Deciphering the Role of Pumilio1 in Two New Neurological Diseases

解读 Pumilio1 在两种新神经系统疾病中的作用

• 批准号: 10605223
• 负责人: Vincenzo Alessandro Gennarino
• 金额: $ 43.48万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-15 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10605223
• 关键词: Abnormal coordination; Adult; Age; Ataxia; Brain; Brain region; Candidate Disease Gene; Cell Line; Cerebellar Ataxia; Cerebellar degeneration; Cerebellum; Clinical; Cognitive; Complex; Data; Development; Disease; Evaluation; FMRP; Family; Future; Gene Expression; Heterozygote; Hippocampus; Homologous Gene; Human; Knock-out; Knockout Mice; Maps; MicroRNAs; Modeling; Molecular; Motor; Mus; Mutant Strains Mice; Mutation; Nervous System Physiology; Neurons; Pathway Analysis; Pathway interactions; Patients; Phenotype; Post-Transcriptional Regulation; Proteins; Publishing; Purkinje Cells; RNA-Binding Proteins; Role; Seizures; Substantia nigra structure; Testing; behavioral study; bioinformatics tool; crosslinking and immunoprecipitation sequencing; developmental disease; early onset; early onset disorder; falls; gain of function; in vivo; insight; interest; loss of function; nervous system disorder; neuropsychiatric symptom; transcriptome sequencing; validation studies

Little is known about the role of RNA-binding proteins in brain development or disease, but accumulating evidence indicates their involvement in neurological disorders. In fact, we found that the RNA-binding protein Pum1 is crucial for neurological function in both mice and humans. Pum1-haploinsufficient mice develop ataxia at 5 weeks of age and show Purkinje cell degeneration at 10 weeks. Pum1 knockout mice are sicker: they are born at a lower mendelian ratio, are smaller than wild-type, have early seizures, and more severe cerebellar degeneration. We then found that mutations in human PUM1 also cause two very different diseases that parallel what we observed in mice: a mild, adult-onset pure cerebellar ataxia in patients with a mutation that reduces PUM1 levels by ~25%, and an early-onset disorder that causes several cognitive and physical delays, smaller size, motor incoordination, and seizures in patients with PUM1 mutations that reduce its levels ~40-60%. But how do the specific mutations alter PUM1 function, aside from making it less stable? The most obvious place to look is at PUM1 targets. The only published neuronal targets are ATXN1 and E2F3, and their abundance is increased by similar amounts (~50%) in both the adult-onset ataxia and early-onset cases. The ataxia might be explained by elevated abundance of cerebellar ATXN1, but the broader phenotype of the developmental disorder must involve dysregulation of other PUM1 targets. There is, however, more of a puzzle here than is first apparent. The mildest mutation, T1035S, which reduces PUM1 levels by only 25%, is in homology domain (HD); of the mutations that produce the severe, early onset phenotype, R1139W is in HD8, and R1147W is just outside this domain. Why, then, do R1139W and R1147W produce equally severe phenotypes, when only the former abolishes PUM1's repressor activity? And why is T1035S so mild, when it also abolishes PUM1's repressor activity? We propose that the milder disease results from target dysregulation, whereas more severe disease results when levels of PUM1 fall below a certain point (perhaps 30-40%), because its interacting partners either cannot form their normal complexes or the complexes fall apart quickly, causing loss of function of those interactors (and loss or gain of function of downstream targets). To test this two-part hypothesis, we will: 1) map the Pum1 targetome in the mouse brain as well as that of Pum2, its homolog (there may be regulatory overlap between the two proteins); 2) identify PUM1 protein interactors, and 3) study the cross-talk between Pum1 and Pum2 in mice. In sum, our recent discoveries not only define two new neurological diseases, they demonstrate that understanding the post- transcriptional regulation of disease-related proteins, like the PUF family, can lead to the identification of new candidate disease genes.

人们对rna结合蛋白在大脑发育或疾病中的作用知之甚少，但越来越多的证据表明它们与神经系统疾病有关。事实上，我们发现rna结合蛋白Pum1对小鼠和人类的神经功能都至关重要。pum1 -单倍体不足的小鼠在5周龄时出现共济失调，在10周龄时出现浦肯野细胞变性。Pum1基因敲除小鼠的病情更严重：它们出生时的孟德尔比率更低，体型比野生型更小，有早期癫痫发作，以及更严重的小脑退化。然后，我们发现人类PUM1的突变也会导致两种非常不同的疾病，与我们在小鼠中观察到的相似：一种是轻度的，成年发作的纯小脑性共济失调，患者的PUM1突变使其水平降低了25%；另一种是早发性疾病，导致PUM1突变使其水平降低了40-60%，患者的认知和身体发育迟缓，体型变小，运动不协调和癫痫发作。但是，除了降低PUM1的稳定性之外，这些特定的突变是如何改变其功能的呢？最明显的地方是PUM1目标。唯一公布的神经元靶点是ATXN1和E2F3，它们的丰度在成人发病的共济失调和早期发病的病例中都增加了相似的量（约50%）。共济失调可能由小脑ATXN1丰度升高来解释，但发育障碍的更广泛表型必须涉及其他PUM1靶点的失调。然而，这里有一个比最初看起来更令人困惑的问题。最温和的突变T1035S，使PUM1水平仅降低25%，位于同源结构域（HD）；在产生严重早发表型的突变中，R1139W位于HD8中，而R1147W恰好位于该结构域之外。那么，为什么R1139W和R1147W产生同样严重的表型，而只有前者能消除PUM1的抑制活性？为什么T1035S如此温和，而它却能消除PUM1的抑制活性？我们认为，较轻的疾病是由靶标失调引起的，而当PUM1水平低于某一点（可能是30-40%）时，更严重的疾病就会发生，因为PUM1的相互作用伙伴要么不能形成正常的复合物，要么复合物迅速分解，导致这些相互作用体的功能丧失（以及下游靶标的功能丧失或获得）。为了验证这个由两部分组成的假设，我们将：1)绘制小鼠大脑中的Pum1靶组及其同源物Pum2靶组（这两种蛋白之间可能存在调控重叠）；2)鉴定PUM1蛋白相互作用因子；3)研究小鼠PUM1与Pum2的串扰。总之，我们最近的发现不仅定义了两种新的神经系统疾病，而且表明了解疾病相关蛋白的转录后调控，如PUF家族，可以导致新的候选疾病基因的鉴定。