Department of Molecular and Cellular Neuroscience, Dorris Neuroscience Center, The Scripps Research Institute, La Jolla, California, USA.
Department of Cell and Molecular Biology, The Scripps Research Institute, La Jolla, California, USA.
Nat Biotechnol. 2017 Oct;35(10):960-968. doi: 10.1038/nbt.3963. Epub 2017 Sep 11.
The reprogramming of differentiated cells into induced pluripotent stem cells (iPSCs) is usually achieved by exogenous induction of transcription by factors acting in the nucleus. In contrast, during development, signaling pathways initiated at the membrane induce differentiation. The central idea of this study is to identify antibodies that can catalyze cellular de-differentiation and nuclear reprogramming by acting at the cell surface. We screen a lentiviral library encoding ∼100 million secreted and membrane-bound single-chain antibodies and identify antibodies that can replace either Sox2 and Myc (c-Myc) or Oct4 during reprogramming of mouse embryonic fibroblasts into iPSCs. We show that one Sox2-replacing antibody antagonizes the membrane-associated protein Basp1, thereby de-repressing nuclear factors WT1, Esrrb and Lin28a (Lin28) independent of Sox2. By manipulating this pathway, we identify three methods to generate iPSCs. Our results establish unbiased selection from autocrine combinatorial antibody libraries as a robust method to discover new biologics and uncover membrane-to-nucleus signaling pathways that regulate pluripotency and cell fate.
将分化细胞重编程为诱导多能干细胞（iPSCs）通常通过核内作用的转录因子的外源诱导来实现。相比之下，在发育过程中，起始于细胞膜的信号通路诱导分化。本研究的核心思想是鉴定能够通过作用于细胞表面来催化细胞去分化和核重编程的抗体。我们筛选了一个编码约 1 亿个分泌型和膜结合型单链抗体的慢病毒文库，并鉴定出能够在将小鼠胚胎成纤维细胞重编程为 iPSCs 的过程中替代 Sox2 和 Myc（c-Myc）或 Oct4 的抗体。我们表明，一种替代 Sox2 的抗体拮抗膜相关蛋白 Basp1，从而使核因子 WT1、Esrrb 和 Lin28a（Lin28）去抑制，而不依赖 Sox2。通过操纵这条通路，我们确定了三种生成 iPSCs 的方法。我们的结果确立了从自分泌组合抗体文库中进行无偏选择作为发现新生物制剂和揭示调节多能性和细胞命运的膜到核信号通路的有效方法。
