De Henau S et al. (2015) Nat Commun "A redox signalling globin is essential for reproduction in Caenorhabditis elegans."

Moens L, Pauwels M, Dewilde S, Tilleman L, De Henau S, Germani F, Vanfleteren JR, Vangheel M, Bolognesi M, Nardini M, Pesce A, Bert W, De Wael K, Vlaeminck C, Luyckx E, Trashin S, Braeckman BP

[Nat Commun, 2015]

Moderate levels of reactive oxygen species (ROS) are now recognized as redox signalling molecules. However, thus far, only mitochondria and NADPH oxidases have been identified as cellular sources of ROS in signalling. Here we identify a globin (GLB-12) that produces superoxide, a type of ROS, which serves as an essential signal for reproduction in C. elegans. We find that GLB-12 has an important role in the regulation of multiple aspects in germline development, including germ cell apoptosis. We further describe how GLB-12 displays specific molecular, biochemical and structural properties that allow this globin to act as a superoxide generator. In addition, both an intra- and extracellular superoxide dismutase act as key partners of GLB-12 to create a transmembrane redox signal. Our results show that a globin can function as a driving factor in redox signalling, and how this signal is regulated at the subcellular level by multiple control layers.

De Henau S et al. (2020) Dev Cell "Mitochondria-Derived H2O2 Promotes Symmetry Breaking of the C.elegans Zygote."

De Henau S, Pages-Gallego M, Dansen TB, Pannekoek WJ

[Dev Cell, 2020]

Symmetry breaking is an essential step in cell differentiation and early embryonic development. However, the molecular cues that trigger symmetry breaking remain largely unknown. Here, we show that mitochondrial H₂O₂ acts as a symmetry-breaking cue in the C.elegans zygote. We find that symmetry breaking is marked by a local H₂O₂ increase and coincides with a relocation of mitochondria to the cell cortex. Lowering endogenous H₂O₂ levels delays the onset of symmetry breaking, while artificially targeting mitochondria to the cellular cortex using a light-induced heterodimerization technique is sufficient to initiate symmetry breaking in a H₂O₂-dependent manner. In wild-type development, both sperm and maternal mitochondria contribute to symmetry breaking. Our findings reveal that mitochondrial H₂O₂-signaling promotes the onset of polarization, a fundamental process in development and cell differentiation, and this is achieved by both mitochondrial redistribution and differential H₂O₂-production.

Dansen TB et al. (2021) STAR Protoc "Modulating organelle distribution using light-inducible heterodimerization in C.elegans."

De Henau S, Dansen TB

[STAR Protoc, 2021]

The relative positioning of organelles underlies fundamental cellular processes, including signaling, polarization, and cellular growth. Here, we describe the usage of a light-dependent heterodimerization system, LOVpep-ePDZ, to alter organelle positioning locally and reversibly in order to study the functional consequences of organelle positioning. The protocol gives details on how to accomplish expression of fusion proteins encoding this system, describes the imaging parameters to achieve subcellular activation in <i>C.elegans</i>, and may be adapted for use in other model systems. For complete details on the use and execution of this protocol, please refer to De Henau etal. (2020).

Schwartz AZA et al. (2020) Dev Cell "Stimulating Embryo Polarization with Mitochondrial Peroxide."

Nance J, Schwartz AZA

[Dev Cell, 2020]

Centrosomes break symmetry in the C.elegans one-cell embryo, triggering its anterior-posterior polarization and initiating segregation of somatic and germline cell lineages. In this issue of Developmental Cell, De Henau etal. show that mitochondria also contribute to symmetry breaking by producing hydrogen peroxide at the egg's future posterior pole.

Eberhardt AG et al. (2008) Vet Parasitol "The Strongyloides (Nematoda) of sheep and the predominant Strongyloides of cattle form at least two different, genetically isolated populations."

Mayer WE, Eberhardt AG, Streit A, Bonfoh B

[Vet Parasitol, 2008]

