RESUMO
In the pathophysiologic setting of acute and chronic kidney injury, the excessive activation and recruitment of blood-borne monocytes prompts their differentiation into inflammatory macrophages, a process that leads to progressive glomerulosclerosis and interstitial fibrosis. Importantly, this differentiation of monocytes into macrophages requires the meticulous coordination of gene expression at both the transcriptional and post-transcriptional level. The transcriptomes of these cells are ultimately determined by RNA-binding proteins such as QUAKING (QKI), that define their pre-mRNA splicing and mRNA transcript patterns. Using two mouse models, namely (1) quaking viable mice (qkv) and (2) the conditional deletion in the myeloid cell lineage using the lysozyme 2-Cre (QKIFL/FL;LysM-Cre mice), we demonstrate that the abrogation of QKI expression in the myeloid cell lineage reduces macrophage infiltration following kidney injury induced by unilateral urethral obstruction (UUO). The qkv and QKIFL/FL;LysM-Cre mice both showed significant diminished interstitial collagen deposition and fibrosis in the UUO-damaged kidney, as compared to wild-type littermates. We show that macrophages isolated from QKIFL/FL;LysM-Cre mice are associated with defects in pre-mRNA splicing. Our findings demonstrate that reduced expression of the alternative splice regulator QKI in the cells of myeloid lineage attenuates renal interstitial fibrosis, suggesting that inhibition of this splice regulator may be of therapeutic value for certain kidney diseases.
RESUMO
The oligodendrocyte lineage is responsible for myelination of the central nervous system. Post-translational modifications are known to regulate oligodendrocyte precursor cell (OPC) differentiation into mature myelinating oligodendrocytes. The role of arginine methylation during oligodendrocyte differentiation and myelination is still poorly understood. We generated mice depleted of PRMT5 in OPCs using Olig2-Cre, and these mice developed severe hypomyelination and died at the third post-natal week. PRMT5-deficient cells have lower levels of PDGFRα at the plasma membrane due to increased degradation by the Cbl E3 ligase. Mechanistically, the loss of arginine methylation at R554 of the PDGFRα intracellular domain unmasks a Cbl binding site at Y555. We observed the progressive decrease in PRMT5 during oligodendrocyte differentiation, and we show that one role of this decrease is to downregulate growth signals provided by PDGFRα to initiate oligodendrocyte differentiation and myelination. More broadly, the inhibition of PRMT5 may be used therapeutically to manipulate PDGFRα bioavailability.
Assuntos
Diferenciação Celular/fisiologia , Oligodendroglia/citologia , Proteína-Arginina N-Metiltransferases/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Células Cultivadas , Camundongos , Bainha de Mielina/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neurogênese/fisiologiaRESUMO
Quiescent muscle stem cells (MSCs) become activated in response to skeletal muscle injury to initiate regeneration. Activated MSCs proliferate and differentiate to repair damaged fibers or self-renew to maintain the pool and ensure future regeneration. The balance between self-renewal, proliferation, and differentiation is a tightly regulated process controlled by a genetic cascade involving determinant transcription factors such as Pax7, Myf5, MyoD, and MyoG. Recently, there have been several reports about the role of arginine methylation as a requirement for epigenetically mediated control of muscle regeneration. Here we report that the protein arginine methyltransferase 1 (PRMT1) is expressed in MSCs and that conditional ablation of PRMT1 in MSCs using Pax7CreERT2 causes impairment of muscle regeneration. Importantly, PRMT1-deficient MSCs have enhanced cell proliferation after injury but are unable to terminate the myogenic differentiation program, leading to regeneration failure. We identify the coactivator of Six1, Eya1, as a substrate of PRMT1. We show that PRMT1 methylates Eya1 in vitro and that loss of PRMT1 function in vivo prevents Eya1 methylation. Moreover, we observe that PRMT1-deficient MSCs have reduced expression of Eya1/Six1 target MyoD due to disruption of Eya1 recruitment at the MyoD promoter and subsequent Eya1-mediated coactivation. These findings suggest that arginine methylation by PRMT1 regulates muscle stem cell fate through the Eya1/Six1/MyoD axis.
