Department of Molecular and Translational Medicine, Division of Biology and Genetics, University of Brescia, Brescia, Italy.
PLoS One. 2019 Feb 4;14(2):e0211647. doi: 10.1371/journal.pone.0211647. eCollection 2019.
Classical Ehlers-Danlos syndrome (cEDS) is a dominant inherited connective tissue disorder mainly caused by mutations in the COL5A1 and COL5A2 genes encoding type V collagen (COLLV), which is a fibrillar COLL widely distributed in a variety of connective tissues. cEDS patients suffer from skin hyperextensibility, abnormal wound healing/atrophic scars, and joint hypermobility. Most of the causative variants result in a non-functional COL5A1 allele and COLLV haploinsufficiency, whilst COL5A2 mutations affect its structural integrity. To shed light into disease mechanisms involved in cEDS, we performed gene expression profiling in skin fibroblasts from four patients harboring haploinsufficient and structural mutations in both disease genes. Transcriptome profiling revealed significant changes in the expression levels of different extracellular matrix (ECM)-related genes, such as SPP1, POSTN, EDIL3, IGFBP2, and C3, which encode both matricellular and soluble proteins that are mainly involved in cell proliferation and migration, and cutaneous wound healing. These gene expression changes are consistent with our previous protein findings on in vitro fibroblasts from other cEDS patients, which exhibited reduced migration and poor wound repair owing to COLLV disorganization, altered deposition of fibronectin into ECM, and an abnormal integrin pattern. Microarray analysis also indicated the decreased expression of DNAJB7, VIPAS39, CCPG1, ATG10, SVIP, which encode molecular chaperones facilitating protein folding, enzymes regulating post-Golgi COLLs processing, and proteins acting as cargo receptors required for endoplasmic reticulum (ER) proteostasis and implicated in the autophagy process. Patients' cells also showed altered mRNA levels of many cell cycle regulating genes including CCNE2, KIF4A, MKI67, DTL, and DDIAS. Protein studies showed that aberrant COLLV expression causes the disassembly of itself and many structural ECM constituents including COLLI, COLLIII, fibronectin, and fibrillins. Our findings provide the first molecular evidence of significant gene expression changes in cEDS skin fibroblasts highlighting that defective ECM remodeling, ER homeostasis and autophagy might play a role in the pathogenesis of this connective tissue disorder.
经典型埃勒斯-当洛斯综合征（cEDS）是一种显性遗传性结缔组织疾病，主要由编码 V 型胶原（COLLV）的 COL5A1 和 COL5A2 基因突变引起，COLLV 是一种广泛分布于多种结缔组织中的纤维状 COLL。cEDS 患者表现为皮肤过度伸展、伤口愈合异常/萎缩性瘢痕和关节过度活动。大多数致病变异导致 COL5A1 等位基因无功能和 COLLV 单倍不足，而 COL5A2 突变则影响其结构完整性。为了深入了解 cEDS 相关的疾病机制，我们对来自四个患者的皮肤成纤维细胞进行了基因表达谱分析，这些患者在两个疾病基因中均存在单倍不足和结构突变。转录组谱分析显示，不同细胞外基质（ECM）相关基因的表达水平发生了显著变化，如 SPP1、POSTN、EDIL3、IGFBP2 和 C3，这些基因编码的基质和可溶性蛋白主要参与细胞增殖和迁移以及皮肤伤口愈合。这些基因表达的变化与我们之前在来自其他 cEDS 患者的体外成纤维细胞中的蛋白发现结果一致，这些细胞由于 COLLV 结构紊乱、纤连蛋白在 ECM 中的沉积改变以及整合素模式异常，表现出迁移减少和伤口修复不良。微阵列分析还表明，DNAJB7、VIPAS39、CCPG1、ATG10、SVIP 的表达减少，这些基因编码的分子伴侣有助于蛋白质折叠、调节高尔基体后 COLL 加工的酶，以及作为内质网（ER）蛋白稳态所必需的货物受体并参与自噬过程的蛋白质。患者的细胞还显示出许多细胞周期调节基因的 mRNA 水平改变，包括 CCNE2、KIF4A、MKI67、DTL 和 DDIAS。蛋白质研究表明，异常的 COLLV 表达导致其自身和许多结构 ECM 成分（包括 COLLI、COLLIII、纤连蛋白和原纤维蛋白）的解体。我们的研究结果提供了 cEDS 皮肤成纤维细胞中显著基因表达变化的首个分子证据，突出表明 ECM 重塑、ER 稳态和自噬异常可能在这种结缔组织疾病的发病机制中发挥作用。
