Lee Ting-Hein, McKleroy William, Khalifeh-Soltani Amin, Sakuma Stephen, Lazarev Stanislav, Riento Kirsi, Nishimura Stephen L, Nichols Ben J, Atabai Kamran
Cardiovascular Research Institute, University of California, San Francisco, San Francisco, CA 94158 Department of Medicine, University of California, San Francisco, San Francisco, CA 94158 Lung Biology Center, University of California, San Francisco, San Francisco, CA 94158 Department of Pathology, University of California, San Francisco, San Francisco, CA 94158 MRC Laboratory of Molecular Biology, Cambridge CB2 0QH, United Kingdom.
Mol Biol Cell. 2014 Mar;25(5):583-93. doi: 10.1091/mbc.E13-07-0382. Epub 2014 Jan 8.
Tissue fibrosis occurs when matrix production outpaces matrix degradation. Degradation of collagen, the main component of fibrotic tissue, is mediated through an extracellular proteolytic pathway and intracellular pathway of cellular uptake and lysosomal digestion. Recent studies demonstrate that disruption of the intracellular pathways can exacerbate fibrosis. These pathways are poorly characterized. Here we identify novel mediators of the intracellular pathway of collagen turnover through a genome-wide RNA interference screen in Drosophila S2 cells. Screening of 7505 Drosophila genes conserved among metazoans identified 22 genes that were required for efficient internalization of type I collagen. These included proteins involved in vesicle transport, the actin cytoskeleton, and signal transduction. We show further that the flotillin genes have a conserved and central role in collagen uptake in Drosophila and human cells. Short hairpin RNA-mediated silencing of flotillins in human monocyte and fibroblasts impaired collagen uptake by promoting lysosomal degradation of the endocytic collagen receptors uPARAP/Endo180 and mannose receptor. These data provide an initial characterization of intracellular pathways of collagen turnover and identify the flotillin genes as critical regulators of this process. A better understanding of these pathways may lead to novel therapies that reduce fibrosis by increasing collagen turnover.
当基质产生超过基质降解时，组织纤维化就会发生。纤维化组织的主要成分胶原蛋白的降解是通过细胞外蛋白水解途径以及细胞摄取和溶酶体消化的细胞内途径介导的。最近的研究表明，细胞内途径的破坏会加剧纤维化。这些途径的特征尚不明确。在这里，我们通过在果蝇S2细胞中进行全基因组RNA干扰筛选，确定了胶原蛋白周转细胞内途径的新型介质。对后生动物中保守的7505个果蝇基因进行筛选，确定了22个基因是I型胶原蛋白有效内化所必需的。这些基因包括参与囊泡运输、肌动蛋白细胞骨架和信号转导的蛋白质。我们进一步表明，浮舰蛋白基因在果蝇和人类细胞的胶原蛋白摄取中具有保守且核心的作用。短发夹RNA介导的人类单核细胞和成纤维细胞中浮舰蛋白的沉默，通过促进内吞胶原蛋白受体uPARAP/Endo180和甘露糖受体的溶酶体降解，损害了胶原蛋白的摄取。这些数据提供了胶原蛋白周转细胞内途径的初步特征，并确定浮舰蛋白基因是这一过程的关键调节因子。对这些途径的更好理解可能会带来通过增加胶原蛋白周转来减少纤维化的新疗法。
