Peroxisomes are ubiquitous organelles formed by peroxisome biogenesis (PB). During PB, peroxisomal matrix proteins harboring a peroxisome targeting signal (PTS) are imported inside peroxisomes by peroxins, encoded by PEX genes. Genetic alterations in PEX genes lead to a spectrum of incurable diseases called Zellweger spectrum disorders (ZSD). In vitro drug screening is part of the quest for a cure in ZSD by restoring PB in ZSD cell models. In vitro PB evaluation is commonly achieved by immunofluorescent staining or transient peroxisome fluorescent reporter expression. Both techniques have several drawbacks (cost, time-consuming technique, etc.) which we overcame by developing a third-generation lentiviral transfer plasmid expressing an enhanced green fluorescent protein fused to PTS1 (eGFP-PTS1). By eGFP-PTS1 lentiviral transduction, we quantified PB and peroxisome motility in ZSD and control mouse and human fibroblasts. We confirmed the stable eGFP-PTS1 expression along cell passages. eGFP signal analysis distinguished ZSD from control eGFP-PTS1-transduced cells. Live eGFP-PTS1 transduced cells imaging quantified peroxisomes motility. In conclusion, we developed a lentiviral transfer plasmid allowing stable eGFP-PTS1 expression to study PB (deposited on Addgene: #133282). This tool meets the needs for in vitro PB evaluation and ZSD drug discovery.

中文翻译：

---

通过慢病毒表达与过氧化物酶体靶向信号1融合的绿色荧光蛋白可实现准确而实时的过氧化物酶体生物发生评估。

过氧化物酶体是由过氧化物酶体生物发生（PB）形成的普遍存在的细胞器。在PB期间，过氧化物酶体靶向信号（PTS）的过氧化物酶体基质蛋白通过PEX基因编码的过氧化物被导入过氧化物酶体内部。PEX基因的遗传改变会导致一系列无法​​治愈的疾病，称为Zellweger谱系疾病（ZSD）。体外药物筛选是通过恢复ZSD细胞模型中的PB来治愈ZSD的部分任务。体外PB评估通常通过免疫荧光染色或瞬时过氧化物酶体荧光报告基因表达来实现。两种技术都有一些缺点（成本，耗时的技术等），我们通过开发第三代慢病毒转移质粒克服了这些缺点，该质粒表达与PTS1（eGFP-PTS1）融合的增强型绿色荧光蛋白。通过eGFP-PTS1慢病毒转导，我们量化了ZSD以及对照小鼠和人类成纤维细胞中的PB和过氧化物酶活力。我们证实了沿细胞传代的稳定的eGFP-PTS1表达。eGFP信号分析将ZSD与对照eGFP-PTS1转导的细胞区分开。实时eGFP-PTS1转导的细胞对定量的过氧化物酶体运动性成像。总之，我们开发了一种慢病毒转移质粒，可以稳定地表达eGFP-PTS1来研究PB（存放在Addgene：＃133282上）。该工具可满足体外PB评估和ZSD药物发现的需求。我们开发了一种慢病毒转移质粒，可以稳定地表达eGFP-PTS1来研究PB（存放在Addgene：＃133282上）。该工具可满足体外PB评估和ZSD药物发现的需求。我们开发了一种慢病毒转移质粒，可以稳定地表达eGFP-PTS1来研究PB（存放在Addgene：＃133282上）。该工具可满足体外PB评估和ZSD药物发现的需求。