Pokidysheva Elena N, Tufa Sara F, Keene Douglas R, Hudson Billy G, Boudko Sergei P
Division of Nephrology and Hypertension, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA; Center for Matrix Biology, Vanderbilt University Medical Center, Nashville, Tennessee, USA; Aspirnaut Program, Vanderbilt University Medical Center, Nashville, Tennessee, USA.
Micro-Imaging Center, Shriners Children's, Portland, Oregon, USA.
J Biol Chem. 2025 Jun 14;301(7):110384. doi: 10.1016/j.jbc.2025.110384.
The collagen IV scaffold serves as a fundamental structural unit of the basement membrane (BM). Understanding its structure, assembly, and function is essential for tissue engineering, the design of organoid models, and developing therapies for diseases such as Alport syndrome, Gould syndrome, psoriasis, eye abnormalities, hearing loss, and others, where collagen IV is required for structural integrity and functionality of the BM. The collagen IV molecule is a 400 nm long heterotrimer, comprising non-collagenous 1 (NC1), collagenous, and 7S domains. The assembly of the collagen IV scaffold involves oligomerization of the C-terminal NC1 and the N-terminal 7S domains, along with lateral associations within the collagenous domain. However, the detailed architecture and assembly mechanisms of the collagen IV scaffold remain unclear. Here, we investigated the potency and mechanism of recombinant single-chain NC1 trimer incorporation into the collagen IV scaffold. We discovered that the NC1 trimer influences the overall assembly of the basement membrane by affecting the quality of the developing collagen IV scaffold in a dose-dependent manner, without impacting already established scaffolds. This interference occurs through the hexamerization of supplemented NC1 trimers with endogenous NC1 domains, as the NC1 trimer becomes sulfilimine crosslinked with the existing chains. Overall, the single-chain NC1 trimer of collagen IV is crucial for developing novel extracellular therapies in two main ways: (1) facilitating the delivery and incorporation of functional replacements like collagen IV fragments and (2) inhibiting the formation of new basement membranes in conditions such as tumor growth and detrimental vascularization.
IV型胶原蛋白支架是基底膜（BM）的基本结构单元。了解其结构、组装和功能对于组织工程、类器官模型的设计以及开发针对诸如阿尔波特综合征、古尔德综合征、银屑病、眼部异常、听力损失等疾病的治疗方法至关重要，在这些疾病中，IV型胶原蛋白对于基底膜的结构完整性和功能是必需的。IV型胶原蛋白分子是一种400纳米长的异源三聚体，由非胶原蛋白1（NC1）、胶原蛋白和7S结构域组成。IV型胶原蛋白支架的组装涉及C端NC1和N端7S结构域的寡聚化，以及胶原蛋白结构域内的横向关联。然而，IV型胶原蛋白支架的详细结构和组装机制仍不清楚。在这里，我们研究了重组单链NC1三聚体掺入IV型胶原蛋白支架的效力和机制。我们发现，NC1三聚体通过以剂量依赖的方式影响正在形成的IV型胶原蛋白支架的质量，从而影响基底膜的整体组装，而不会影响已经建立的支架。这种干扰是通过补充的NC1三聚体与内源性NC1结构域的六聚化发生的，因为NC1三聚体与现有链形成亚磺酰亚胺交联。总体而言，IV型胶原蛋白的单链NC1三聚体以两种主要方式对开发新型细胞外疗法至关重要：（1）促进诸如IV型胶原蛋白片段等功能性替代物的递送和掺入；（2）在肿瘤生长和有害血管生成等情况下抑制新基底膜的形成。
