Abstract
Deficiency of antiquitin (ATQ), an enzyme involved in lysine degradation, is the major cause of vitamin B6 -dependent epilepsy. Accumulation of the potentially neurotoxic α-aminoadipic semialdehyde (AASA) may contribute to frequently associated developmental delay. AASA is formed by α-aminoadipic semialdehyde synthase (AASS) via the saccharopine pathway of lysine degradation, or, as has been postulated, by the pipecolic acid (PA) pathway, and then converted to α-aminoadipic acid by ATQ. The PA pathway has been considered to be the predominant pathway of lysine degradation in mammalian brain; however, this was refuted by recent studies in mouse. Consequently, inhibition of AASS was proposed as a potential new treatment option for ATQ deficiency. It is therefore of utmost importance to determine whether the saccharopine pathway is also predominant in human brain cells. The route of lysine degradation was analyzed by isotopic tracing studies in cultured human astrocytes, ReNcell CX human neuronal progenitor cells and human fibroblasts, and expression of enzymes of the two lysine degradation pathways was determined by Western blot. Lysine degradation was only detected through the saccharopine pathway in all cell types studied. The enrichment of 15 N-glutamate as a side product of AASA formation through AASS furthermore demonstrated activity of the saccharopine pathway. We provide first evidence that the saccharopine pathway is the major route of lysine degradation in cultured human brain cells. These results support inhibition of the saccharopine pathway as a new treatment option for ATQ deficiency.
Highlights
Lysine degradation has been considered to have two distinct pathways, through saccharopine formation by ε-deamination, or through pipecolic acid (PA) formation by α-deamination or transamination (Figure 1), the first enzyme of the PA pathway has not yet been identified.[1]
It has been shown that ATQ is expressed in human astrocytes but not in neurons[20]; we examined the formation of aminoadipic acid (AAA) in astrocytes as well as in a human neural progenitor cell (NPC) line derived from the cortical region of human fetal brain (ReNcell CX NPCs), which we demonstrate express aminoadipic semialdehyde synthase (AASS) and ATQ
We examined the route of production of AAA in human astrocytes and in a human NPC line derived from the cortical region of human fetal brain (ReNcell CX NPCs), which express AASS, pipecolic acid oxidase (PIPOX), and ATQ, but not in neurons, as it has been shown that in human brain, neurons do not express ATQ.[20]
Summary
Lysine degradation has been considered to have two distinct pathways, through saccharopine formation by ε-deamination, or through pipecolic acid (PA) formation by α-deamination or transamination (Figure 1), the first enzyme of the PA pathway has not yet been identified.[1]. It was recently shown that in mouse brain, liver, and kidney, lysine is not degraded to AAA by the PA pathway but only through the saccharopine pathway.[13,14] Development of inhibitors of α-aminoadipic semialdehyde synthase (AASS), the enzyme which converts lysine to saccharopine, was proposed as a possible new treatment strategy in patients with ATQ deficiency.[13,14] As any activity through the PA pathway could result in accumulation of AASA/P6C even if AASS is inhibited, it is important to determine whether the saccharopine pathway is the major lysine degradation pathway in human brain, as the lysine degradation pathways appear to show species-specific activities.[15,16,17,18,19,33]. We examined whether activity through the PA pathway might increase when the saccharopine pathway is impaired, by comparing AASS-deficient and ATQ-deficient human fibroblasts to control fibroblasts
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