Structural basis and regulation of the reductive stress response.

Although oxidative phosphorylation is best known for producing ATP, it also yields reactive oxygen species (ROS) as invariant byproducts. Depletion of ROS below their physiological levels, a phenomenon known as reductive stress, impedes cellular signaling and has been linked to cancer, diabetes, and cardiomyopathy. Cells alleviate reductive stress by ubiquitylating and degrading the mitochondrial gatekeeper FNIP1, yet it is unknown how the responsible E3 ligase CUL2FEM1B can bind its target based on redox state and how this is adjusted to changing cellular environments. Here, we show that CUL2FEM1B relies on zinc as a molecular glue to selectively recruit reduced FNIP1 during reductive stress. FNIP1 ubiquitylation is gated by pseudosubstrate inhibitors of the BEX family, which prevent premature FNIP1 degradation to protect cells from unwarranted ROS accumulation. FEM1B gain-of-function mutation and BEX deletion elicit similar developmental syndromes, showing that the zinc-dependent reductive stress response must be tightly regulated to maintain cellular and organismal homeostasis.

Ubiquitin-dependent remodeling of the actin cytoskeleton drives cell fusion.

Cell-cell fusion is a frequent and essential event during development, and its dysregulation causes diseases ranging from infertility to muscle weakness. Fusing cells need to repeatedly remodel their plasma membrane through orchestrated formation and disassembly of actin filaments, but how the dynamic reorganization of the cortical actin cytoskeleton is controlled is still poorly understood. Here, we identified a ubiquitin-dependent toggle switch that establishes reversible actin bundling during mammalian cell fusion. We found that EPS8-IRSp53 complexes stabilize cortical actin bundles at sites of cell contact to promote close membrane alignment. EPS8 monoubiquitylation by CUL3KCTD10 displaces EPS8-IRSp53 from membranes and counteracts actin bundling, a dual activity that restricts actin bundling to allow paired cells to progress with fusion. We conclude that cytoskeletal rearrangements during development are precisely controlled by ubiquitylation, raising the possibility of modulating the efficiency of cell-cell fusion for therapeutic benefit.

A Cellular Mechanism to Detect and Alleviate Reductive Stress.

Metazoan organisms rely on conserved stress response pathways to alleviate adverse conditions and preserve cellular integrity. Stress responses are particularly important in stem cells that provide lifetime support for tissue formation and repair, but how these protective systems are integrated into developmental programs is poorly understood. Here we used myoblast differentiation to identify the E3 ligase CUL2FEM1B and its substrate FNIP1 as core components of the reductive stress response. Reductive stress, as caused by prolonged antioxidant signaling or mitochondrial inactivity, reverts the oxidation of invariant Cys residues in FNIP1 and allows CUL2FEM1B to recognize its target. The ensuing proteasomal degradation of FNIP1 restores mitochondrial activity to preserve redox homeostasis and stem cell integrity. The reductive stress response is therefore built around a ubiquitin-dependent rheostat that tunes mitochondrial activity to redox needs and implicates metabolic control in coordination of stress and developmental signaling.

Investigation of candidate genes involved in the rhodoquinone biosynthetic pathway in Rhodospirillum rubrum.

The lipophilic electron-transport cofactor rhodoquinone (RQ) facilitates anaerobic metabolism in a variety of bacteria and selected eukaryotic organisms in hypoxic environments. We have shown that an intact rquA gene in Rhodospirillum rubrum is required for RQ production and efficient growth of the bacterium under anoxic conditions. While the explicit details of RQ biosynthesis have yet to be fully delineated, ubiquinone (Q) is a required precursor to RQ in R. rubrum, and the RquA gene product is homologous to a class I methyltransferase. In order to identify any additional requirements for RQ biosynthesis or factors influencing RQ production in R. rubrum, we performed transcriptome analysis to identify differentially expressed genes in anoxic, illuminated R. rubrum cultures, compared with those aerobically grown in the dark. To further select target genes, we employed a bioinformatics approach to assess the likelihood that a given differentially expressed gene under anoxic conditions may also have a direct role in RQ production or regulation of its levels in vivo. Having thus compiled a list of candidate genes, nine were chosen for further study by generation of knockout strains. RQ and Q levels were quantified using liquid chromatography-mass spectrometry, and rquA gene expression was measured using the real-time quantitative polymerase chain reaction. In one case, Q and RQ levels were decreased relative to wild type; in another case, the opposite effect was observed. These results comport with the crucial roles of rquA and Q in RQ biosynthesis, and reveal the existence of potential modulators of RQ levels in R. rubrum.

Unlocking a dark past.

A transcription factor called SALL4 could be the missing link between thalidomide and the limb defects caused by the drug.