Rahman Abir A, Soto-Avellaneda Alejandro, Yong Jin Hyun, Stojkovska Iva, Lai Nathan K, Albright Joshua E, Webb Abby R, Oe Emily, Valarde Jacob P, Oxford Alexandra E, Urquhart Paige E, Wagner Brandon, Brown Connor, Amado Isabella, Vasquez Peyton, Lehning Nicholas, Grozdanov Veselin, Pu Xinzhu, Danzer Karin M, Morrison Brad E
Department of Biological Sciences, Boise State University, Boise, ID 83725, USA; Biomolecular Ph.D. Program, Boise State University, Boise, ID 83725, USA.
Department of Urology, School of Medicine, University of California, San Francisco, San Francisco, CA 94158, USA.
Neuroscience. 2020 Aug 10;441:33-45. doi: 10.1016/j.neuroscience.2020.06.009. Epub 2020 Jun 13.
The motor features of Parkinson's disease (PD) result from the loss of dopaminergic (DA) neurons in the substantia nigra with autophagy dysfunction being closely linked to this disease. A PD-causing familial mutation in VPS35 (D620N) has been reported to inhibit autophagy. In order to identify signaling pathways responsible for this autophagy defect, we performed an unbiased screen using RNA sequencing (RNA-Seq) of wild-type or VPS35 D620N-expressing retinoic acid-differentiated SH-SY5Y cells. We report that VPS35 D620N-expressing cells exhibit transcriptome changes indicative of alterations in extracellular matrix (ECM)-receptor interaction as well as PI3K-AKT signaling, a pathway known to regulate autophagy. Hyaluronan (HA) is a major component of brain ECM and signals via the ECM receptors CD44, a top RNA-Seq hit, and HA-mediated motility receptor (HMMR) to the autophagy-regulating PI3K-AKT pathway. We find that high (>950 kDa), but not low (15-40 kDa), molecular weight HA treatment inhibits autophagy. In addition, VPS35 D620N facilitated enhanced HA-AKT signaling. Transcriptomic assessment and validation of protein levels identified the differential expression of CD44 and HMMR isoforms in VPS35 D620N mutant cells. We report that knockdown of HMMR or CD44 results in upregulated autophagy in cells expressing wild-type VPS35. However, only HMMR knockdown resulted in rescue of autophagy dysfunction by VPS35 D620N indicating a potential pathogenic role for this receptor and HA signaling in Parkinson's disease.
帕金森病（PD）的运动特征源于黑质中多巴胺能（DA）神经元的丧失，而自噬功能障碍与该疾病密切相关。据报道，VPS35基因（D620N）中的一个导致PD的家族性突变会抑制自噬。为了确定导致这种自噬缺陷的信号通路，我们使用野生型或表达VPS35 D620N的视黄酸分化的SH-SY5Y细胞进行RNA测序（RNA-Seq），进行了一项无偏筛选。我们报告称，表达VPS35 D620N的细胞表现出转录组变化，表明细胞外基质（ECM）-受体相互作用以及PI3K-AKT信号通路发生改变，PI3K-AKT信号通路是已知的调节自噬的通路。透明质酸（HA）是脑ECM的主要成分，通过ECM受体CD44（RNA-Seq的主要命中靶点）和HA介导的运动受体（HMMR）向自噬调节PI3K-AKT信号通路发出信号。我们发现，高分子量（>950 kDa）而非低分子量（15-40 kDa）的HA处理会抑制自噬。此外，VPS35 D620N促进了HA-AKT信号增强。转录组评估和蛋白质水平验证确定了VPS35 D620N突变细胞中CD44和HMMR异构体的差异表达。我们报告称，敲低HMMR或CD44会导致表达野生型VPS35的细胞中自噬上调。然而，只有敲低HMMR才能挽救VPS35 D620N引起的自噬功能障碍，这表明该受体和HA信号在帕金森病中具有潜在的致病作用。