Assuntos
Arginina/metabolismo , Linhagem da Célula , Proteína-Arginina N-Metiltransferases/metabolismo , Células-Tronco/citologia , Animais , Diferenciação Celular , Proliferação de Células , Autorrenovação Celular , Células Cultivadas , Proteínas de Homeodomínio/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Metilação , Camundongos Endogâmicos C57BL , Células Musculares/citologia , Desenvolvimento Muscular , Proteína MyoD/metabolismo , Proteínas Nucleares/metabolismo , Peptídeos/metabolismo , Análise Serial de Proteínas , Proteínas Tirosina Fosfatases/metabolismo , Regeneração , Especificidade por Substrato , Transcrição GênicaRESUMO
RNA binding proteins required for the maintenance of myelin and axoglial junctions are unknown. Herein, we report that deletion of the Quaking (QKI) RNA binding proteins in oligodendrocytes (OLs) using Olig2-Cre results in mice displaying rapid tremors at postnatal day 10, followed by death at postnatal week 3. Extensive CNS hypomyelination was observed as a result of OL differentiation defects during development. The QKI proteins were also required for adult myelin maintenance, because their ablation using PLP-CreERT resulted in hindlimb paralysis with immobility at â¼30 d after 4-hydroxytamoxifen injection. Moreover, deterioration of axoglial junctions of the spinal cord was observed and is consistent with a loss of Neurofascin 155 (Nfasc155) isoform that we confirmed as an alternative splice target of the QKI proteins. Our findings define roles for the QKI RNA binding proteins in myelin development and maintenance, as well as in the generation of Nfasc155 to maintain healthy axoglial junctions. SIGNIFICANCE STATEMENT: Neurofascin 155 is responsible for axoglial junction formation and maintenance. Using a genetic mouse model to delete Quaking (QKI) RNA-binding proteins in oligodendrocytes, we identify QKI as the long-sought regulator of Neurofascin alternative splicing, further establishing the role of QKI in oligodendrocyte development and myelination. We establish a new role for QKI in myelin and axoglial junction maintenance using an inducible genetic mouse model that deletes QKI in mature oligodendrocytes. Loss of QKI in adult oligodendrocytes leads to phenotypes reminiscent of the experimental autoimmune encephalomyelitis mouse model with complete hindlimb paralysis and death by 30 d after induction of QKI deletion.
Assuntos
Axônios , Moléculas de Adesão Celular/biossíntese , Moléculas de Adesão Celular/genética , Bainha de Mielina/genética , Fatores de Crescimento Neural/biossíntese , Fatores de Crescimento Neural/genética , Neuroglia , Proteínas de Ligação a RNA/genética , Processamento Alternativo , Animais , Animais Recém-Nascidos , Ataxia/genética , Regulação da Expressão Gênica/genética , Camundongos , Camundongos Knockout , Oligodendroglia , Paralisia/genética , Ratos , Ratos Sprague-DawleyRESUMO
Regeneration of skeletal muscle requires the continued presence of quiescent muscle stem cells (satellite cells), which become activated in response to injury. Here, we report that whole-body protein arginine methyltransferase PRMT7(-/-) adult mice and mice conditionally lacking PRMT7 in satellite cells using Pax7-CreERT2 both display a significant reduction in satellite cell function, leading to defects in regenerative capacity upon muscle injury. We show that PRMT7 is preferentially expressed in activated satellite cells and, interestingly, PRMT7-deficient satellite cells undergo cell-cycle arrest and premature cellular senescence. These defects underlie poor satellite cell stem cell capacity to regenerate muscle and self-renew after injury. PRMT7-deficient satellite cells express elevated levels of the CDK inhibitor p21CIP1 and low levels of its repressor, DNMT3b. Restoration of DNMT3b in PRMT7-deficient cells rescues PRMT7-mediated senescence. Our findings define PRMT7 as a regulator of the DNMT3b/p21 axis required to maintain muscle stem cell regenerative capacity.
