FRINK Linked Data Fragments server

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2024704480> ?p ?o ?g. }

• W2024704480 endingPage "772" @default.
• W2024704480 startingPage "759" @default.
• W2024704480 abstract "The αβ and γδ T lineages are thought to arise from a common precursor; however, the regulation of separation and development of these lineages is not fully understood. We report here that development of αβ and γδ precursors was differentially affected by elimination of ribosomal protein L22 (Rpl22), which is ubiquitously expressed but not essential for translation. Rpl22 deficiency selectively arrested development of αβ-lineage T cells at the β-selection checkpoint by inducing their death. The death was caused by induction of p53 expression, because p53 deficiency blocked death and restored development of Rpl22-deficient thymocytes. Importantly, Rpl22 deficiency led to selective upregulation of p53 in αβ-lineage thymocytes, at least in part by increasing p53 synthesis. Taken together, these data indicate that Rpl22 deficiency activated a p53-dependent checkpoint that produced a remarkably selective block in αβ T cell development but spared γδ-lineage cells, suggesting that some ribosomal proteins may perform cell-type-specific or stage-specific functions. The αβ and γδ T lineages are thought to arise from a common precursor; however, the regulation of separation and development of these lineages is not fully understood. We report here that development of αβ and γδ precursors was differentially affected by elimination of ribosomal protein L22 (Rpl22), which is ubiquitously expressed but not essential for translation. Rpl22 deficiency selectively arrested development of αβ-lineage T cells at the β-selection checkpoint by inducing their death. The death was caused by induction of p53 expression, because p53 deficiency blocked death and restored development of Rpl22-deficient thymocytes. Importantly, Rpl22 deficiency led to selective upregulation of p53 in αβ-lineage thymocytes, at least in part by increasing p53 synthesis. Taken together, these data indicate that Rpl22 deficiency activated a p53-dependent checkpoint that produced a remarkably selective block in αβ T cell development but spared γδ-lineage cells, suggesting that some ribosomal proteins may perform cell-type-specific or stage-specific functions. T cells mature in the thymus through a well-defined series of stages that can be delineated by changes in expression of the coreceptors CD4 and CD8. The least mature T cell precursors enter the thymus as CD4−CD8− (double-negative or DN) thymocytes, which progress to the CD4+CD8+ (double-positive or DP) stage before selectively silencing one of these coreceptors to become mature CD4+ or CD8+ single-positive (SP) cells. During thymopoiesis, two major types of mature T cells are generated that can be distinguished by the clonotypic subunits contained within their T cell receptor complexes (TCR): αβ T cells and γδ T cells. These two lineages are thought to derive from a common DN precursor (Petrie et al., 1992Petrie H.T. Scollay R. Shortman K. Commitment to the T cell receptor-alpha beta or -gamma delta lineages can occur just prior to the onset of CD4 and CD8 expression among immature thymocytes.Eur. J. Immunol. 1992; 22: 2185-2188Crossref PubMed Scopus (101) Google Scholar) and to separate from one another between the CD44+CD25+ (DN2) and CD44−CD25+ (DN3) stages of development (Ciofani et al., 2006Ciofani M. Knowles G.C. Wiest D.L. von Boehmer H. Zuniga-Pflucker J.C. Stage-specific and differential notch dependency at the alphabeta and gammadelta T lineage bifurcation.Immunity. 2006; 25: 105-116Abstract Full Text Full Text PDF PubMed Scopus (171) Google Scholar). DN thymocytes that productively rearrange the T cell receptor (TCR) γ and TCRδ loci and express a mature γδTCR (TCRγδ heterodimer associated with CD3γɛ and ζ) are capable of differentiating along the γδ-lineage pathway while remaining DN (Kang et al., 1998Kang J. Coles M. Cado D. Raulet D.H. The developmental fate of T cells is critically influenced by TCRgammadelta expression.Immunity. 1998; 8: 427-438Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar, Passoni et al., 1997Passoni L. Hoffman E.S. Kim S. Crompton T. Pao W. Dong M.Q. Owen M.J. Hayday A.C. Intrathymic delta selection events in gammadelta cell development.Immunity. 