Altered iron-sulfur cluster transfer in Arabidopsis mitochondria reveals lipoyl synthase as a Janus-faced enzyme that generates toxic sulfide

Abstract Iron–sulfur (Fe–S) cluster are vital cofactors in all domains of life. Mitochondrial Fe–S cluster assembly occurs in two major steps to first build [2Fe–2S] clusters and subsequently assemble these into [4Fe–4S] clusters. The two assembly machineries are interconnected by glutaredoxin S15 (GRXS15) that transfers [2Fe–2S] clusters to the second machinery. Diminished cluster transfer activity of GRXS15 in Arabidopsis mitochondria causes specific defects associated with lipoyl synthase (LIP1) activity. Conversely, overexpression of LIP1 in wild-type plants causes the release of toxic amounts of sulfide that can be detoxified by increasing the capacity for sulfide fixation through overexpression of O -acetylserine-(thiol)-lyase. The release of sulfide by lipoyl synthase causes a disturbance of mitochondrial sulfide homeostasis resulting in distinct and readily observable macroscopic phenotypes. These phenotypes enable a direct readout of consequences resulting from defects in Fe–S cluster assembly or targeted modulation of Fe–S cluster flux in mitochondria.