DNA Demethylation and Muller Glia Reprogramming During Retina Regeneration

描述(申请人提供)：与哺乳动物不同，斑马鱼能够再生受损的视网膜，恢复失明的视力。这种再生能力依赖于Muller glia(MG)，MG在去分化、增殖、最终分化以再生新的神经元和胶质细胞的过程中，通过经历身份的多次转变来应对视网膜损伤。虽然MG可以被诱使在受损的哺乳动物视网膜中增殖，但它们并不具有多能性，而且很少能再生受损的神经元。因此，了解推动斑马鱼MG重编程为多能的机制可能会为从哺乳动物MG中产生多能祖细胞提供新的策略。最近的研究表明，MG的重新编程伴随着基因表达程序的激活，这些程序类似于作用于胚胎干细胞和视网膜前体细胞的程序。我们假设，驱动MG去分化和多能性的遗传程序是由DNA甲基化控制的。在动物中，DNA甲基化主要发生在CpG二核苷酸上，并控制转录调控过程，如印迹、X染色体失活、转座子沉默和基因活性的稳定沉默。基因编码区附近DNA甲基化与基因沉默相关。重要的是，DNA甲基化的变化与基因表达程序的激活和抑制有关，这些程序发生在早期发育过程中，并伴随着体细胞的重新编程来产生诱导的多能干细胞。很可能基因组甲基化的删除和重建，在关键位置， 伴随着基因表达的变化，驱动MG去分化和多能性，并随后再生新的视网膜细胞类型。在这里，我们建议识别MG基因组中在视网膜再生过程中经历甲基化变化的区域，并确定这些变化是否与以前使用微阵列技术表征的基因表达变化相关。此外，我们还提出了一种假设，即DNA去甲基化在MG的去分化过程中是一个由Apobec2a和2b蛋白驱动的活跃过程。

DESCRIPTION (provided by applicant): Unlike mammals, zebrafish are able to regenerate a damaged retina and restore lost sight. This regenerative ability depends on Muller glia (MG) that respond to retinal injury by undergoing multiple shifts in identity as they dedifferentiate, proliferate, and finally differentiate to regenerate new neurons and glia. Although MG can be coaxed to proliferate in the injured mammalian retina, they do not exhibit multipotency and only rarely regenerate damaged neurons. Therefore understanding the mechanisms driving zebrafish MG reprogramming to mutlipotency may suggest novel strategies for generating multipotent progenitors from mammalian MG. Recent studies suggest that MG reprogramming is accompanied by activation of gene expression programs that are similar to those acting in embryonic stem cells and retinal progenitors. We hypothesize that genetic programs driving MG dedifferentiation and multipotency are controlled by DNA methylation. In animals, DNA methylation predominantly occurs at CpG dinucleotides and controls transcriptional regulatory processes like imprinting, X-chromosome inactivation, transposon silencing, and stable silencing of gene activity. Methylation of DNA proximal to gene-coding regions is correlated with gene silencing. Importantly, changes in DNA methylation have been correlated with the activation and suppression of gene expression programs that take place during early development and accompany the reprogramming of somatic cells to yield induced pluripotent stem cells. It is likely that erasure and reestablishment of genomic methylation, at key locations, accompanies the gene expression changes that drive MG dedifferentiation and multipotency and subsequently the regeneration of new retinal cell types. Here we propose to identify regions of the MG genome that are undergoing methylation changes during retina regeneration and determine if these changes correlate with gene expression changes that have previously been characterized using microarray technology. In addition, we propose to test the hypothesis that DNA demethylation in dedifferentiating MG is an active process driven by Apobec2a and 2b proteins.