AbstractsBiology & Animal Science

Ubiquinone (Coenzyme Q, UQ or CoQ) is a small lipophilic molecule which is essential for all life forms that rely on mitochondrial respiration for energy production. It functions as an electron carrier in the respiratory chain and plays a role in many other cellular functions as well. In the past decade, an increasing number of patients have been described to have a genetic defect in UQ biosynthesis and present with severe and diverse clinical manifestations, making UQ research relevant to patient care. This thesis focuses on a particular enzyme of the UQ biosynthetic pathway in mice, which is encoded by the Mclk1/Coq7 gene and is responsible for the penultimate step of UQ biosynthesis: the hydroxylation of DMQ (demethoxy-UQ). I generated three conditional Mclk1 knockout (KO) models which are able to bypass the embryonic lethality of Mclk1 and allow us to examine the consequences of complete loss-of-function of Mclk1 in vitro and in vivo in adult animals. First, we created Mclk1 KO mouse embryonic fibroblasts (MEFs) in vitro. These cells do not synthesize UQ and accumulate DMQ. They are viable under standard (glucose) culture conditions. More interestingly, despite lacking UQ, they still have a functionally active electron transport chain. To understand this surprising phenomenon, we generated Pdss2 KO MEFs which are devoid of both DMQ and UQ. To our surprise, Pdss2 KO MEFs also are still capable of carrying out mitochondrial respiration. These observations suggest that 1) the respiratory phenotype of Mclk1 mutants can be considered essentially as a UQ-deficiency phenotype, and 2) mitochondrial respiration can occur in the absence of UQ. Mclk1 KO MEFs cannot survive in medium containing galactose instead of glucose as this forces cells to mostly rely on mitochondrial ATP production to sustain their viability. These characteristics make them a unique tool for testing the efficacy of UQ analogues in promoting electron transport in mammalian mitochondria. Next, we generated a mouse model with liver-specific loss of Mclk1. A large depletion of UQ in hepatocytes was found to cause only a mild impairment of respiratory chain function and no gross abnormalities. Therefore, liver mitochondrial function appears to have a high tolerance of severe UQ deficiency. Using this model, we also demonstrated that dietary UQ10 can functionally rescue the endogenous UQ deficiency of Mclk1 KO liver at the respiratory chain level. Furthermore, we generated an inducible Mclk1 KO model in which Mclk1 was knocked out in adult mice. These animals have a median survival time of 9 months after induction of global KO of the Mclk1 gene. We observed a very slow gradual decrement of tissue UQ contents after inducing the ablation of Mclk1, which could explain the slowly progressive phenotype in these mice. In the heart, kidneys and skeletal muscles severe deficits of UQ were found to severely impair mitochondrial respiration. Therefore, unlike the liver, the kidneys and muscle tissues need high levels of UQ to sustain sufficient respiratory function.…