Collagen type IV (Col IV) is a basement membrane protein associated with early blood vessel morphogenesis and is essential for blood vessel stability. Defects in vascular Col IV deposition are the basis of heritable disorders, such as small vessel disease, marked by cerebral hemorrhage and drastically shorten lifespan. To date, little is known about how endothelial cells regulate the intracellular transport and selective secretion of Col IV in response to angiogenic cues, leaving a void in our understanding of this critical process. Our aim was to identify trafficking pathways that regulate Col IV deposition during angiogenic blood vessel development. We have identified the GTPase Rab10 as a major regulator of Col IV vesicular trafficking during vascular development using both in vitro imaging and biochemistry as well as in vivo models. Knockdown of Rab10 reduced de novo Col IV secretion in vivo and in vitro. Mechanistically, we determined that Rab10 is an indirect mediator of Col IV secretion, partnering with atypical Rab25 to deliver the enzyme lysyl hydroxylase 3 (LH3) to Col IV-containing vesicles staged for secretion. Loss of Rab10 or Rab25 results in depletion of LH3 from Col IV-containing vesicles and rapid lysosomal degradation of Col IV. Furthermore, we demonstrate that Rab10 is Notch responsive, indicating a novel connection between permissive Notch-based vessel maturation programs and vesicle trafficking. Our results illustrate both a new trafficking-based component in the regulated secretion of Col IV and how this vesicle trafficking program interfaces with Notch signaling to fine-tune basement membrane secretion during blood vessel development.

IV 型胶原 (Col IV) 是一种与早期血管形态发生相关的基底膜蛋白，对于血管稳定性至关重要。血管 Col IV 沉积缺陷是遗传性疾病的基础，例如以脑出血为特征的小血管疾病，并大大缩短寿命。迄今为止，人们对内皮细胞如何响应血管生成信号调节细胞内运输和选择性分泌 Col IV 知之甚少，这使得我们对这一关键过程的理解存在空白。我们的目的是确定在血管生成过程中调节 Col IV 沉积的运输途径。我们利用体外成像、生物化学以及体内模型，确定 GTPase Rab10 是血管发育过程中 Col IV 囊泡运输的主要调节因子。Rab10 的敲除减少了体内和体外 Col IV 的从头分泌。从机制上讲，我们确定 Rab10 是 Col IV 分泌的间接介导物，与非典型 Rab25 合作，将赖氨酰羟化酶 3 (LH3) 传递至处于分泌阶段的含有 Col IV 的囊泡。Rab10 或 Rab25 的缺失导致含有 Col IV 的囊泡中 LH3 的消耗以及 Col IV 的快速溶酶体降解。此外，我们证明 Rab10 对 Notch 具有反应性，表明基于 Notch 的许可性血管成熟程序与囊泡运输之间存在新的联系。我们的结果说明了 Col IV 调节分泌中基于运输的新成分，以及该囊泡运输程序如何与 Notch 信号相互作用以微调血管发育过程中的基底膜分泌。