Assuntos
Inibidor de Quinase Dependente de Ciclina p21/genética , DNA (Citosina-5-)-Metiltransferases/genética , Músculo Esquelético/metabolismo , Proteína-Arginina N-Metiltransferases/genética , Regeneração/genética , Células Satélites de Músculo Esquelético/metabolismo , Células-Tronco/metabolismo , Animais , Pontos de Checagem do Ciclo Celular/genética , Diferenciação Celular , Senescência Celular , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , DNA (Citosina-5-)-Metiltransferases/metabolismo , Feminino , Regulação da Expressão Gênica , Integrases/genética , Integrases/metabolismo , Masculino , Camundongos , Camundongos Knockout , Músculo Esquelético/citologia , Fator de Transcrição PAX7/genética , Fator de Transcrição PAX7/metabolismo , Proteína-Arginina N-Metiltransferases/deficiência , Células Satélites de Músculo Esquelético/citologia , Transdução de Sinais , Células-Tronco/citologia , DNA Metiltransferase 3BRESUMO
The quaking (qkI) gene encodes 3 major alternatively spliced isoforms that contain unique sequences at their C termini dictating their cellular localization. QKI-5 is predominantly nuclear, whereas QKI-6 is distributed throughout the cell and QKI-7 is cytoplasmic. The QKI isoforms are sequence-specific RNA binding proteins expressed mainly in glial cells modulating RNA splicing, export, and stability. Herein, we identify a new role for the QKI proteins in the regulation of microRNA (miRNA) processing. We observed that small interfering RNA (siRNA)-mediated QKI depletion of U343 glioblastoma cells leads to a robust increase in miR-7 expression. The processing from primary to mature miR-7 was inhibited in the presence QKI-5 and QKI-6 but not QKI-7, suggesting that the nuclear localization plays an important role in the regulation of miR-7 expression. The primary miR-7-1 was bound by the QKI isoforms in a QKI response element (QRE)-specific manner. We observed that the pri-miR-7-1 RNA was tightly bound to Drosha in the presence of the QKI isoforms, and this association was not observed in siRNA-mediated QKI or Drosha-depleted U343 glioblastoma cells. Moreover, the presence of the QKI isoforms led to an increase presence of pri-miR-7 in nuclear foci, suggesting that pri-miR-7-1 is retained in the nucleus by the QKI isoforms. miR-7 is known to target the epidermal growth factor (EGF) receptor (EGFR) 3' untranslated region (3'-UTR), and indeed, QKI-deficient U343 cells had reduced EGFR expression and decreased ERK activation in response to EGF. Elevated levels of miR-7 are associated with cell cycle arrest, and it was observed that QKI-deficient U343 that harbor elevated levels of miR-7 exhibited defects in cell proliferation that were partially rescued by the addition of a miR-7 inhibitor. These findings suggest that the QKI isoforms regulate glial cell function and proliferation by regulating the processing of certain miRNAs.
Assuntos
MicroRNAs/genética , Neuroglia/fisiologia , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Processos de Crescimento Celular/fisiologia , Linhagem Celular , Núcleo Celular/genética , Núcleo Celular/metabolismo , Fator de Crescimento Epidérmico/genética , Fator de Crescimento Epidérmico/metabolismo , Receptores ErbB/genética , Receptores ErbB/metabolismo , Células HEK293 , Humanos , Sistema de Sinalização das MAP Quinases/genética , MicroRNAs/biossíntese , Neuroglia/metabolismo , Ligação Proteica , Isoformas de Proteínas , Ribonuclease III/genética , Ribonuclease III/metabolismoRESUMO
Sam68, the Src-associated substrate during mitosis of 68 kDa, belongs to the large class of heteronuclear ribonucleoprotein particle K (hnRNP K) homology (KH) domain family of RNA-binding proteins. Sam68 contains a single KH domain harboring conserved N- and C-terminal sequences required for RNA binding and homodimerization. The KH domain is one of the most prevalent RNA binding domains that directly contacts single-stranded RNA. Sam68 has been implicated in numerous aspects of RNA metabolism including alternative splicing and polysomal recruitment of mRNAs. Studies in mice have revealed physiological roles linking Sam68 to osteoporosis, obesity, cancer, infertility and ataxia. Recent publications have greatly enhanced our understanding of Sam68 mechanism of action in addition to its cellular role. Herein, we will discuss the latest advances in the literature pertaining to obesity and neuronal development.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adipogenia/fisiologia , Proteínas de Ligação a DNA/metabolismo , Neurogênese/fisiologia , Proteínas de Ligação a RNA/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Processamento Alternativo/fisiologia , Animais , Ataxia/genética , Ataxia/metabolismo , Proteínas de Ligação a DNA/genética , Humanos , Infertilidade/genética , Infertilidade/metabolismo , Camundongos , Neoplasias/genética , Neoplasias/metabolismo , Obesidade/genética , Obesidade/metabolismo , Osteoporose/genética , Osteoporose/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genéticaRESUMO
