Yegambaram Manivannan, Pokharel Marissa D, Sun Xutong, Lu Qing, Soto Jamie, Aggarwal Saurabh, Maltepe Emin, Fineman Jeffery R, Wang Ting, Black Stephen M
Center for Translational Science, Florida International University, 11350 SW Village Parkway, Port St. Lucie, FL 34987-2352, USA.
Department of Cellular Biology & Pharmacology, Herbert Wertheim College of Medicine, Florida International University, Miami, FL 33199, USA.
Int J Mol Sci. 2025 Apr 17;26(8):3815. doi: 10.3390/ijms26083815.
It is well-established that mitochondrial dysfunction plays a critical role in the development of pulmonary hypertension (PH). However, the molecular mechanisms and how the individual electron transport complexes (ETC) may be affected are poorly understood. In this study, we identified decreased ETC Complex I activity and assembly and linked these changes to disrupted mitochondrial bioenergetics in pulmonary arterial endothelial cells (PAECs) isolated from a lamb model of PH with increased pulmonary blood flow (Shunt). These derangements were associated with decreased mitochondrial activity of the protein tyrosine kinase, pp60. Treating Control PAECs with either the Src family kinase inhibitor, PP2, or the siRNA-mediated knockdown of pp60 was able to recapitulate the adverse effects on ETC Complex I activity and assembly and mitochondrial bioenergetics. Conversely, restoring pp60 activity in lamb PH PAECs re-established ETC Complex I activity, improved ETC Complex I assembly and enhanced mitochondrial bioenergetics. Phosphoprotein enrichment followed by two-dimensional gel electrophoresis and tandem mass spectrometry was used to identify three ETC Complex I subunits (NDUFS1, NDUFAF5, and NDUFV2) as pp60 substrates. Finally, we demonstrated that the pY levels of NDUFS1, NDUFAF5, and NDUFV2 are decreased in lamb PH PAECs. Enhancing mitochondrial pp60 activity could be a therapeutic strategy to reverse PH-related mitochondrial dysfunction.
线粒体功能障碍在肺动脉高压（PH）的发展中起关键作用，这一点已得到充分证实。然而，其分子机制以及各个电子传递复合体（ETC）可能如何受到影响却知之甚少。在本研究中，我们发现从具有增加肺血流量（分流）的PH羔羊模型中分离出的肺动脉内皮细胞（PAECs）中，ETC复合体I的活性和组装降低，并将这些变化与线粒体生物能量学紊乱联系起来。这些紊乱与蛋白酪氨酸激酶pp60的线粒体活性降低有关。用Src家族激酶抑制剂PP2处理对照PAECs，或通过siRNA介导敲低pp60，都能够重现对ETC复合体I活性、组装和线粒体生物能量学的不利影响。相反，恢复羔羊PH PAECs中的pp60活性可重新建立ETC复合体I活性，改善ETC复合体I组装并增强线粒体生物能量学。通过二维凝胶电泳和串联质谱进行磷蛋白富集，以鉴定三个ETC复合体I亚基（NDUFS1、NDUFAF5和NDUFV2）作为pp60底物。最后，我们证明在羔羊PH PAECs中NDUFS1、NDUFAF5和NDUFV2的磷酸化酪氨酸（pY）水平降低。增强线粒体pp60活性可能是一种逆转PH相关线粒体功能障碍的治疗策略。
