentof erythropoiesis beyond the erythroid colony-forming unit stage.Thismay be because the long formofGATA-1 is difficult to translate and isimpaired inDBA cells.46 These translational defects of GATA-1werefound because of the rarity of GATA-1 mutations in DBA. However,additional translation defects in DBA and other ribosomopathieshave probably not been discovered because of a lack of thoroughcomparative proteomic studies of normal vs ribosomopathic cells.Unfortunately, this is a classic example of a problem awaiting tech-nological advancement; proteomics technologies are still evolving andanalyzing an entire proteome of human cells is not yet within reachof research laboratories. At this juncture, sequencing of polysome-associated RNA undergoing translation is an option.47-49 The bestapproximation at this point is ribosome profiling technology, whichallows not only the identification of ribosome protected (or translated)mRNA fragments, but also the analysis of alternative start codon usage,read-through beyond stop codons, and of translational kinetics.50,51

Additionally, although the need for cooperating mutations appears tobe clear, questions remain regarding the nature of these mutations, themoment when they are acquired, and the amount required to stimulatehyperproliferation.One attractive possibility accounting for the varyingpenetrance of cancers in different ribosomopathy patients may lie intheir genetic backgrounds (ie, cooperating mutations may be inheritedas allelic variants that are normally maintained in the population).

Yet another intriguing paradox regarding ribosomopathies is theissue of phenotypic tissue specificity: why are such a wide spectrum ofclinical presentations with typical tissue-specific pathologies observedfor each disease, despite the fact that they all share a common defectin the same biochemical process? Although patients with congeni-tal ribosomopathies are bornwith the same ribosome defect in each cellof their body, not all tissues are affected and only particular diseasesymptoms are developed.5 Interestingly, although mutations in severalribosomal proteins and biogenesis factors give rise to anemia indifferent ribosomopathies, each disease also exhibits additional uniquefeatures. Even within the same ribosomopathy, mutations in differentribosomal protein genes promote different types of birth defects. Forinstance, most patients with RPL5/uL18 mutations have a cleft palate,whereas most individuals with RPL11/uL5mutations do not have anycraniofacial defects.52 As such, patients with genetic disorders that arelinked to mutations in ribosomal proteins show remarkably specificphenotypes, suggesting that ribosomal proteins have unique functionsin different tissues. Dissecting the molecular and biochemical defectsof different types of mutant ribosomes can contribute to the excitingemergingfield of “specialized ribosomes,” thosewhose specificitymay

be controlled and fine-tuned by tissue-specific and developmen-tally regulated signals. Alternatively, essential tissue-specific roles ofribosomal proteins outside of the ribosome (extra-ribosomal functions)may explain the tissue-specific phenotypes. Indeed, there is a growingbody of evidence that show ribosomal proteins do not only function intranslation, but that they can also regulate transcription and enzymeactivity in the cell as well.53

In conclusion, many potential explanations for cellular trans-formation exist and are important subjects for investigation. Wespeculate that the ribosomal protein and biogenesis mutants inribosomopathies allow the ribosome to become mature enoughto support some degree of translation, yet still cause sufficientperturbations of ribosomal function to effect the changes in geneexpression associated with ribosomopathic phenotypes. Indeed, weshowed that yeast cells expressing a T-ALL–associated RPL10/uL16R98S mutant or a Cbf5p-D95A mutant promote specific defects intranslation fidelity.35,38 Based on our work on RPL10/uL16 R98S,35

we also hypothesize that compensatory mutations might be likelydrivers of transformation, and are hence particularly important becausethey allow bypass of the cellular proliferation defects due to defectiveribosome biogenesis. One class of compensatory mutations comprisesmutant biogenesis factors, which suppress the assembly but not thefunctional defects of mutant ribosomes. This leads to increased cellularutilization of defective ribosomes with altered fidelity. Additionalexperimentalwork is needed to validate thismodel in other systemsandfor other ribosomal defects. Moreover, while bypassing the ribosomalassembly quality control through mutated biogenesis factors mayprovide one window to oncogenesis, it is likely that other classes ofoncogenic secondary mutations exist. Indeed, it is becoming clear thatcooperating secondary mutations/suppressors are frequently requiredfor disease development.54 Transformation merely represents an endpoint that is likely made more accessible by evolutionary selection forsecondarymutations in a variety of cellular pathways.Determining thenature of the pathways and the mutations is an exciting and promisingfield for investigation.