1997; 7: 83-95Abstract Full Text Full Text PDF PubMed Scopus (100) Google Scholar). In contrast, cells that bear an in-frame TCRβ rearrangement express the pre-TCR (TCRβ-pTα heterodimer associated with CD3γδɛ and ζ) and are able to differentiate along the αβ-lineage pathway to the DP stage. Precursors that have matured to the DP stage in response to pre-TCR signals are said to have been β-selected. β-selection stipulates that only those DN3 thymocytes in which V(D)J recombination produces a functional TCRβ protein will survive and differentiate; those failing to do so die by apoptosis (Dudley et al., 1994Dudley E.C. Petrie H.T. Shah L.M. Owen M.J. Hayday A.C. T cell receptor beta chain gene rearrangement and selection during thymocyte development in adult mice.Immunity. 1994; 1: 83-93Abstract Full Text PDF PubMed Scopus (252) Google Scholar, Hoffman et al., 1996Hoffman E.S. Passoni L. Crompton T. Leu T.M. Schatz D.G. Koff A. Owen M.J. Hayday A.C. Productive T-cell receptor beta-chain gene rearrangement: coincident regulation of cell cycle and clonality during development in vivo.Genes Dev. 1996; 10: 948-962Crossref PubMed Scopus (271) Google Scholar). Pre-TCR signals that induce traversal of the β-selection checkpoint produce four developmental outcomes: (1) rescue of those DN3 thymocytes from apoptosis; (2) extensive proliferative expansion (Hoffman et al., 1996Hoffman E.S. Passoni L. Crompton T. Leu T.M. Schatz D.G. Koff A. Owen M.J. Hayday A.C. Productive T-cell receptor beta-chain gene rearrangement: coincident regulation of cell cycle and clonality during development in vivo.Genes Dev. 1996; 10: 948-962Crossref PubMed Scopus (271) Google Scholar); (3) allelic exclusion at the TCRβ locus, i.e., termination of V(D)J recombination at the remaining β allele (Aifantis et al., 1997Aifantis I. Buer J. von Boehmer H. Azogui O. Essential role of the pre-T cell receptor in allelic exclusion of the T cell receptor beta locus.Immunity. 1997; 7: 601-607Abstract Full Text Full Text PDF PubMed Scopus (117) Google Scholar); and (4) differentiation to the DP stage (Kruisbeek et al., 2000Kruisbeek A.M. Haks M.C. Carleton M. Michie A.M. Zuniga-Pflucker J.C. Wiest D.L. Branching out to gain control: how the pre-TCR is linked to multiple functions.Immunol. Today. 2000; 21: 637-644Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar). Although signaling through the pre-TCR and γδTCR complexes facilitates development of αβ- and γδ-lineage precursors, respectively, the precise role that these receptors play in selection of the αβ and γδ lineages has been controversial (reviewed in Hayes et al., 2003Hayes S.M. Shores E.W. Love P.E. An architectural perspective on signaling by the pre-, alphabeta and gammadelta T cell receptors.Immunol. Rev. 2003; 191: 28-37Crossref PubMed Scopus (60) Google Scholar, MacDonald and Wilson, 1998MacDonald H.R. Wilson A. The role of the T-cell receptor (TCR) in alpha beta/gamma delta lineage commitment: clues from intracellular TCR staining.Immunol. Rev. 1998; 165: 87-94Crossref PubMed Scopus (30) Google Scholar). We and others have recently provided support for the signal strength model of αβ- versus γδ-lineage commitment that posits that strong signals direct development to the γδ lineage whereas relatively weaker signals promote commitment to the αβ lineage (Haks et al., 2005Haks M.C. Lefebvre J.M. Lauritsen J.P. Carleton M. Rhodes M. Miyazaki T. Kappes D.J. Wiest D.L. Attenuation of gammadeltaTCR signaling efficiently diverts thymocytes to the alphabeta lineage.Immunity. 2005; 22: 595-606Abstract Full Text Full Text PDF PubMed Scopus (171) Google Scholar, Hayes et al., 2005Hayes S.M. Li L. Love P.E. TCR signal strength influences alphabeta/gammadelta lineage fate.Immunity. 2005; 22: 583-593Abstract Full Text Full Text PDF PubMed Scopus (192) Google Scholar). In particular, we showed that differential activation of the extracellular signal-regulated kinase (ERK)-early growth response (Egr)-inhibitor of differentiation 3 (Id3) pathway is an important element of the TCR signals that regulate αβ- versus γδ-lineage choice and development (Haks et al., 2005Haks M.C. Lefebvre J.M. Lauritsen J.P. Carleton M. Rhodes M. Miyazaki T. Kappes D.J. Wiest D.L. Attenuation of gammadeltaTCR signaling efficiently diverts thymocytes to the alphabeta lineage.Immunity. 