Arginine methylation of histones is a well-known regulator of gene expression. Protein arginine methyltransferase 6 (PRMT6) has been shown to function as a transcriptional repressor by methylating the histone H3 arginine 2 [H3R2(me2a)] repressive mark; however, few targets are known. To define the physiological role of PRMT6 and to identify its targets, we generated PRMT6(-/-) mouse embryo fibroblasts (MEFs). We observed that early passage PRMT6(-/-) MEFs had growth defects and exhibited the hallmarks of cellular senescence. PRMT6(-/-) MEFs displayed high transcriptional levels of p53 and its targets, p21 and PML. Generation of PRMT6(-/-); p53(-/-) MEFs prevented the premature senescence, suggesting that the induction of senescence is p53-dependent. Using chromatin immunoprecipitation assays, we observed an enrichment of PRMT6 and H3R2(me2a) within the upstream region of Trp53. The PRMT6 association and the H3R2(me2a) mark were lost in PRMT6(-/-) MEFs and an increase in the H3K4(me3) activator mark was observed. Our findings define a new regulator of p53 transcriptional regulation and define a role for PRMT6 and arginine methylation in cellular senescence.
Assuntos
Proteínas de Drosophila/fisiologia , Regulação da Expressão Gênica , Proteína-Arginina N-Metiltransferases/fisiologia , Transcrição Gênica , Proteína Supressora de Tumor p53/genética , Animais , Células Cultivadas , Senescência Celular/genética , Proteínas de Drosophila/genética , Camundongos , Camundongos Knockout , Proteína-Arginina N-Metiltransferases/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas ras/genéticaRESUMO
We report that mice ablated for the Sam68 RNA-binding protein exhibit a lean phenotype as a result of increased energy expenditure, decreased commitment to early adipocyte progenitors, and defects in adipogenic differentiation. The Sam68(-/-) mice were protected from obesity, insulin resistance, and glucose intolerance induced with a high-fat diet. To identify the alternative splice events regulated by Sam68, genome-wide exon usage profiling in white adipose tissue was performed. Adipocytes from Sam68(-/-) mice retained intron 5 within the mTOR transcript introducing a premature termination codon, leading to an unstable mRNA. Consequently, Sam68-depleted cells had reduced mTOR levels resulting in lower levels of insulin-stimulated S6 and Akt phosphorylation leading to defects in adipogenesis, and this defect was rescued by the exogenous expression of full-length mTOR. Sam68 bound intronic splice elements within mTOR intron 5 required for the usage of the 5' splice site. We propose that Sam68 regulates alternative splicing during adipogenesis.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Adipogenia/genética , Processamento Alternativo , Proteínas de Ligação a RNA/genética , Serina-Treonina Quinases TOR/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Composição Corporal/genética , Metabolismo Energético/genética , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Complexos Multiproteicos , Células NIH 3T3 , Fenótipo , Proteínas/metabolismo , Proteínas/fisiologia , Proteínas de Ligação a RNA/fisiologia , Transdução de Sinais/genéticaRESUMO
The MRE11/RAD50/NBS1 complex is the primary sensor rapidly recruited to DNA double-strand breaks (DSBs). MRE11 is known to be arginine methylated by PRMT1 within its glycine-arginine-rich (GAR) motif. In this study, we report a mouse knock-in allele of Mre11 that substitutes the arginines with lysines in the GAR motif and generates the MRE11(RK) protein devoid of methylated arginines. The Mre11(RK/RK) mice were hypersensitive to γ-irradiation (IR) and the cells from these mice displayed cell cycle checkpoint defects and chromosome instability. Moreover, the Mre11(RK/RK) MEFs exhibited ATR/CHK1 signaling defects and impairment in the recruitment of RPA and RAD51 to the damaged sites. The M(RK)RN complex formed and localized to the sites of DNA damage and normally activated the ATM pathway in response to IR. The M(RK)RN complex exhibited exonuclease and DNA-binding defects in vitro responsible for the impaired DNA end resection and ATR activation observed in vivo in response to IR. Our findings provide genetic evidence for the critical role of the MRE11 GAR motif in DSB repair, and demonstrate a mechanistic link between post-translational modifications at the MRE11 GAR motif and DSB processing, as well as the ATR/CHK1 checkpoint signaling.