Acknowledgments

This studywas partially supported by grants from theNational Institutesof Health National Heart, Lung, and Blood Institute (R01 HL119438)(J.D.D.), FondsWetenschappelijk Onderzoek–Vlaanderen (G.0840.13),Stichting tegen Kanker (F25), and the European Research Council (GA334946) (K.D.K).K.D.K. received a postdoctoral fellowship fromFondsWetenschappelijk Onderzoek–Vlaanderen. S.O.S. received a long-termfellowship from the European Molecular Biology Organization.

Authorship

Contribution: K.D.K., S.O.S., and J.D.D. wrote and edited themanuscript.

Conflict-of-interest disclosure: The authors declare no competingfinancial interests.

Correspondence: Kim De Keersmaecker, Campus GasthuisbergO&N4, Box 602, Herestraat 49, 3000 Leuven, Belgium; e-mail:kim.dekeersmaecker@cme.vib-kuleuven.be; and JonathanD. Dinman,Microbiology Building, #231, Department of Cell Biology andMolecular Genetics, University of Maryland, College Park, MD20742; e-mail: dinman@umd.edu.

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References

1. Vlachos A, Rosenberg PS, Atsidaftos E, Alter BP,Lipton JM. Incidence of neoplasia in DiamondBlackfan anemia: a report from the DiamondBlackfan Anemia Registry. Blood. 2012;119(16):3815-3819.

2. Rosenberg PS, Alter BP, Bolyard AA, et al;Severe Chronic Neutropenia InternationalRegistry. The incidence of leukemia and mortalityfrom sepsis in patients with severe congenitalneutropenia receiving long-term G-CSF therapy.Blood. 2006;107(12):4628-4635.

3. Alter BP, Giri N, Savage SA, et al. Malignancies andsurvival patterns in the National Cancer Instituteinherited bone marrow failure syndromes cohortstudy. Br J Haematol. 2010;150(2):179-188.

4. Narla A, Ebert BL. Ribosomopathies: humandisorders of ribosome dysfunction. Blood. 2010;115(16):3196-3205.

5. Armistead J, Triggs-Raine B. Diverse diseasesfrom a ubiquitous process: the ribosomopathyparadox. FEBS Lett. 2014;588(9):1491-1500.

6. Ruggero D, Shimamura A. Marrow failure:a window into ribosome biology. Blood. 2014;124(18):2784-2792.

7. Longerich S, Li J, Xiong Y, Sung P, Kupfer GM.Stress and DNA repair biology of the Fanconianemia pathway. Blood. 2014;124(18):2812-2819.

8. Townsley DM, Dumitriu B, Young NS. Bonemarrow failure and the telomeropathies. Blood.2014;124(18):2775-2783.

9. Bejar R, Steensma DP. Recent developments inmyelodysplastic syndromes. Blood. 2014;124(18):2793-2803.

10. Ban N, Beckmann R, Cate JHD, et al. A newsystem for naming ribosomal proteins. Curr OpinStruct Biol. 2014;24:165-169.

11. Draptchinskaia N, Gustavsson P, Andersson B,et al. The gene encoding ribosomal protein S19 ismutated in Diamond-Blackfan anaemia. NatGenet. 1999;21(2):169-175.

12. Bolze A, Mahlaoui N, Byun M, et al. Ribosomalprotein SA haploinsufficiency in humans withisolated congenital asplenia. Science. 2013;340(6135):976-978.

13. De Keersmaecker K, Atak ZK, Li N, et al. Exomesequencing identifies mutation in CNOT3 andribosomal genes RPL5 and RPL10 in T-cell acutelymphoblastic leukemia. Nat Genet. 2013;45(2):186-190.

14. Tzoneva G, Perez-Garcia A, Carpenter Z, et al.Activating mutations in the NT5C2 nucleotidasegene drive chemotherapy resistance in relapsedALL. Nat Med. 2013;19(3):368-371.

15. Rao S, Lee SY, Gutierrez A, et al. Inactivation ofribosomal protein L22 promotes transformation byinduction of the stemness factor, Lin28B. Blood.2012;120(18):3764-3773.

16. Wang K, Kan J, Yuen ST, et al. Exomesequencing identifies frequent mutation ofARID1A in molecular subtypes of gastric cancer.Nat Genet. 2011;43(12):1219-1223.