2005; 22: 595-606Abstract Full Text Full Text PDF PubMed Scopus (171) Google Scholar). Nevertheless, the regulatory cascades that control γδ-lineage commitment and development, and how these differ from those involved in adoption of the αβ fate, remain to be established. Rpl22 is a component of the 60S large ribosomal subunit and colocalizes with ribosomal RNA in the nucleolus and the cytoplasm (Lavergne et al., 1987Lavergne J.P. Conquet F. Reboud J.P. Reboud A.M. Role of acidic phosphoproteins in the partial reconstitution of the active 60 S ribosomal subunit.FEBS Lett. 1987; 216: 83-88Crossref PubMed Scopus (39) Google Scholar, Shu-Nu et al., 2000Shu-Nu C. Lin C.H. Lin A. An acidic amino acid cluster regulates the nucleolar localization and ribosome assembly of human ribosomal protein L22.FEBS Lett. 2000; 484: 22-28Abstract Full Text Full Text PDF PubMed Scopus (31) Google Scholar, Toczyski et al., 1994Toczyski D.P. Matera A.G. Ward D.C. Steitz J.A. The Epstein-Barr virus (EBV) small RNA EBER1 binds and relocalizes ribosomal protein L22 in EBV-infected human B lymphocytes.Proc. Natl. Acad. Sci. USA. 1994; 91: 3463-3467Crossref PubMed Scopus (133) Google Scholar). Although ubiquitously expressed and associated with the ribosome, Rpl22 is not required for translation in vitro (Lavergne et al., 1987Lavergne J.P. Conquet F. Reboud J.P. Reboud A.M. Role of acidic phosphoproteins in the partial reconstitution of the active 60 S ribosomal subunit.FEBS Lett. 1987; 216: 83-88Crossref PubMed Scopus (39) Google Scholar). Nevertheless, it remains possible that Rpl22 may play a cell type- or developmental stage-specific role in protein synthesis or ribosome assembly. It has also been suggested that Rpl22 may play a role in assembly or function of other multisubunit ribonucleoprotein (RNP) particles (Dobbelstein and Shenk, 1995Dobbelstein M. Shenk T. In vitro selection of RNA ligands for the ribosomal L22 protein associated with Epstein-Barr virus-expressed RNA by using randomized and cDNA-derived RNA libraries.J. Virol. 1995; 69: 8027-8034PubMed Google Scholar, Le et al., 2000Le S. Sternglanz R. Greider C.W. Identification of two RNA-binding proteins associated with human telomerase RNA.Mol. Biol. Cell. 2000; 11: 999-1010Crossref PubMed Scopus (111) Google Scholar, Toczyski and Steitz, 1991Toczyski D.P. Steitz J.A. EAP, a highly conserved cellular protein associated with Epstein-Barr virus small RNAs (EBERs).EMBO J. 1991; 10: 459-466PubMed Google Scholar, Wood et al., 2001Wood J. Frederickson R.M. Fields S. Patel A.H. Hepatitis C virus 3′X region interacts with human ribosomal proteins.J. Virol. 2001; 75: 1348-1358Crossref PubMed Scopus (68) Google Scholar). Accordingly, Rpl22 has been found to associate with both viral RNAs and proteins in infected cells and to be a component of the telomerase holoenzyme complex (Le et al., 2000Le S. Sternglanz R. Greider C.W. Identification of two RNA-binding proteins associated with human telomerase RNA.Mol. Biol. Cell. 2000; 11: 999-1010Crossref PubMed Scopus (111) Google Scholar, Leopardi et al., 1997Leopardi R. Ward P.L. Ogle W.O. Roizman B. Association of herpes simplex virus regulatory protein ICP22 with transcriptional complexes containing EAP, ICP4, RNA polymerase II, and viral DNA requires posttranslational modification by the U(L)13 proteinkinase.J. Virol. 1997; 71: 1133-1139PubMed Google Scholar, Toczyski and Steitz, 1991Toczyski D.P. Steitz J.A. EAP, a highly conserved cellular protein associated with Epstein-Barr virus small RNAs (EBERs).EMBO J. 1991; 10: 459-466PubMed Google Scholar, Wood et al., 2001Wood J. Frederickson R.M. Fields S. Patel A.H. Hepatitis C virus 3′X region interacts with human ribosomal proteins.J. Virol. 