Criticality of the phosphate carrier SLC25A3 for mitochondrial inorganic phosphate uptake to sustain striated muscle function
Abstract
Significance: Mitochondria are the main source of energy (ATP) production in almost every cell in the body. Mitochondria are also very important for regulating calcium (Ca2+), which is a fundamental molecule that cells use for signaling purposes. The uptake by mitochondria of inorganic phosphate (Pi) is needed to make ATP and for the ability of mitochondria to regulate Ca2+ (which is a fundamental aspect for overall cellular Ca2+ regulation). The mitochondrial phosphate carrier (PiC) has long been considered as the primary mechanism of Pi uptake into mitochondria. However, only very recently have mutations in human PiC been discovered and a PiC-deficient mouse model has been generated, allowing PiC function to be directly studied in live organisms for the first time. Human PiC mutations cause severe symptoms soon after birth, and muscle is a key affected tissue. Some individuals die after a few months of life, but surprisingly, others recover despite having very little PiC in the muscle. This suggests that very little PiC is needed for ATP production and Ca2+ regulation or that mitochondria can find another way to take up Pi. We have obtained muscle cells from PiC-deficient humans and have generated mice with PiC removed in skeletal muscle (SkM). Thus, we are able, for the first time, to study the pathogenesis of PiC loss in SkM. These models will allow us to understand the (mal)adaptive mechanisms due to severe mitochondrial dysfunction, which, more generally, are poorly understood for mutations in nuclear DNA-encoded mitochondrial proteins such as PiC. The study of PiC is also relevant in light of the reported first human mutations in a protein (MICU1) needed for mitochondrial Ca2+ regulation. Mutations causing MICU1 loss cause severe muscle disease in humans; mitochondrial Pi uptake may partially rescue problems caused by MICU1 loss. Finally, with advances in DNA sequencing technologies, the incidence of pathogenic PiC variants in humans is expected to grow and thus the demand for treatment strategies will increase. Hypotheses: We will test if loss of PiC in SkM causes severe defects in energy production and Ca2+ regulation, affecting the ability of SkM to contract. We further propose that PiC loss is partly compensated by alternate routes of Pi uptake into mitochondria; these routes are known but have never been considered as important because this role has rarely been considered. We predict that these compensatory mechanisms will be able to take up enough Pi when energy demand is low, but not when demand is high. Model Systems: Mice and mouse cells with SkM-specific PiC loss, muscle cells from PiC-deficient and age-matched control humans. Aim 1: To test the role of PiC in SkM and for critical mechanisms counteracting PiC deficiency. We will investigate if PiC is the major route for Pi uptake into mitochondria under PiC replete conditions and when PiC is deficient or whether alternate routes of Pi uptake are operative. Behavioral and physiologic testing will assess SkM and diaphragm function in mice with SkM-specific PiC loss. To evaluate bioenergetic consequences and test for alternate Pi uptake pathways, studies in mitochondria and cells will test Pi transport, ATP production, and compensation by glycolytically derived ATP. To compare PiC needs at low- and high-energy demand, mice will be challenged with mild or vigorous exercise. Overall, we expect to learn (1) the importance of PiC vs. alternate Pi uptake pathways and (2) how SkM adapts to an energy crisis due to PiC loss. Aim 2: To test if PiC deficiency causes dysregulation of cellular Ca2+ and Ca2+ regulated functions. PiC-mediated Pi influx is coupled to mitochondrial Ca2+ uptake, affecting the amount of Ca2+ uptake and Ca2+ binding in mitochondria. Thus, PiC loss might alter important Ca2+ mediated functions in mitochondria. In particular, mitochondrial Ca2+ is important for regulating the Ca2+ needed for muscle contracti
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Oct 29, 2018
- Source ID
- W81XWH1710203
Entities
People
- Erin L Seifert
Organizations
- Thomas Jefferson University
- United States Army