Physiological function of HB-EGF : Study of the knock-in mice of the mutant form of HB-EGF

EOF家族生长因子通过膜锚定前体的蛋白水解处理从细胞表面释放，称为外膜结构域脱落；然而，体外结构域脱落在体内的生物学意义尚不清楚。肝素结合egf样生长因子（HB-EGF）也来源于膜锚定前体（proHB-EGF）。我们证明控制proHB-EGF的外域脱落是正常小鼠发育所必需的过程。在小鼠胚胎皮肤转染SHB-EGF和proHB-EGF实验中，外源过表达SHB-EGF导致胚胎表皮增生，而proHB-EGF表达未引起胚胎表皮异常。这表明，即使在过表达后，胚胎表皮也严格控制着proHB-EGF外畴脱落。接下来，我们通过靶向基因替换制备了携带编码跨膜结构域截断的proHB-EGF （HB4^ATM）基因的小鼠。这些小鼠产生可溶性形式的β - egf (SHB-EGF)，而不是prohb - egf。携带HB^ATM及其Fl杂合子的嵌合小鼠表现出严重的皮肤增生，加速了角质形成细胞的增殖和分化。我们还观察到心脏心室孔增生，大多数动物在胚胎或新生儿阶段死亡。这表明proHB-EGF外畴的蛋白水解过程在体内受到严格控制。我们的研究结果还表明，胞外结构域脱落是生长因子活性的重要翻译后控制。

The EOF family of growth factors is released from the cell surface by proteolytic processing of the membrane-anchored precursors, dubbed ectodomain shedding ; the biological significance of ectodomain shedding in vivo, however, remains unknown. Heparin-binding EGF-like growth factor (HB-EGF) is also derived from a membrane-anchored precursor (proHB-EGF).We demonstrate that the control of ectodomain shedding of proHB-EGF is a process essential for normal mouse development. In the transfection experiments of SHB-EGF and proHB-EGF into mouse embryonic skins, exogenous overexpression of SHB-EGF resulted in embryonic epidermal hyperplasia, while proHB-EGF expression did not cause any abnormalities in embryonic epidermis. These suggest that even following overexpression, proHB-EGF ectodomain shedding is strictly controlled in the embryonic epidermis.Next we prepared mice carrying a gene encoding transmembrane-domain-truncated proHB-EGF (HB4^ATM) by targeted gene replacement. These mice produce a soluble form offtB-EGF (SHB-EGF), instead ofproHB-EGF. Chimeric mice carrying HB^ATM and their Fl heterozygotes exhibit severe skin hyperplasia with accelerated proliferation and perturbed differentiation of keratinocytes. We also observed ventricular well hyperplasia in the heart, with most of the animals dying in either the embryonic or neonatal stages. These indicate that the proteolytic processing of proHB-EGF ectodomain is strictly controlled in vivo. Our results also indicate that ectodomain shedding is a vital post-translational control of growth factor activity.