2001; 75: 1348-1358Crossref PubMed Scopus (68) Google Scholar). However, to date Rpl22 has not been implicated in any specific developmental pathway, mechanism, or disease. We report here that despite the ubiquitous expression of Rpl22, Rpl22-deficient (Rpl22−/−) mice exhibited a profound and remarkably selective defect in the development of αβ, but not γδ, T lymphocytes. Rpl22 deficiency arrested development of αβ T cell precursors at the β-selection checkpoint. The developmental arrest was caused by activation of a p53-dependent checkpoint, because p53 deficiency blocked death and rescued thymocyte development. Importantly, the selectivity of the blockade of αβ T cell development resulted from selective induction of p53, as indicated by the fact that p53 amounts were not elevated in Rpl22−/− γδ-lineage cells. The selective increase in p53 expression caused by Rpl22 deficiency was not associated with detectable increases in p53 stability, but was associated with increased p53 biosynthesis. Therefore, despite the germline disruption of the Rpl22 gene, its absence activated a p53-dependent checkpoint only in developing αβ-lineage T cells, suggesting that despite its ubiquitous expression, Rpl22 performs a cell-type-specific or stage-specific function that is particularly important for executing the β-selection differentiation program. Mouse embryonic stem (ES) cells carrying a mutation in the Rpl22 gene were obtained from OmniBank, a library of gene-trapped ES cell clones identified by a corresponding OmniBank Sequence Tag (Zambrowicz et al., 2003Zambrowicz B.P. Abuin A. Ramirez-Solis R. Richter L.J. Piggott J. BeltrandelRio H. Buxton E.C. Edwards J. Finch R.A. Friddle C.J. et al.Wnk1 kinase deficiency lowers blood pressure in mice: a gene-trap screen to identify potential targets for therapeutic intervention.Proc. Natl. Acad. Sci. USA. 2003; 100: 14109-14114Crossref PubMed Scopus (275) Google Scholar). Inverse genomic PCR revealed that the gene-trapping retroviral vector had inserted into chromosome 4 between coding exons 3 and 4 of the Rpl22 gene (Figure S1 in the Supplemental Data available online). ES cells carrying this mutation were used to generate mice heterozygous for the Rpl22 mutation by standard methods (Zambrowicz et al., 2003Zambrowicz B.P. Abuin A. Ramirez-Solis R. Richter L.J. Piggott J. BeltrandelRio H. Buxton E.C. Edwards J. Finch R.A. Friddle C.J. et al.Wnk1 kinase deficiency lowers blood pressure in mice: a gene-trap screen to identify potential targets for therapeutic intervention.Proc. Natl. Acad. Sci. USA. 2003; 100: 14109-14114Crossref PubMed Scopus (275) Google Scholar). Interbreeding of Rpl22 heterozygotes (Rpl22+/−) gave rise to the expected Mendelian ratios of Rpl22+/+, Rpl22+/−, and Rpl22−/− animals (Figure S1; data not shown). To confirm that insertion of the gene-trap vector disrupted Rpl22 expression, mRNA and protein expression was assessed by RT-PCR and immunoblotting. Intact Rpl22 mRNA was not detected in tissues from Rpl22−/− mice (data not shown). As expected from the systemic disruption of the Rpl22 gene, Rpl22 protein was absent from thymus and spleen of Rpl22−/− mice (Figure S1). Remarkably, relative to control littermates (LM), Rpl22−/− mice exhibited no substantial difference in growth rate and size. Likewise, comprehensive clinical diagnostic and pathologic analysis (including tests of inflammation, behavior, obesity, diabetes, bone, cell proliferation, and cardiovascular function; described in detail in Beltrandelrio et al., 2003Beltrandelrio H. Kern F. Lanthorn T. Oravecz T. Piggott J. Powell D. Ramirez-Solis R. Sands A.T. Zambrowicz B.P. Saturation screening of the druggable mammalian genome.in: Carrol P. Fitzgerald K. Model Organisms in Drug Discovery. John Wiley & Sons, Ltd., Chichester, UK2003: 251Crossref Google Scholar) revealed few differences in Rpl22−/− mice relative to those expressing Rpl22, although there was evidence of mild glomerulonephritis in 3 of 3 Rpl22−/− mice examined. Hematologic analysis of Rpl22−/− mice revealed a decrease in peripheral blood leukocytes relative to LM controls, which was caused by a reduction in lymphocytes (Figure 1A). No obvious changes in other blood lineages were observed (Figure 1A). Rpl22+/− mice did not differ substantially from Rpl22+/+ mice for any of the evaluated cell subsets (Figure 1A). Flow-cytometric analysis indicated that the decrease in lymphocytes was, in turn, due to a substantial decrease in both CD4+ and CD8+ T cells (Figure 1B). The reduction in peripheral blood CD4+ and CD8+ T cells did not appear to be caused by altered trafficking patterns; the αβ T cell content of most peripheral lymphoid organs analyzed was similarly reduced (Figures 1C and 1D). Interestingly, the defect in generation of mature T cells was selective for the αβ lineage, as shown by the fact that the proportion and absolute number of γδ-lineage T cells in peripheral lymphoid organs was increased (Figures 1C and 1D). This was readily apparent in