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Quebras de DNA de Cadeia Dupla , Enzimas Reparadoras do DNA/metabolismo , Reparo do DNA/fisiologia , Proteínas de Ligação a DNA/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Motivos de Aminoácidos , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Pontos de Checagem do Ciclo Celular , Células Cultivadas , Quinase 1 do Ponto de Checagem , Instabilidade Cromossômica , Enzimas Reparadoras do DNA/química , Proteínas de Ligação a DNA/química , Ativação Enzimática , Raios gama , Técnicas de Introdução de Genes , Proteína Homóloga a MRE11 , Camundongos , Proteínas Quinases/metabolismo , Rad51 Recombinase/metabolismo , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismoRESUMO
PRMT5 is a type II protein arginine methyltranferase that catalyzes monomethylation and symmetric dimethylation of arginine residues. PRMT5 is functionally involved in a variety of biological processes including embryo development and circadian clock regulation. However, the role of PRMT5 in oligodendrocyte differentiation and central nervous system myelination is unknown. Here we show that PRMT5 expression gradually increases throughout postnatal brain development, coinciding with the period of active myelination. PRMT5 expression was observed in neurons, astrocytes, and oligodendrocytes. siRNA-mediated depletion of PRMT5 in mouse primary oligodendrocyte progenitor cells abrogated oligodendrocyte differentiation. In addition, the PRMT5-depleted oligodendrocyte progenitor and C6 glioma cells expressed high levels of the inhibitors of differentiation/DNA binding, Id2 and Id4, known repressors of glial cell differentiation. We observed that CpG-rich islands within the Id2 and Id4 genes were bound by PRMT5 and were hypomethylated in PRMT5-deficient cells, suggesting that PRMT5 plays a role in gene silencing during glial cell differentiation. Our findings define a role of PRMT5 in glial cell differentiation and link PRMT5 to epigenetic changes during oligodendrocyte differentiation.
Assuntos
Diferenciação Celular/fisiologia , Inativação Gênica/fisiologia , Proteína 2 Inibidora de Diferenciação/biossíntese , Proteínas Inibidoras de Diferenciação/biossíntese , Bainha de Mielina/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteínas Metiltransferases/metabolismo , Animais , Astrócitos/citologia , Astrócitos/metabolismo , Linhagem Celular Tumoral , Ilhas de CpG/fisiologia , Proteína 2 Inibidora de Diferenciação/genética , Proteínas Inibidoras de Diferenciação/genética , Metilação , Camundongos , Bainha de Mielina/genética , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Neurônios/metabolismo , Proteínas Metiltransferases/genética , Proteína-Arginina N-Metiltransferases , Ratos , Ratos Sprague-DawleyRESUMO
Sam68, Src associated in mitosis of 68 kDa, is a known RNA-binding protein and a signaling adaptor protein for tyrosine kinases. However, the proteins associated with Sam68 and the existence of a Sam68 complex, its mass, and regulation are, however, unknown. Herein we identify a large Sam68 complex with a mass >1 MDa in HeLa cells that is composed of approximately 40 proteins using an immunoprecipitation followed by a mass spectrometry approach. Many of the proteins identified are RNA-binding proteins and are known components of a previously identified structure termed the spreading initiation center. The large Sam68 complex is a ribonucleoprotein complex, as treatment with RNases caused a shift in the molecular mass of the complex to 200-450 kDa. Moreover, treatment of HeLa cells with phorbol 12-myristate 13-acetate or epidermal growth factor induced the disassociation of Sam68 from the large complex and the appearance of Sam68 within the smaller complex. Actually, in certain cell lines such as breast cancer cell lines MCF-7 and BT-20, Sam68 exists in equilibrium between a large and a small complex. The appearance of the small Sam68 complex in cells correlates with the ability of Sam68 to promote the alternative splicing of CD44 and cell migration. Our findings show that Sam68 exists in equilibrium in transformed cells between two complexes and that extracellular signals, such as epidermal growth factor stimulation, promote alternative splicing by modulating the composition of the Sam68 complex.