Findings could aid development of novel therapeutics for conditions ranging from heart failure and stroke to cancer and neurodegeneration.
A protein embedded in the surface of mitochondria – the energy-producing batteries of living cells – opens the door to cell death, causing cells to experience severe power failures, according to new work by researchers at Temple University School of Medicine. The new study, appearing online September 17 in the journal Molecular Cell, suggests that blocking the door with a small molecule inhibitor could be key to the treatment of cardiovascular diseases such as heart attack and stroke, where extensive mitochondrial dysfunction and cell death hinder tissue recovery.
The study, led by Muniswamy Madesh, PhD, Associate Professor in the Department of Biochemistry, the Cardiovascular Research Center, and the Center for Translational Medicine at Temple University School of Medicine (TUSM), shows that the protein, spastic paraplegia 7 (SPG7), is the central component of the so-called permeability transition pore (PTP), a protein complex in the mitochondrial membrane that mediates necrotic cell death (death caused by cell injury).
The identification of SPG7 marks a major advance in scientists’ understanding of how the PTP affects necrosis. Although first described in 1976, the molecular parts of the pore have eluded discovery. “The only known molecular component of the PTP prior to our discovery of SPG7 was a protein called CypD, which is necessary for pore function,” Dr. Madesh explained.
To identify genes that modulate PTP opening induced by calcium overload or increased levels of reactive oxygen species (ROS) – the two primary factors that cause mitochondrial dysfunction and cell death via pore opening – Dr. Madesh’s team devised an RNA interference-based screen in which the activity of each gene under investigation was knocked down, or silenced, to examine its effects on mitochondrial calcium levels. The researchers began with a panel of 128 different genes but after initial screening narrowed the field to just 14 candidate PTP components. Subsequent experiments showed that the loss of only one of them – SPG7 – prevented pore opening.
Much of what is known about the PTP comes from studies of mitochondria in disease. In pathological states, particularly those involving hypoxia (oxygen deficiency), calcium and ROS accumulate within mitochondria, causing them to swell and prompting the PTP to open. Because pore opening disrupts the flow of electrons and protons across the mitochondrial membranes, which normally sustains energy production, it results in a catastrophic drop in cellular energy levels.
In the absence of disease, precisely how the PTP helps to mediate normal cellular physiology remains unclear. According to Dr. Madesh, “Under physiological conditions, SPG7 may function through transient pore openings to release toxic metabolites that have accumulated in mitochondria.” He plans to explore this possibility with knockout animal models.
Dr. Madesh also explained that the new findings could aid the development of novel therapeutics for conditions ranging from heart failure and stroke to cancer and neurodegeneration – all of which involve hypoxia and mitochondrial dysfunction to varying degrees. In these diseases, if the PTP could be prevented from opening, mitochondria could potentially continue to function, and cell death could be averted. With colleagues at TUSM, Dr. Madesh plans to explore the effects of SPG7 inhibitors in animals and, potentially, human patients.
Other researchers contributing to this work include Santhanam Shanmughapriya, Sudarsan Rajan, Nicholas E. Hoffman, Andrew M. Higgins, Dhanendra Tomar, Neeharika Nemani, Kevin J. Hines, Dylan J. Smith, Akito Eguchi, Sandhya Vallem, Farah Shaikh, Maggie Cheung, Nicole J. Leonard, Ryan S. Stolakis, Matthew P. Wolfers, and Neelakshi R. Jog in the Department of Biochemistry and Center for Translational Medicine at TUSM; Jessica Ibetti, J. Kurt Chuprun, Walter J. Koch, and John W. Elrod at the Center for Translational Medicine and Department of Pharmacology at TUSM; and Steven R. Houser at the Cardiovascular Research Center at TUSM.
Funding: The research was funded by the National Institutes of Health grants R01GM109882, R01HL086699, R01HL119306, and 1S10RR027327.
Source: Jeremy Walter – Temple University
Image Credit: The image is in the public domain
Original Research: Abstract for “SPG7 Is an Essential and Conserved Component of the Mitochondrial Permeability Transition Pore” by Santhanam Shanmughapriya, Sudarsan Rajan, Nicholas E. Hoffman, Andrew M. Higgins, Dhanendra Tomar, Neeharika Nemani, Kevin J. Hines, Dylan J. Smith, Akito Eguchi, Sandhya Vallem, Farah Shaikh, Maggie Cheung, Nicole J. Leonard, Ryan S. Stolakis, Matthew P. Wolfers, Jessica Ibetti, J. Kurt Chuprun, Neelakshi R. Jog, Steven R. Houser, Walter J. Koch, John W. Elrod, and Muniswamy Madesh in Molecular Cell. Published online September 17 2015 doi:10.1016/j.molcel.2015.08.009
Abstract
SPG7 Is an Essential and Conserved Component of the Mitochondrial Permeability Transition Pore
Highlights
•Mitochondrial SPG7 is essential for the PTP complex formation in multiple cell types
•SPG7 interacts with CypD and VDAC at the IMM and OMM contact site
•C terminus of SPG7 and CsA-binding region of CypD are necessary for PTP formation
•Loss of SPG7 protects mitochondria from Ca2+- and ROS-induced PTP-dependent necrosis
Summary
Mitochondrial permeability transition is a phenomenon in which the mitochondrial permeability transition pore (PTP) abruptly opens, resulting in mitochondrial membrane potential (ΔΨm) dissipation, loss of ATP production, and cell death. Several genetic candidates have been proposed to form the PTP complex, however, the core component is unknown. We identified a necessary and conserved role for spastic paraplegia 7 (SPG7) in Ca2+- and ROS-induced PTP opening using RNAi-based screening. Loss of SPG7 resulted in higher mitochondrial Ca2+ retention, similar to cyclophilin D (CypD, PPIF) knockdown with sustained ΔΨm during both Ca2+ and ROS stress. Biochemical analyses revealed that the PTP is a heterooligomeric complex composed of VDAC, SPG7, and CypD. Silencing or disruption of SPG7-CypD binding prevented Ca2+- and ROS-induced ΔΨm depolarization and cell death. This study identifies an ubiquitously expressed IMM integral protein, SPG7, as a core component of the PTP at the OMM and IMM contact site.
“SPG7 Is an Essential and Conserved Component of the Mitochondrial Permeability Transition Pore” by Santhanam Shanmughapriya, Sudarsan Rajan, Nicholas E. Hoffman, Andrew M. Higgins, Dhanendra Tomar, Neeharika Nemani, Kevin J. Hines, Dylan J. Smith, Akito Eguchi, Sandhya Vallem, Farah Shaikh, Maggie Cheung, Nicole J. Leonard, Ryan S. Stolakis, Matthew P. Wolfers, Jessica Ibetti, J. Kurt Chuprun, Neelakshi R. Jog, Steven R. Houser, Walter J. Koch, John W. Elrod, and Muniswamy Madesh in Molecular Cell. Published online September 17 2015 doi:10.1016/j.molcel.2015.08.009