spleen, axillary, and inguinal lymph nodes (pooled; LN), mesenteric lymph nodes (Mes.LN), Peyer's Patches (PP), and peritoneal exudates (PE) (Figure 1D). These data suggest that Rpl22 deficiency selectively impairs the generation of αβ-lineage cells, while sparing that of γδ-lineage cells. Consistent with this hypothesis, thymic cellularity of Rpl22−/− mice was markedly reduced to about 1% of control. This reduction was caused by a severe impairment in development of αβ-lineage precursors to the more mature DP and SP stages (Figure 2A). In particular, Rpl22 deficiency appeared to arrest development at the β-selection checkpoint. Indeed, relative to Rpl22-expressing LM, the proportion of CD44−CD25+ DN3 thymocytes at the β-selection checkpoint was increased in Rpl22−/− mice, and there was a corresponding decrease in the proportion that had developed beyond the β-selection checkpoint to the CD44−CD25− DN4 stage (Figure 2B). In contrast to the severe impairment in development of αβ-lineage T cells, development of γδ-lineage cells appeared to be relatively resistant to the effects of Rpl22 deficiency. The proportion of γδ-lineage cells was increased and the absolute number was only moderately reduced (Figure 2C). The effect of Rpl22 deficiency on thymocyte development is reminiscent of that of pre-Tα (pTα) deficiency. pTα deficiency abrogates pre-TCR signaling, thereby selectively impairing development of αβ-lineage T cells by arresting their maturation at the β-selection checkpoint (Fehling et al., 1995Fehling H.J. Krotkova A. Saint-Ruf C. von Boehmer H. Crucial role of the pre-T-cell receptor alpha gene in development of alpha beta but not gamma delta T cells.Nature. 1995; 375: 795-798Crossref PubMed Scopus (457) Google Scholar). Like Rpl22−/− mice, pTα-deficient (Ptrca−/−) mice exhibited similar decreases in CD4+ and CD8+ αβ-lineage T cells and an increase in γδ-lineage cells. However, there were some important differences. Thymic cellularity of Ptrca−/− mice was not reduced as severely as in Rpl22−/− mice; more DP thymocytes were produced in Ptrca−/− mice; and, whereas the absolute number of DN3 thymocytes was decreased in Rpl22−/− mice, the number of DN3 cells was increased in Ptrca−/− mice (Figures 2A and 2B). Taken together, these data suggest that Rpl22 deficiency impairs development of αβ-lineage cells by a different mechanism than pTα deficiency. Because of the ubiquitous expression of Rpl22, it was important to determine whether the defect in T cell development in Rpl22−/− mice was intrinsic to the hematopoietic compartment or the surrounding stroma. To distinguish these possibilities, we constructed bone-marrow (BM) chimeras. We found that the profile of the developing thymocytes reflected the source of the donor BM. Chimeras produced with BM from Rpl22+/+ mice exhibited normal proportions of thymic CD4 and CD8 subsets, irrespective of the host genotype (Figure 3A; Figure S2). In contrast, chimeras generated with Rpl22−/− BM exhibited the abnormal Rpl22−/− thymic phenotype, with a striking reduction in DP thymocytes and increased representation of DN thymocytes, many of which are of the γδ lineage (Figures 2 and 3A). Because the defect in thymocyte development was cell autonomous, we sought to determine whether it could be complemented by ectopic expression of Rpl22. To do so, fetal liver hematopoietic precursors from Rpl22+/+, Rpl22+/−, and Rpl22−/− mice were cultured on monolayers of OP9-DL1 cells, which support development of T cells in vitro from hematopoietic precursors (Schmitt and Zuniga-Pflucker, 2002Schmitt T.M. Zuniga-Pflucker J.C. Induction of T cell development from hematopoietic progenitor cells by delta-like-1 in vitro.Immunity. 2002; 17: 749-756Abstract Full Text Full Text PDF PubMed Scopus (851) Google Scholar). Precursors from Rpl22−/− mice that were infected with an empty IRES-GFP retrovirus (pMiG) were impaired in their ability to develop from the DN to the DP stage (Figure 3B; left panels of d8 and d16 columns). Importantly, the ability of Rpl22−/− precursors to develop to the DP stage was dramatically improved upon retroviral transduction with an Rpl22 cDNA (pMiG-Rpl22) (Figure 3B; right panels of d8 and d16 columns). Retroviral transduction with pMiG-Rpl22 did not completely restore development of DP thymocytes to the extent observed in control LM, perhaps because the