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias da Mama/metabolismo , Movimento Celular , Proteínas de Ligação a DNA/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Proteínas de Ligação a RNA/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Processamento Alternativo , Western Blotting , Núcleo Celular/metabolismo , Proteínas de Ligação a DNA/genética , Feminino , Imunofluorescência , Células HeLa , Humanos , Receptores de Hialuronatos/genética , Receptores de Hialuronatos/metabolismo , Imunoprecipitação , Ligação Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Acetato de Tetradecanoilforbol/farmacologia , Células Tumorais CultivadasRESUMO
Sam68 is a KH-type RNA-binding protein involved in several steps of RNA metabolism with potential implications in cell differentiation and cancer. However, its physiological roles are still poorly understood. Herein, we show that Sam68(-/-) male mice are infertile and display several defects in spermatogenesis, demonstrating an essential role for Sam68 in male fertility. Sam68(-/-) mice produce few spermatozoa, which display dramatic motility defects and are unable to fertilize eggs. Expression of a subset of messenger mRNAs (mRNAs) is affected in the testis of knockout mice. Interestingly, Sam68 is associated with polyadenylated mRNAs in the cytoplasm during the meiotic divisions and in round spermatids, when it interacts with the translational machinery. We show that Sam68 is required for polysomal recruitment of specific mRNAs and for accumulation of the corresponding proteins in germ cells and in a heterologous system. These observations demonstrate a novel role for Sam68 in mRNA translation and highlight its essential requirement for the development of a functional male gamete.
Assuntos
Células Germinativas/fisiologia , Biossíntese de Proteínas , RNA Mensageiro/metabolismo , Espermatogênese/fisiologia , Animais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Feminino , Fertilidade/fisiologia , Regulação da Expressão Gênica , Genes Reporter , Células Germinativas/citologia , Masculino , Meiose/fisiologia , Camundongos , Camundongos Knockout , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Gravidez , RNA Mensageiro/genética , Espermatozoides/citologia , Espermatozoides/fisiologia , Testículo/citologia , Testículo/fisiologiaRESUMO
The Src substrate associated in mitosis of 68 kDa (Sam68) is a KH-type RNA binding protein that has been shown to regulate several aspects of RNA metabolism; however, its physiologic role has remained elusive. Herein we report the generation of Sam68-null mice by homologous recombination. Aged Sam68-/- mice preserved their bone mass, in sharp contrast with 12-month-old wild-type littermates in which bone mass was decreased up to approximately 75%. In fact, the bone volume of the 12-month-old Sam68-/- mice was virtually indistinguishable from that of 4-month-old wild-type or Sam68-/- mice. Sam68-/- bone marrow stromal cells had a differentiation advantage for the osteogenic pathway. Moreover, the knockdown of Sam68 using short hairpin RNA in the embryonic mesenchymal multipotential progenitor C3H10T1/2 cells resulted in more pronounced expression of the mature osteoblast marker osteocalcin when differentiation was induced with bone morphogenetic protein-2. Cultures of mouse embryo fibroblasts generated from Sam68+/+ and Sam68-/- littermates were induced to differentiate into adipocytes with culture medium containing pioglitazone and the Sam68-/- mouse embryo fibroblasts shown to have impaired adipocyte differentiation. Furthermore, in vivo it was shown that sections of bone from 12-month-old Sam68-/- mice had few marrow adipocytes compared with their age-matched wild-type littermate controls, which exhibited fatty bone marrow. Our findings identify endogenous Sam68 as a positive regulator of adipocyte differentiation and a negative regulator of osteoblast differentiation, which is consistent with Sam68 being a modulator of bone marrow mesenchymal cell differentiation, and hence bone metabolism, in aged mice.