Strongyloides sp. (Nematoda) are very wide spread small intestinal parasites of vertebrates that can form a facultative free-living generation. Most authors considered all Strongyloides of farm ruminants to belong to the same species, namely Strongyloides papillosus (Wedl, 1856). Here we show that, at least in southern Germany, the predominant Strongyloides found in cattle and the Strongyloides found in sheep belong to separate, genetically isolated populations. While we did find mixed infections in cattle, one form clearly dominated. This variety, in turn, was never found in sheep, indicating that the two forms have different host preferences. We also present molecular tools for distinguishing the two varieties, and an analysis of their phylogenetic relationship with the human parasite Strongyloides stercoralis and the major laboratory model species Strongyloides ratti. Based on our findings we propose that Strongyloides from sheep and the predominant Strongyloides from cattle should be considered separate species as it had already been proposed by [Brumpt, E., 1921. Recherches sur le determinisme des sexes et de l''evolution des Anguillules parasites (Strongyloides). Comptes rendu hebdomadaires des seances et memoires de la Societe de Biologie et de ses filiales 85, 149-152], but was largely ignored by later authors. For nomenclature, we follow [Brumpt, E., 1921. Recherches sur le determinisme des sexes et de l''evolution des Anguillules parasites (Strongyloides). Comptes rendu hebdomadaires des seances et memoires de la Societe de Biologie et de ses filiales 85, 149-152] and use the name S. papillosus for the Strongyloides of sheep and the name Strongyloides vituli for the predominant Strongyloides of cattle.

Braeckman BP et al. (2016) Antioxid Redox Signal "<i>In vivo</i> detection of ROS and redox status in <i>Caenorhabditis elegans</i>."

Back P, Smolders A, De Henau S, Braeckman BP

[Antioxid Redox Signal, 2016]

SIGNIFICANCE: Due to its large families of redox active enzymes, genetic amenability and complete transparency, the nematode &lt;i&gt;Caenorhabditis elegans&lt;/i&gt; has the potential to become an important model for the &lt;i&gt;in vivo&lt;/i&gt; study of redox biology. RECENT ADVANCES: The recent development of several genetically-encoded ratiometric ROS and redox sensors has revolutionized the quantification and precise localization of ROS and redox signals in living organisms. Only few exploratory studies have applied these sensors in &lt;i&gt;C. elegans&lt;/i&gt; and undoubtedly, much remains to be discovered in this model. As a follow-up to our recent findings that the &lt;i&gt;C. elegans&lt;/i&gt; somatic gonad uses superoxide and hydrogen peroxide signals to communicate with the germline, we here analyze the patterns of hydrogen peroxide inside the &lt;i&gt;C. elegans&lt;/i&gt; germline. CRITICAL ISSUES: Despite the advantages of genetically encoded ROS and redox sensors over classic chemical sensors, still several general as well as &lt;i&gt;C. elegans&lt;/i&gt; specific issues need to be addressed. The major concerns for the application of these sensors in &lt;i&gt;C. elegans&lt;/i&gt; are (1) decreased vitality of some reporter strains, (2) interference of autofluorescent compartments with the sensor signal and (3) the use of immobilization methods that do not influence the worm's redox physiology. FUTURE DIRECTIONS: We propose that several of the current issues may be solved by designing reporter strains carrying single copies of codon-optimized sensors. Preferably, these sensors should have their emission wavelengths in the red region, where autofluorescence is absent. Worm analysis could be optimized using 4D ratiometric fluorescence microscopy of worms immobilized in microfluidic chips.

Kelley CA et al. (2020) Mol Biol Cell "Redox signaling modulates Rho activity and tissue contractility in the C. elegans spermatheca."

Dansen TB, Cram EJ, Kelley CA, De Henau S, Bell L

[Mol Biol Cell, 2020]

Actomyosin based contractility in smooth muscle and non-muscle cells is regulated by signaling through the small GTPase Rho and by calcium-activated pathways. We use the myoepithelial cells of the <i>Caenorhabditis elegans</i> spermatheca to study the mechanisms of coordinated myosin activation <i>in vivo</i>. Here, we show that redox signaling modulates RHO-1/Rho activity in this contractile tissue. Exogenously added as well as endogenously generated hydrogen peroxide decreases spermathecal contractility by inhibition of RHO-1, which depends on a conserved cysteine in its nucleotide binding site (C20). Further, we identify an endogenous gradient of H₂O₂ across the spermathecal tissue, which depends on the activity of the cytosolic superoxide dismutase, SOD-1. Collectively, we show that SOD-1 mediated H₂O₂ production regulates the redox environment and fine-tunes Rho activity across the spermatheca through oxidation of RHO-1 C20.

Gerber J et al. (2004) Mol Cell Biol "The yeast scaffold proteins Isu1p and Isu2p are required inside mitochondria for maturation of cytosolic Fe/S proteins."

