Huang Shuan Shian, Chen Chun-Lin, Huang Franklin W, Hou Wei-Hsien, Huang Jung San
Auxagen Inc., St. Louis, Missouri, 63132.
Department of Biological Science, National Sun Yat-sen University and Doctoral Degree Program in Marine Biotechnology, National Sun Yat-sen University and Academia Sinica, Kaohsiung, 804, Taiwan.
J Cell Biochem. 2016 Jul;117(7):1568-79. doi: 10.1002/jcb.25448. Epub 2016 Feb 18.
Dimethyl sulfoxide (DMSO) is used to treat many diseases/symptoms. The molecular basis of the pharmacological actions of DMSO has been unclear. We hypothesized that DMSO exerts some of these actions by enhancing TGF-β activity. Here we show that DMSO enhances TGF-β activity by ∼3-4-fold in Mv1Lu and NMuMG cells expressing Smad-dependent luciferase reporters. In Mv1Lu cells, DMSO enhances TGF-β-stimulated expression of P-Smad2 and PAI-1. It increases cell-surface expression of TGF-β receptors (TβR-I and/or TβR-II) by ∼3-4-fold without altering their cellular levels as determined by (125) I-labeled TGF-β-cross-linking/Western blot analysis, suggesting the presence of large intracellular pools in these cells. Sucrose density gradient ultracentrifugation/Western blot analysis reveals that DMSO induces recruitment of TβR-II (but not TβR-I) from its intracellular pool to plasma-membrane microdomains. It induces more recruitment of TβR-II to non-lipid raft microdomains than to lipid rafts/caveolae. Mv1Lu cells transiently transfected with TβR-II-HA plasmid were treated with DMSO and analyzed by indirect immunofluoresence staining using anti-HA antibody. In these cells, TβR-II-HA is present as a vesicle-like network in the cytoplasm as well as in the plasma membrane. DMSO causes depletion of TβR-II-HA-containing vesicles from the cytoplasm and co-localization of TβR-II-HA and cveolin-1 at the plasma membrane. These results suggest that DMSO, a fusogenic substance, enhances TGF-β activity presumably by inducing fusion of cytoplasmic vesicles (containing TβR-II) and the plasma membrane, resulting in increased localization of TβR-II to non-lipid raft microdomains where canonical signaling occurs. Fusogenic activity of DMSO may play a pivotal role in its pharmacological actions involving membrane proteins with large cytoplasmic pools. J. Cell. Biochem. 117: 1568-1579, 2016. © 2015 Wiley Periodicals, Inc.
二甲基亚砜（DMSO）用于治疗多种疾病/症状。DMSO药理作用的分子基础尚不清楚。我们推测DMSO通过增强转化生长因子-β（TGF-β）活性发挥其中一些作用。在此我们表明，在表达依赖Smad的荧光素酶报告基因的Mv1Lu和NMuMG细胞中，DMSO可使TGF-β活性增强约3至4倍。在Mv1Lu细胞中，DMSO增强TGF-β刺激的磷酸化Smad2（P-Smad2）和纤溶酶原激活物抑制剂-1（PAI-1）的表达。通过¹²⁵I标记的TGF-β交联/蛋白质印迹分析确定，它使TGF-β受体（TβR-I和/或TβR-II）的细胞表面表达增加约3至4倍，而不改变其细胞内水平，这表明这些细胞中存在大量细胞内池。蔗糖密度梯度超速离心/蛋白质印迹分析显示，DMSO诱导TβR-II（而非TβR-I）从其细胞内池募集到质膜微结构域。与脂筏/小窝相比，它诱导更多的TβR-II募集到非脂筏微结构域。用抗HA抗体通过间接免疫荧光染色对瞬时转染TβR-II-HA质粒的Mv1Lu细胞进行处理并分析。在这些细胞中，TβR-II-HA以囊泡样网络形式存在于细胞质以及质膜中。DMSO导致含TβR-II-HA的囊泡从细胞质中耗竭，并使TβR-II-HA与小窝蛋白-1在质膜处共定位。这些结果表明，DMSO这种促融合物质可能通过诱导细胞质囊泡（含TβR-II）与质膜融合，从而增强TGF-β活性，导致TβR-II在发生经典信号传导的非脂筏微结构域中的定位增加。DMSO的促融合活性可能在其涉及具有大量细胞质池的膜蛋白的药理作用中起关键作用。《细胞生物化学杂志》117: 1568 - 1579, 2016。© Wiley期刊公司，2015年。
