LncRNA-HIT Functions as an Epigenetic Regulator of Chondrogenesis through Its Recruitment of p100/CBP Complexes.
LncRNA-HIT Functions as an Epigenetic Regulator of Chondrogenesis through Its Recruitment of p100/CBP Complexes.
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DOI：
10.1371/journal.pgen.1005680
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发表时间：
2015-12
期刊：
影响因子：
4.5
通讯作者：
Stadler HS
中科院分区：
文献类型：
--
作者：
Carlson HL;Quinn JJ;Yang YW;Thornburg CK;Chang HY;Stadler HS
在E11小鼠胚胎中的基因表达谱鉴定了长非编码RNA（lncRNA）、LNCRNA-HIT在未分化的肢体间充质、肠道和发育中的生殖结节中的高表达。在肢体间充质中，发现LncRNA-HIT保留在细胞核中，与p100和CBP形成复合物。通过ChIRP-seq对LncRNA-HIT-p100/CBP复合物的全基因组分布的分析揭示了在小鼠基因组中的多个基因座处的LncRNA-HIT相关峰。通过LncRNA-HIT-p100/CBP复合物接触的基因的本体论分析表明这些位点在软骨形成分化中的主要作用。使用siRNA介导的LncRNA-HIT或p100转录物减少的功能分析显示，许多LncRNA-HIT相关基因座的表达显著降低。LncRNA-HIT siRNA治疗还影响肢体间充质形成软骨的能力，减少间充质细胞凝聚和软骨结节的形成。LncRNA-HIT siRNA处理通过降低H3 K27 ac或p100活性在机制上影响促软骨形成基因表达，证实LncRNA-HIT对于肢体间充质中的软骨形成分化是必需的。总之，这些发现揭示了在早期肢体发育过程中起作用的基本表观遗传机制，使用LncRNA-HIT及其相关蛋白质来促进多个基因的表达，这些基因的产物是软骨形成所必需的。骨骼生物学家研究的一个基本问题是再生疗法的发展，以取代受损伤或疾病影响的软骨组织，对于受骨关节炎影响的个体来说，占65岁以上所有成年人的近一半。迄今为止，还没有促进持续软骨再生的疗法，因为我们还不能概括指导细胞形成关节软骨所需的编程事件，而这些细胞不继续分化成骨。我们对软骨形成过程中发生的早期编程事件的分析导致了LncRNA-HIT的鉴定，LncRNA-HIT是一种长的非编码RNA，对于胚胎肢体间充质分化为软骨是必不可少的。对LncRNA-HIT在间充质中分布的全基因组分析显示，LncRNA-HIT与许多其产物促进软骨形成的基因之间存在强相关性。在缺乏LncRNA-HIT的情况下，这些软骨形成基因的表达严重降低，影响这些细胞分化成软骨。从机制上讲，LncRNA-HIT通过将p100和CBP募集到这些位点来调节这些促软骨形成基因，促进H3 K27 ac和转录激活。LncRNA-HIT似乎也存在于大多数脊椎动物物种中，这表明由该lncRNA调节的表观遗传程序可能代表许多物种用于促进软骨形成的基本机制。
Gene expression profiling in E 11 mouse embryos identified high expression of the long noncoding RNA (lncRNA), LNCRNA-HIT in the undifferentiated limb mesenchyme, gut, and developing genital tubercle. In the limb mesenchyme, LncRNA-HIT was found to be retained in the nucleus, forming a complex with p100 and CBP. Analysis of the genome-wide distribution of LncRNA-HIT-p100/CBP complexes by ChIRP-seq revealed LncRNA-HIT associated peaks at multiple loci in the murine genome. Ontological analysis of the genes contacted by LncRNA-HIT-p100/CBP complexes indicate a primary role for these loci in chondrogenic differentiation. Functional analysis using siRNA-mediated reductions in LncRNA-HIT or p100 transcripts revealed a significant decrease in expression of many of the LncRNA-HIT-associated loci. LncRNA-HIT siRNA treatments also impacted the ability of the limb mesenchyme to form cartilage, reducing mesenchymal cell condensation and the formation of cartilage nodules. Mechanistically the LncRNA-HIT siRNA treatments impacted pro-chondrogenic gene expression by reducing H3K27ac or p100 activity, confirming that LncRNA-HIT is essential for chondrogenic differentiation in the limb mesenchyme. Taken together, these findings reveal a fundamental epigenetic mechanism functioning during early limb development, using LncRNA-HIT and its associated proteins to promote the expression of multiple genes whose products are necessary for the formation of cartilage. A fundamental problem studied by skeletal biologists is the development of regenerative therapies to replace cartilage tissues impacted by injury or disease, which for individuals affected by osteoarthritis represents nearly half of all of all adults over the age of sixty five. To date, no therapies exist to promote sustained cartilage regeneration, as we have not been able to recapitulate the programming events necessary to instruct cells to form articular cartilage without these cells continuing to differentiate into bone. Our analysis of the early programming events occurring during cartilage formation led to the identification of LncRNA-HIT a long noncoding RNA that is essential for the differentiation of the embryonic limb mesenchyme into cartilage. A genome wide analysis of LncRNA-HIT’s distribution in the mesenchyme revealed strong association between LncRNA-HIT and numerous genes whose products facilitate cartilage formation. In the absence of LncRNA-HIT, the expression of these chondrogenic genes is severely reduced, impacting the differentiation of these cells into cartilage. Mechanistically, LncRNA-HIT regulates these pro-chondrogenic genes by recruiting p100 and CBP to these loci, facilitating H3K27ac and transcriptional activation. LncRNA-HIT also appears to be present in most vertebrate species, suggesting that the epigenetic program regulated by this lncRNA may represent a fundamental mechanism used by many species to promote cartilage formation.