Prohl C, Gerber J, Lill R, Muhlenhoff U, Neumann K

[Mol Cell Biol, 2004]

Iron-sulfur (Fe/S) proteins are located in mitochondria, cytosol, and nucleus. Mitochondrial Fe/S proteins are matured by the iron-sulfur cluster (ISC) assembly machinery. Little is known about the formation of Fe/S proteins in the cytosol and nucleus. A function of mitochondria in cytosolic Fe/S protein maturation has been noted, but small amounts of some ISC components have been detected outside mitochondria. Here, we studied the highly conserved yeast proteins Isu1p and Isu2p, which provide a scaffold for Fe/S cluster synthesis. We asked whether the Isu proteins are needed for biosynthesis of cytosolic Fe/S clusters and in which subcellular compartment the Isu proteins are required. The Isu proteins were found to be essential for de novo biosynthesis of both mitochondrial and cytosolic Fe/S proteins. Several lines of evidence indicate that Isu1p and Isu2p have to be located inside mitochondria in order to perform their function in cytosolic Fe/S protein maturation. We were unable to mislocalize Isu1p to the cytosol due to the presence of multiple, independent mitochondrial targeting signals in this protein. Further, the bacterial homologue IscU and the human Isu proteins (partially) complemented the defects of yeast Isu protein-depleted cells in growth rate, Fe/S protein biogenesis, and iron homeostasis, yet only after targeting to mitochondria. Together, our data suggest that the Isu proteins need to be localized in mitochondria to fulfill their functional requirement in Fe/S protein maturation in the cytosol.

Platzer K et al. (2018) Am J Hum Genet "De Novo Variants in MAPK8IP3 Cause Intellectual Disability with Variable Brain Anomalies."

Stegmann APA, Bonati MT, Panis B, Smith-Hicks C, Lemke JR, Pepler A, Wilson C, Iascone M, McWalter K, Brasington C, Allen W, Di Donato N, Platzer K, Ramos L, Edwards SL, Jamra R, Gamble CN, Mandel H, Stobe P, Mahida S, Marquardt T, Demmer LA, Miller KG, Falik-Zaccai T, Pinz H, Hellenbroich Y, Sticht H, Kok F, Cho MT, Stumpel CTRM, Shinde DN, Angione KM

[Am J Hum Genet, 2018]

Using exome sequencing, we have identified de novo variants in MAPK8IP3 in 13 unrelated individuals presenting with an overlapping phenotype of mild to severe intellectual disability. The de novo variants comprise six missense variants, three of which are recurrent, and three truncating variants. Brain anomalies such as perisylvian polymicrogyria, cerebral or cerebellar atrophy, and hypoplasia of the corpus callosum were consistent among individuals harboring recurrent de novo missense variants. MAPK8IP3 has been shown to be involved in the retrograde axonal-transport machinery, but many of its specific functions are yet to be elucidated. Using the CRISPR-Cas9 system to target six conserved amino acid positions in Caenorhabditis elegans, we found that two of the six investigated human alterations led to a significantly elevated density of axonal lysosomes, and five variants were associated with adverse locomotion. Reverse-engineering normalized the observed adverse effects back to wild-type levels. Combining genetic, phenotypic, and functional findings, as well as the significant enrichment of de novo variants in MAPK8IP3 within our total cohort of 27,232 individuals who underwent exome sequencing, we implicate de novo variants in MAPK8IP3 as a cause of a neurodevelopmental disorder with intellectual disability and variable brain anomalies.

Vinci CR et al. (2007) J Biol Chem "Recognition of age-damaged (R,S)-adenosyl-L-methionine by two methyltransferases in the yeast Saccharomyces cerevisiae."

Vinci CR, Clarke SG

[J Biol Chem, 2007]

The biological methyl donor, S adenosylmethionine (AdoMet), can exist in two diastereoisomeric states with respect to its sulfonium ion. The "S" configuration, (S,S)AdoMet, is the only form that is produced enzymatically as well as the only form used in almost all biological methylation reactions. Under physiological conditions, however, the sulfonium ion can spontaneously racemize to the "R" form, producing (R,S)AdoMet. As of yet, (R,S)AdoMet has no known physiological function and may inhibit cellular reactions. In this study, two enzymes have been found in Saccharomyces cerevisiae that are capable of recognizing (R,S)AdoMet and using it to methylate homocysteine to form methionine. These enzymes are the products of the SAM4 and MHT1 genes, previously identified as homocysteine methyltransferases dependent upon AdoMet and S-methylmethionine respectively. We find here that Sam4 recognizes both (S,S) and (R,S)AdoMet, but its activity is much higher with the R,S form. Mht1 reacts with only the R,S form of AdoMet while no activity is seen with the S,S form. R,S-specific homocysteine methyltransferase activity is also shown here to occur in extracts of Arabidopsis thaliana, Drosophila melanogaster, and Caenorhabditis elegans, but has not been detected in several tissue extracts of Mus musculus. Such activity may function to prevent the accumulation of (R,S)AdoMet in these organisms.