A novel SATB2 mutation illuminates bone anabolism

一种新的 SATB2 突变阐明了骨合成代谢

• 批准号: 8758799
• 负责人: KARL Leonard INSOGNA
• 金额: $ 21.98万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8758799
• 关键词: Adolescence; Adult; Affect; Alleles; Amino Acids; Anabolism; Animals; Asparagine; Back; Base Pairing; Binding Proteins; Biomechanics; Bone Density; Bone Resorption; Bone remodeling; Calvaria; Cartilage; Cells; Childhood; Clinical Trials; Cyclophosphamide; DNA; DNA Binding; Data; Data Set; Databases; Development; Dideoxy Chain Termination DNA Sequencing; Dual-Energy X-Ray Absorptiometry; Engineering; Experimental Models; Family member; Female; Genes; Genetic Recombination; Genetic Transcription; Genomic DNA; Genotype; In Situ Hybridization; In Vitro; Individual; Knock-in Mouse; Lead; Lysine; Marrow; Matrix Attachment Regions; Measures; Mesenchymal Stem Cells; Metabolic Pathway; Metabolism; Methodology; Molecular; Mus; Mutation; Osteoblasts; Osteoclasts; Osteocytes; Osteogenesis; Osteoporosis; Parents; Pathway interactions; Patients; Phase; Phenotype; Play; Positioning Attribute; Protein Binding; RNA; Rare Diseases; Recording of previous events; Regulator Genes; Reporting; Role; Signal Transduction; Spectroscopy, Fourier Transform Infrared; Testing; Thick; Variant; Woman; Work; bone; bone mass; bone metabolism; bone strength; bone turnover; clinical phenotype; drug discovery; embryonic stem cell; exome; exome sequencing; fly; gain of function mutation; genetic variant; insight; kindred; mineralization; mouse genome; mutant; neutralizing antibody; novel; novel therapeutics; older women; overexpression; public health relevance; research study; skeletal; trait

DESCRIPTION (provided by applicant): The study of uncommon clinical phenotypes can sometimes provide important insights into common genetically influenced traits, such as bone mass. A 50 yo white female was evaluated because of extremely high bone mass (T-scores +6 - +10), radiographically normal appearing bones except for very thick cortices, and chronically and markedly elevated markers of both bone formation and bone resorption. High turnover is usually associated with low bone mass, particularly in adult women, so this patient is very unusual. (It is worth noting that patients with gain-of-function mutations in LRP5 do not have elevated levels of P1NP or CTX.) Whole exome sequencing identified a novel variant in the gene Special AT rich Binding Protein 2 (SATB2) in this individual. A single base pair change, G to C, at position chr2:200193607 (hg19) resulted in an amino acid change, lysine 400 to asparagine (K400N). This allelic variant was not seen in either parent, in 2500 normal individuals or in publicly available databases. SATB2 is a member of the family of proteins that bind to matrix attachment regions (MAR) in DNA and SATB2 has been previously reported to have a critical role in bone formation. Lysine 400 is in the highly conserved CUT1 domain of SATB2 that binds DNA. Lysine 400 is strictly evolutionarily conserved back to flies, indicating a functionally important role for this amino acid. In situ hybridization studies documented high-level specific expression of SABT2 in osteoblasts and osteocytes, but not in cartilage or marrow. Overexpression of the K400N variant in MC3T3-E1 cells led to spontaneous in vitro mineralization. Since at present there are very few targets for bone anabolic therapies, exploring the mechanisms by which the SATB2 K400N mutation results in high bone mass is important. The only way to faithfully recapitulate the genotype of this individual in an experimental model is to replace one wild type SATB2 allele in a mouse with the mutant allele, i.e., by creating a knock-in mouse. To begin to identify the metabolic pathways entrained by this mutation in SATB2, we will pursue the following Specific Aims: 1. Generate knock-in mice bearing the SATB2-K400N mutation using an engineered ES cell clone in which the mutant allele has already been introduced by recombination. 2. Characterize the phenotype of the SATB2- K400N knock-in mice by using densitometric, histomorphometric and biomechanical methodologies and by studying MSCs, osteoblasts and osteoclasts isolated from these animals. 3. Undertake gene profiling studies using RNA isolated from primary calvarial osteoblast cultures from wild type and knock-in mice and Affymetrix whole mouse genome 430 2.0 arrays. High rates of skeletal turnover associated with increasing bone mass are normally only seen during childhood and adolescence, a period of rapid skeletal accrual. The molecular and metabolic pathways by which this occurs are unclear. Whether pathways similar to those are entrained by the SATB2 K400N mutation remains to be determined, but is an intriguing possibility. The proposed studies will help identify new molecular pathways regulating bone anabolism and new targets for drug discovery.

描述（由申请人提供）：对不常见临床表型的研究有时可以提供对常见遗传影响特征（如骨量）的重要见解。一名50岁的白人女性因骨量极高（t评分+6 - +10），x线摄影显示骨骼正常，除了非常厚的皮质，骨形成和骨吸收标志物长期显著升高而被评估。高更替通常与低骨量有关，特别是在成年女性中，所以这个病人很不寻常。（值得注意的是，LRP5功能获得性突变的患者没有P1NP或CTX水平升高。）全外显子组测序在该个体中发现了特殊AT丰富结合蛋白2 （SATB2）基因的新变体。chr2:200193607 （hg19）的碱基对从G到C的变化导致了氨基酸从赖氨酸400到天冬酰胺（K400N）的变化。这种等位基因变异在父母双方、2500名正常个体或公开数据库中均未见。SATB2是DNA中与基质附着区（MAR）结合的蛋白家族的一员，SATB2在骨形成中起关键作用。赖氨酸400位于结合DNA的SATB2的高度保守的CUT1结构域。赖氨酸400在果蝇中具有严格的进化保守性，表明该氨基酸具有重要的功能作用。原位杂交研究证实SABT2在成骨细胞和骨细胞中有高水平的特异性表达，但在软骨或骨髓中没有。MC3T3-E1细胞中K400N变异体的过表达导致自发的体外矿化。由于目前骨合成代谢治疗的靶点很少，因此探索SATB2 K400N突变导致高骨量的机制非常重要。在实验模型中忠实地概括这个个体的基因型的唯一方法是