amount of Rpl22 protein expressed by pMiG-Rpl22-transduced Rpl22−/− precursors was not restored to that expressed by control LM (Figure 3C). Taken together, these data demonstrate that the defect in thymocyte development in Rpl22−/− mice is cell autonomous and results from the lack of Rpl22 protein expression. Because T cell development in Rpl22−/− mice was blocked at the β-selection checkpoint at DN3, we asked whether this resulted from the inability to produce TCRβ protein and initiate pre-TCR signaling. Accordingly, we performed intracellular staining to detect expression of the TCRβ protein. In Rpl22-expressing DN3 (CD25+) thymocytes, the fraction of cells that expressed TCRβ protein was between 15% and 20%, consistent with previous analysis (Figure 4A; Wilson et al., 1999Wilson A. Capone M. MacDonald H.R. Unexpectedly late expression of intracellular CD3epsilon and TCR gammadelta proteins during adult thymus development.Int. Immunol. 1999; 11: 1641-1650Crossref PubMed Scopus (56) Google Scholar). The fraction of TCRβ-expressing cells was increased further to 55%–60% among Rpl22-expressing (CD25−) DN4 thymocytes, as is expected for cells that have traversed the β-selection checkpoint (Figure 4A; Dudley et al., 1994Dudley E.C. Petrie H.T. Shah L.M. Owen M.J. Hayday A.C. T cell receptor beta chain gene rearrangement and selection during thymocyte development in adult mice.Immunity. 1994; 1: 83-93Abstract Full Text PDF PubMed Scopus (252) Google Scholar, Hoffman et al., 1996Hoffman E.S. Passoni L. Crompton T. Leu T.M. Schatz D.G. Koff A. Owen M.J. Hayday A.C. Productive T-cell receptor beta-chain gene rearrangement: coincident regulation of cell cycle and clonality during development in vivo.Genes Dev. 1996; 10: 948-962Crossref PubMed Scopus (271) Google Scholar, Wilson et al., 1999Wilson A. Capone M. MacDonald H.R. Unexpectedly late expression of intracellular CD3epsilon and TCR gammadelta proteins during adult thymus development.Int. Immunol. 1999; 11: 1641-1650Crossref PubMed Scopus (56) Google Scholar). Interestingly, in Rpl22−/− mice, the fraction of TCRβ-expressing DN3 thymocytes was actually higher (∼55%); however, very few of those TCRβ-expressing cells downregulated CD25 and differentiated to the DN4 stage, suggesting that there was selection against differentiation of TCRβ-expressing cells in Rpl22−/− mice (Figure 4A). Consistent with this possibility, most of the DN4 thymocytes in Rpl22−/− mice lacked TCRβ expression, but many did express the γδTCR complex, showing that the γδTCR complex was capable of promoting development to the DN4 stage (Figure 4B). The failure of most Rpl22−/− DN3 thymocytes to mature to the DN4 stage did not appear to result from the absence of signaling, because about 30% of Rpl22−/− DN3 exhibited increased expression of the CD5 activation marker (Figure 4C; right) indicating that they have received a signal. These data suggest that the developmental arrest in Rpl22−/− mice was not caused by a defect in pre-TCR expression or function, because most Rpl22−/− DN3 expressed TCRβ protein and appeared to have received a TCR signal. Moreover, the enrichment of γδTCR-expressing cells and depletion of TCRβ-expressing cells among the DN4 subpopulation demonstrates that the impaired development of DN3 thymocytes to the DN4 stage caused by Rpl22 deficiency was selective for cells of the αβ lineage. Interestingly, despite the fact that B cell progenitors undergo a similar transition in response to signals by the pre-B cell receptor, neither that transition nor B cell development in general was substantially altered in Rpl22−/− mice (Figure S3; Melchers, 2005Melchers F. The pre-B-cell receptor: selector of fitting immunoglobulin heavy chains for the B-cell repertoire.Nat. Rev. Immunol. 2005; 5: 578-584Crossref PubMed Scopus (136) Google Scholar). We reasoned that αβ-lineage precursors might fail to develop in Rpl22−/− mice because they undergo apoptosis. Consistent with this hypothesis, flow-cytometry analysis revealed that significantly fewer Rpl22−/− thymocytes fall in the FSC-SSC live cell gate and there was a commensurate increase in those that stain positive for the apoptotic marker Annexin V (Figures 4D and 4E; Fadok et al., 1992Fadok V.A. Voelker D.R. Campbell P.A. Cohen J.J. Bratton D.L. Henson P.M. Exposure of phosphatidylserine on the surface of apoptotic lymphocytes triggers specific recognition and removal by macrophages.J. Immunol. 