Nagai Masayoshi, Porter Robert S, Miyasato Maxwell, Wang Aijia, Gavilan Cecilia M, Hughes Elizabeth D, Wu Michael C, Saunders Thomas L, Iwase Shigeki
Department of Human Genetics, University of Michigan, Ann Arbor, Michigan, USA.
Genetics & Genomics Graduate Program, University of Michigan, Ann Arbor, Michigan, USA.
J Biol Chem. 2024 Nov;300(11):107881. doi: 10.1016/j.jbc.2024.107881. Epub 2024 Oct 11.
PHF21A is a histone-binding protein that recognizes unmethylated histone H3K4, the reaction product of LSD1 histone demethylase. PHF21A and LSD1 form a complex, and both undergo neuron-specific microexon splicing. The PHF21A neuronal microexon interferes with nucleosome binding, whereas the LSD1 neuronal microexon weakens H3K4 demethylation activity and can alter the substrate specificity to H3K9 or H4K20. However, the temporal expression patterns of PHF21A and LSD1 splicing isoforms during brain development and their biological roles remain unknown. In this work, we report that neuronal PHF21A isoform expression precedes neuronal LSD1 expression during human neuron differentiation and mouse brain development. The asynchronous splicing events resulted in stepwise deactivation of the LSD1-PHF21A complex in reversing H3K4 methylation. An unbiased proteomics survey revealed that the enzymatically inactive LSD1-PHF21A complex interacts with neuron-specific binding partners, including MYT1-family transcription factors and post-transcriptional mRNA processing proteins such as VIRMA. The interaction with the neuron-specific components, however, did not require the PHF21A microexon, indicating that the neuronal proteomic milieu, rather than the microexon-encoded PHF21A segment, is responsible for neuron-specific complex formation. Finally, by using two Phf21a mutant mouse models, we found that Phf21a neuronal splicing prevents excess synapse formation that otherwise would occur when canonical PHF21A is expressed in neurons. These results suggest that the role of the PHF21A microexon is to dampen LSD1-mediated H3K4 demethylation, thereby containing aberrant synaptogenesis.
PHF21A是一种组蛋白结合蛋白，可识别未甲基化的组蛋白H3K4，即LSD1组蛋白去甲基化酶的反应产物。PHF21A和LSD1形成复合物，二者均经历神经元特异性微小外显子剪接。PHF21A神经元微小外显子会干扰核小体结合，而LSD1神经元微小外显子会削弱H3K4去甲基化活性，并可改变对H3K9或H4K20的底物特异性。然而，PHF21A和LSD1剪接异构体在大脑发育过程中的时间表达模式及其生物学作用仍不清楚。在这项研究中，我们报告在人类神经元分化和小鼠大脑发育过程中，神经元PHF21A异构体的表达先于神经元LSD1的表达。异步剪接事件导致LSD1-PHF21A复合物在逆转H3K4甲基化过程中逐步失活。一项无偏向性蛋白质组学调查显示，无酶活性的LSD1-PHF21A复合物与神经元特异性结合伙伴相互作用，包括MYT1家族转录因子和转录后mRNA加工蛋白（如VIRMA）。然而，与神经元特异性成分的相互作用并不需要PHF21A微小外显子，这表明神经元蛋白质组环境而非微小外显子编码的PHF21A片段负责神经元特异性复合物的形成。最后，通过使用两种Phf21a突变小鼠模型，我们发现Phf21a神经元剪接可防止过量突触形成，否则当在神经元中表达典型PHF21A时就会出现这种情况。这些结果表明，PHF21A微小外显子的作用是抑制LSD1介导的H3K4去甲基化，从而抑制异常的突触发生。