17. Novetsky AP, Zighelboim I, Thompson DM Jr,Powell MA, Mutch DG, Goodfellow PJ. Frequentmutations in the RPL22 gene and its clinical andfunctional implications. Gynecol Oncol. 2013;128(3):470-474.

18. Kandoth C, McLellan MD, Vandin F, et al. Mutationallandscape and significance across 12 major cancertypes. Nature. 2013;502(7471):333-339.

19. Ruggero D, Pandolfi PP. Does the ribosome translatecancer? Nat Rev Cancer. 2003;3(3):179-192.

20. Stumpf CR, Ruggero D. The canceroustranslation apparatus. Curr Opin Genet Dev.2011;21(4):474-483.

21. Dameshek W. Riddle: what do aplastic anemia,paroxysmal nocturnal hemoglobinuria (PNH) and“hypoplastic” leukemia have in common? Blood.1967;30(2):251-254.

22. Strunk BS, Karbstein K. Powering throughribosome assembly. RNA. 2009;15(12):2083-2104.

23. Matsuo Y, Granneman S, Thoms M, Manikas RG,Tollervey D, Hurt E. Coupled GTPase andremodelling ATPase activities form a checkpointfor ribosome export. Nature. 2014;505(7481):112-116.

24. Lo KY, Li Z, Bussiere C, Bresson S, Marcotte EM,Johnson AW. Defining the pathway of cytoplasmicmaturation of the 60S ribosomal subunit.Mol Cell.2010;39(2):196-208.

25. Dai MS, Lu H. Inhibition of MDM2-mediated p53ubiquitination and degradation by ribosomalprotein L5. J Biol Chem. 2004;279(43):44475-44482.

26. Marechal V, Elenbaas B, Piette J, Nicolas JC,Levine AJ. The ribosomal L5 protein is associatedwith mdm-2 and mdm-2-p53 complexes. Mol CellBiol. 1994;14(11):7414-7420.

27. Zhang Y, Wolf GW, Bhat K, et al. Ribosomalprotein L11 negatively regulates oncoproteinMDM2 and mediates a p53-dependent ribosomal-stress checkpoint pathway. Mol Cell Biol. 2003;23(23):8902-8912.

28. Lohrum MAE, Ludwig RL, Kubbutat MHG, HanlonM, Vousden KH. Regulation of HDM2 activity bythe ribosomal protein L11. Cancer Cell. 2003;3(6):577-587.

29. Bursac S, Brdovcak MC, Pfannkuchen M, et al.Mutual protection of ribosomal proteins L5 andL11 from degradation is essential for p53activation upon ribosomal biogenesis stress. ProcNatl Acad Sci USA. 2012;109(50):20467-20472.

30. Bursac S, Brdovcak MC, Donati G, Volarevic S.Activation of the tumor suppressor p53 uponimpairment of ribosome biogenesis. BiochimBiophys Acta. 2014;1842(6):817-830.

31. Donati G, Montanaro L, Derenzini M. Ribosomebiogenesis and control of cell proliferation: p53 isnot alone. Cancer Res. 2012;72(7):1602-1607.

32. Narla A, Payne EM, Abayasekara N, et al.L-Leucine improves the anaemia in models ofDiamond Blackfan anaemia and the 5q- syndromein a TP53-independent way. Br J Haematol. 2014;167(4):524-528.

33. Singh SA, Goldberg TA, Henson AL, et al.p53-Independent cell cycle and erythroiddifferentiation defects in murine embryonic stemcells haploinsufficient for Diamond Blackfananemia-proteins: RPS19 versus RPL5. PLoSONE. 2014;9(2):e89098.

34. Heijnen HF, van Wijk R, Pereboom TC, et al.Ribosomal protein mutations induce autophagythrough S6 kinase inhibition of the insulinpathway. PLoS Genet. 2014;10(5):e1004371.

35. Sulima SO, Patchett S, Advani VM, DeKeersmaecker K, Johnson AW, Dinman JD.Bypass of the pre-60S ribosomal quality controlas a pathway to oncogenesis. Proc Natl Acad SciUSA. 2014;111(15):5640-5645.

36. Decatur WA, Fournier MJ. rRNA modificationsand ribosome function. Trends Biochem Sci.2002;27(7):344-351.

37. Baxter-Roshek JL, Petrov AN, Dinman JD.Optimization of ribosome structure and functionby rRNA base modification. PLoS ONE. 2007;2(1):e174.