1992; 148: 2207-2216PubMed Google Scholar). Like the defect in thymocyte development, the excessive apoptosis observed in Rpl22−/− thymocytes was cell autonomous (Figures S2 and S4) and complemented by ectopic expression of Rpl22 (Figure S5). Quantification of annexin staining on electronically gated thymocyte subsets revealed that the DP and CD4+ subpopulations exhibited the greatest increase in apoptosis in Rpl22−/− mice (Figure 4F). Curiously, DN thymocytes did not exhibit an increase in apoptosis (Figure 4F), perhaps because apoptotic cells among this subset may be rapidly eliminated by phagocytosis (Schlegel et al., 2000Schlegel R.A. Callahan M.K. Williamson P. The central role of phosphatidylserine in the phagocytosis of apoptotic thymocytes.Ann. N Y Acad. Sci. 2000; 926: 217-225Crossref PubMed Scopus (35) Google Scholar). Importantly, in contrast to αβ-lineage thymocytes, Rpl22−/− γδ-lineage thymocytes (i.e., γδTCR-expressing cells) did not exhibit an increase in apoptosis (Figure 4G). Differential susceptibility of Rpl22−/− αβ- and γδ-lineage cells to apoptosis was also manifested upon anti-CD3 stimulation of peripheral T cells (Figure 4H). The basis for differential sensitivity is unclear but did not appear to result from the inability to transduce signals, because αβTCR-expressing cells from Rpl22−/− mice exhibited normal induction of activation markers CD25 and CD69 (Figure S6). To investigate whether the differential sensitivity of αβ-lineage precursors to Rpl22 deficiency was associated with differences in Rpl22 expression, we examined Rpl22 protein expression by immunoblotting. As has been reported for other ribosomal structural proteins, anti-CD3 stimulation of mature T cells increased the expression of Rpl22 (Figure 5A; Asmal et al., 2003Asmal M. Colgan J. Naef F. Yu B. Lee Y. Magnasco M. Luban J. Production of ribosome components in effector CD4+ T cells is accelerated by TCR stimulation and coordinated by ERK-MAPK.Immunity. 2003; 19: 535-548Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). Moreover, this was also true for Rag2−/− thymocytes stimulated in vivo with anti-CD3, a stimulus that mimics pre-TCR signaling (Figure 5B; Shinkai and Alt, 1994Shinkai Y. Alt F.W. CD3 epsilon-mediated signals rescue the development of CD4+CD8+ thymocytes in RAG-2−/− mice in the absence of TCR beta chain expression.Int. Immunol. 1994; 6: 995-1001Crossref PubMed Scopus (190) Google Scholar). The increase in Rpl22 protein was not accompanied by increased mRNA (data not shown). Rpl22 protein abundance also increased as DN3 thymocytes traversed the β-selection checkpoint to the DN4 stage (Figure 5C). Importantly, expression of Rpl22 in DN4 thymocytes was greater than in γδ-lineage cells (Figure 5C). The increased expression of Rpl22 occurs during the developmental transition that is blocked by Rpl22 deficiency, suggesting that increased Rpl22 expression is important for αβ-lineage precursors to differentiate in response to pre-TCR signaling. Because a recent report demonstrated that conditional ablation of the Rps6 gene in immature thymocytes completely blocked thymocyte development through activation of a p53-dependent checkpoint, we considered that Rpl22 deficiency might be acting through a similar mechanism (Sulic et al., 2005Sulic S. Panic L. Barkic M. Mercep M. Uzelac M. Volarevic S. Inactivation of S6 ribosomal protein gene in T lymphocytes activates a p53-dependent checkpoint response.Genes Dev. 2005; 19: 3070-3082Crossref PubMed Scopus (112) Google Scholar). In fact, p53 protein expression was found to be higher in thymocytes from Rpl22−/− mice than in Rpl22+/+ LM (Figure 5D). Interestingly, however, p53 expression was not elevated in untreated Rpl22−/− mouse embryonic fibroblasts (MEF), but could be induced upon UV irradiation (Figure 5D). This indicates that Rpl" @default.
• W2024704480 created "2016-06-24" @default.
• W2024704480 creator A5002152110 @default.
• W2024704480 creator A5004177626 @default.
• W2024704480 creator A5016054194 @default.
• W2024704480 creator A5017378893 @default.
• W2024704480 creator A5017721524 @default.
• W2024704480 creator A5026026841 @default.
• W2024704480 creator A5063285107 @default.
• W2024704480 creator A5065560657 @default.
• W2024704480 creator A5073708445 @default.
