Homeoproteins are a class of transcription factors sharing the unexpected property of intercellular trafficking that confers to homeoproteins a paracrine mode of action. Homeoprotein paracrine action participates in the control of patterning processes, including axonal guidance, brain plasticity and boundary formation. Internalization and secretion, the two steps of intercellular transfer, rely on unconventional mechanisms, but the cellular mechanisms at stake still need to be fully characterized. Thanks to the design of new quantitative and sensitive assays dedicated to the study of homeoprotein transfer within HeLa cells in culture, we demonstrate a core role of phosphatidylinositol (4,5)-bisphosphate (PIP₂) together with cholesterol in the translocation of the homeobox protein engrailed-2 (EN2) across the plasma membrane. By using drug and enzyme treatments, we show that both secretion and internalization are regulated according to PIP₂ levels. The requirement for PIP₂ and cholesterol in EN2 trafficking correlates with their selective affinity for this protein in artificial bilayers, which is drastically decreased in a paracrine-deficient mutant of EN2. We propose that the bidirectional plasma membrane translocation events that occur during homeoprotein secretion and internalization are parts of a common process.

同源蛋白是一类转录因子，具有意想不到的细胞间运输特性，赋予同源蛋白旁分泌作用模式。同源蛋白旁分泌作用参与模式形成过程的控制，包括轴突引导、大脑可塑性和边界形成。内化和分泌是细胞间转移的两个步骤，依赖于非常规机制，但仍需要充分表征所涉及的细胞机制。_{由于设计了专门用于研究培养物 HeLa 细胞内同源蛋白转移的新型定量和灵敏检测方法，我们证明了磷脂酰肌醇 (4,5)-二磷酸 (PIP 2} ) 与胆固醇在同源盒易位中的核心作用蛋白质 engrailed-2 (EN2) 穿过质膜。通过使用药物和酶治疗，我们发现分泌和内化均根据 PIP ₂水平进行调节。_{EN2 运输中对 PIP 2}和胆固醇的需求与其在人工双层中对该蛋白质的选择性亲和力相关，而在 EN2 旁分泌缺陷突变体中，这种选择性亲和力大大降低。我们认为，同源蛋白分泌和内化过程中发生的双向质膜易位事件是共同过程的一部分。