Oral and Craniofacial Health Sciences, School of Dentistry, University of North Carolina, Chapel Hill, North Carolina, United States of America.
Nippi Research Institute of Biomatrix, Toride, Ibaraki, Japan.
PLoS Genet. 2019 Jun 7;15(6):e1008196. doi: 10.1371/journal.pgen.1008196. eCollection 2019 Jun.
Covalent intermolecular cross-linking of collagen is essential for tissue stability. Recent studies have demonstrated that cyclophilin B (CypB), an endoplasmic reticulum (ER)-resident peptidyl-prolyl cis-trans isomerase, modulates lysine (Lys) hydroxylation of type I collagen impacting cross-linking chemistry. However, the extent of modulation, the molecular mechanism and the functional outcome in tissues are not well understood. Here, we report that, in CypB null (KO) mouse skin, two unusual collagen cross-links lacking Lys hydroxylation are formed while neither was detected in wild type (WT) or heterozygous (Het) mice. Mass spectrometric analysis of type I collagen showed that none of the telopeptidyl Lys was hydroxylated in KO or WT/Het mice. Hydroxylation of the helical cross-linking Lys residues was almost complete in WT/Het but was markedly diminished in KO. Lys hydroxylation at other sites was also lower in KO but to a lesser extent. A key glycosylation site, α1(I) Lys-87, was underglycosylated while other sites were mostly overglycosylated in KO. Despite these findings, lysyl hydroxylases and glycosyltransferase 25 domain 1 levels were significantly higher in KO than WT/Het. However, the components of ER chaperone complex that positively or negatively regulates lysyl hydroxylase activities were severely reduced or slightly increased, respectively, in KO. The atomic force microscopy-based nanoindentation modulus were significantly lower in KO skin than WT. These data demonstrate that CypB deficiency profoundly affects Lys post-translational modifications of collagen likely by modulating LH chaperone complexes. Together, our study underscores the critical role of CypB in Lys modifications of collagen, cross-linking and mechanical properties of skin.
胶原蛋白的共价分子间交联对于组织稳定性至关重要。最近的研究表明，内质网（ER）驻留的肽基脯氨酰顺反异构酶 cyclophilin B（CypB）调节 I 型胶原蛋白赖氨酸（Lys）羟化，从而影响交联化学。然而，其调节程度、分子机制以及在组织中的功能结果尚不清楚。在这里，我们报告在 CypB 敲除（KO）小鼠皮肤中，形成了两种缺乏 Lys 羟化的异常胶原交联，而在野生型（WT）或杂合型（Het）小鼠中均未检测到。I 型胶原蛋白的质谱分析表明，KO 或 WT/Het 小鼠的胶原肽末端 Lys 均未羟化。WT/Het 的螺旋交联 Lys 残基羟化几乎完全，但 KO 明显减少。KO 中的其他 Lys 羟化程度也较低，但程度较轻。一个关键的糖基化位点，α1(I) Lys-87，发生低聚糖基化，而 KO 中的其他位点主要发生高聚糖基化。尽管存在这些发现，但 KO 中的赖氨酰羟化酶和糖基转移酶 25 结构域 1 水平明显高于 WT/Het。然而，正向或负向调节赖氨酰羟化酶活性的 ER 伴侣复合物的组成部分在 KO 中分别严重减少或轻微增加。KO 皮肤的原子力显微镜基于纳米压痕模量明显低于 WT。这些数据表明 CypB 缺乏会深刻影响胶原蛋白的 Lys 翻译后修饰，可能通过调节 LH 伴侣复合物来实现。总的来说，我们的研究强调了 CypB 在胶原蛋白 Lys 修饰、交联和皮肤机械性能中的关键作用。