• W2024704480 creator A5078673290 @default.
• W2024704480 date "2007-06-01" @default.
• W2024704480 modified "2023-09-28" @default.
• W2024704480 title "Ablation of Ribosomal Protein L22 Selectively Impairs αβ T Cell Development by Activation of a p53-Dependent Checkpoint" @default.
• W2024704480 cites W1222624887 @default.
• W2024704480 cites W1499161600 @default.
• W2024704480 cites W1500023054 @default.
• W2024704480 cites W1522872684 @default.
• W2024704480 cites W1591147683 @default.
• W2024704480 cites W1718537877 @default.
• W2024704480 cites W1922927689 @default.
• W2024704480 cites W1940692702 @default.
• W2024704480 cites W1966249293 @default.
• W2024704480 cites W1968220278 @default.
• W2024704480 cites W1968696005 @default.
• W2024704480 cites W1978686705 @default.
• W2024704480 cites W1996247810 @default.
• W2024704480 cites W1999440327 @default.
• W2024704480 cites W2003097999 @default.
• W2024704480 cites W2005571932 @default.
• W2024704480 cites W2018216682 @default.
• W2024704480 cites W2022162825 @default.
• W2024704480 cites W2028735719 @default.
• W2024704480 cites W2028942940 @default.
• W2024704480 cites W2029350186 @default.
• W2024704480 cites W2030688963 @default.
• W2024704480 cites W2030724377 @default.
• W2024704480 cites W2032917789 @default.
• W2024704480 cites W2039704215 @default.
• W2024704480 cites W2056429839 @default.
• W2024704480 cites W2058288999 @default.
• W2024704480 cites W2060531978 @default.
• W2024704480 cites W2066619117 @default.
• W2024704480 cites W2075566981 @default.
• W2024704480 cites W2078191138 @default.
• W2024704480 cites W2082399676 @default.
• W2024704480 cites W2084459589 @default.
• W2024704480 cites W2088740611 @default.
• W2024704480 cites W2091111894 @default.
• W2024704480 cites W2101091611 @default.
• W2024704480 cites W2112545544 @default.
• W2024704480 cites W2116704889 @default.
• W2024704480 cites W2116755440 @default.
• W2024704480 cites W2118370134 @default.
• W2024704480 cites W2119807671 @default.
• W2024704480 cites W2126464081 @default.
• W2024704480 cites W2128845278 @default.
• W2024704480 cites W2131510524 @default.
• W2024704480 cites W2134937174 @default.
• W2024704480 cites W2135503300 @default.
• W2024704480 cites W2135950183 @default.
• W2024704480 cites W2137764426 @default.
• W2024704480 cites W2140899494 @default.
• W2024704480 cites W2142437744 @default.
• W2024704480 cites W2149641002 @default.
• W2024704480 cites W2150386153 @default.
• W2024704480 cites W2155926744 @default.
• W2024704480 cites W2162297821 @default.
• W2024704480 cites W2168276925 @default.
• W2024704480 cites W2172228770 @default.
• W2024704480 cites W258316223 @default.
• W2024704480 cites W4313335965 @default.
• W2024704480 doi "https://doi.org/10.1016/j.immuni.2007.04.012" @default.
• W2024704480 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/17555992" @default.
• W2024704480 hasPublicationYear "2007" @default.
• W2024704480 type Work @default.
• W2024704480 sameAs 2024704480 @default.
• W2024704480 citedByCount "161" @default.
• W2024704480 countsByYear W20247044802012 @default.
• W2024704480 countsByYear W20247044802013 @default.
• W2024704480 countsByYear W20247044802014 @default.
• W2024704480 countsByYear W20247044802015 @default.
• W2024704480 countsByYear W20247044802016 @default.
• W2024704480 countsByYear W20247044802017 @default.
• W2024704480 countsByYear W20247044802018 @default.
• W2024704480 countsByYear W20247044802019 @default.
• W2024704480 countsByYear W20247044802020 @default.
• W2024704480 countsByYear W20247044802021 @default.
• W2024704480 countsByYear W20247044802022 @default.
• W2024704480 countsByYear W20247044802023 @default.
• W2024704480 crossrefType "journal-article" @default.
• W2024704480 hasAuthorship W2024704480A5002152110 @default.
• W2024704480 hasAuthorship W2024704480A5004177626 @default.
• W2024704480 hasAuthorship W2024704480A5016054194 @default.
• W2024704480 hasAuthorship W2024704480A5017378893 @default.
• W2024704480 hasAuthorship W2024704480A5017721524 @default.
• W2024704480 hasAuthorship W2024704480A5026026841 @default.

• next