38. Jack K, Bellodi C, Landry DM, et al. rRNApseudouridylation defects affect ribosomal ligandbinding and translational fidelity from yeast tohuman cells. Mol Cell. 2011;44(4):660-666.

39. MacInnes AW, Amsterdam A, Whittaker CA,Hopkins N, Lees JA. Loss of p53 synthesis inzebrafish tumors with ribosomal protein genemutations. Proc Natl Acad Sci USA. 2008;105(30):10408-10413.

40. Nagarajan N, Bertrand D, Hillmer AM, et al.Whole-genome reconstruction and mutationalsignatures in gastric cancer. Genome Biol. 2012;13(12):R115.

41. Anderson SJ, Lauritsen JPH, Hartman MG, et al.Ablation of ribosomal protein L22 selectivelyimpairs alphabeta T cell development byactivation of a p53-dependent checkpoint.Immunity. 2007;26(6):759-772.

42. Ebert BL, Pretz J, Bosco J, et al. Identificationof RPS14 as a 5q- syndrome gene by RNAinterference screen. Nature. 2008;451(7176):335-339.

43. Aspesi A, Pavesi E, Robotti E, et al. Dissectingthe transcriptional phenotype of ribosomal proteindeficiency: implications for Diamond-Blackfananemia. Gene. 2014;545(2):282-289.

44. Devlin EE, Dacosta L, Mohandas N, Elliott G,Bodine DM. A transgenic mouse modeldemonstrates a dominant negative effect ofa point mutation in the RPS19 gene associatedwith Diamond-Blackfan anemia. Blood. 2010;116(15):2826-2835.

45. Rujkijyanont P, Adams SL, Beyene J, Dror Y.Bone marrow cells from patients withShwachman-Diamond syndrome abnormallyexpress genes involved in ribosome biogenesisand RNA processing. Br J Haematol. 2009;145(6):806-815.

46. Ludwig LS, Gazda HT, Eng JC, et al. Alteredtranslation of GATA1 in Diamond-Blackfananemia. Nat Med. 2014;20(7):748-753.

47. Horos R, Ijspeert H, Pospisilova D, et al.Ribosomal deficiencies in Diamond-Blackfananemia impair translation of transcripts essentialfor differentiation of murine and humanerythroblasts. Blood. 2012;119(1):262-272.

48. Pereboom TC, Bondt A, Pallaki P, et al.Translation of branched-chain aminotransferase-1 transcripts is impaired in cells haploinsufficientfor ribosomal protein genes. Exp Hematol. 2014;42(5):394-403e4.

49. Hesling C, Oliveira CC, Castilho BA, Zanchin NIT.The Shwachman-Bodian-Diamond syndromeassociated protein interacts with HsNip7 and itsdown-regulation affects gene expression at thetranscriptional and translational levels. Exp CellRes. 2007;313(20):4180-4195.

50. Ingolia NT, Brar GA, Rouskin S, McGeachy AM,Weissman JS. The ribosome profiling strategy formonitoring translation in vivo by deep sequencingof ribosome-protected mRNA fragments. NatProtoc. 2012;7(8):1534-1550.

51. Ingolia NT, Lareau LF, Weissman JS. Ribosomeprofiling of mouse embryonic stem cells revealsthe complexity and dynamics of mammalianproteomes. Cell. 2011;147(4):789-802.

52. Gazda HT, Sheen MR, Vlachos A, et al.Ribosomal protein L5 and L11 mutations areassociated with cleft palate and abnormal thumbsin Diamond-Blackfan anemia patients. Am J HumGenet. 2008;83(6):769-780.

53. Lindstrom MS. Emerging functions of ribosomalproteins in gene-specific transcription andtranslation. Biochem Biophys Res Commun.2009;379(2):167-170.

54. Ishimura R, Nagy G, Dotu I, et al. RNA function.Ribosome stalling induced by mutation of aCNS-specific tRNA causes neurodegeneration.Science. 2014;345(6195):455-459.

1382 DE KEERSMAECKER et al BLOOD, 26 FEBRUARY 2015 x VOLUME 125, NUMBER 9

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online January 9, 2015 originally publisheddoi:10.1182/blood-2014-10-569616

2015 125: 1377-1382  

Kim De Keersmaecker, Sergey O. Sulima and Jonathan D. Dinman Ribosomopathies and the paradox of cellular hypo- to hyperproliferation 

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