Iron is an essential cofactor, but it is also toxic at high levels. In Schizosaccharomyces pombe, the sensor glutaredoxin Grx4 guides the activity of the repressors Php4 and Fep1 to mediate a complex transcriptional response to iron deprivation: activation of Php4 and inactivation of Fep1 leads to inhibition of iron usage/storage, and to promotion of iron import, respectively. However, the molecular events ruling the activity of this double-branched pathway remained elusive. We show here that Grx4 incorporates a glutathione-containing iron-sulfur cluster, alone or forming a heterodimer with the BolA-like protein Fra2. Our genetic study demonstrates that Grx4-Fra2, but not Fep1 nor Php4, participates not only in iron starvation signaling but also in iron-related aerobic metabolism. Iron-containing Grx4 binds and inactivates the Php4 repressor; upon iron deprivation, the cluster in Grx4 is probably disassembled, the proteins dissociate, and Php4 accumulates at the nucleus and represses iron consumption genes. Fep1 is also an iron-containing protein, and the tightly bound iron is required for transcriptional repression. Our data suggest that the cluster-containing Grx4-Fra2 heterodimer constitutively binds to Fep1, and upon iron deprivation the disassembly of the iron cluster between Grx4 and Fra2 promotes reverse metal transfer from Fep1 to Grx4-Fra2, and de-repression of iron-import genes. Our genetic and biochemical study demonstrates that the glutaredoxin Grx4 independently governs the Php4 and Fep1 repressors through metal transfer. Whereas iron loss from Grx4 seems to be sufficient to release Php4 and allow its nuclear accumulation, total or partial disassembly of the Grx4-Fra2 cluster actively participates in iron-containing Fep1 activation by sequestering its iron and decreasing its interaction with promoters.

Oxysterol binding protein-related proteins (ORPs) are conserved lipid binding polypeptides, enriched at ER contacts sites. ORPs promote non-vesicular lipid transport and work as lipid sensors in the context of many cellular tasks, but the determinants of their distinct localization and function are not understood. Here, we demonstrate that the yeast endocytic invaginations associate with the ER and that this association specifically requires the ORPs Osh2 and Osh3, which bridge the endocytic myosin-I Myo5 to the ER integral-membrane VAMP-associated protein (VAP) Scs2. Disruption of the ER contact with endocytic sites using ORP, VAP, myosin-I, or reticulon mutants delays and weakens actin polymerization and interferes with vesicle scission. Finally, we provide evidence suggesting that ORP-dependent sterol transfer facilitates actin polymerization at endocytic sites.

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Cysteine residues, and in particular their thiolate groups, react not only with reactive oxygen species but also with electrophiles and with reactive nitrogen species. Thus, cysteine oxidation has often been linked to the toxic effects of some of these reactive molecules. However, thiol-based switches are common in protein sensors of antioxidant cascades, in both prokaryotic and eukaryotic organisms. We will describe here three redox sensors, the transcription factors OxyR, Yap1 and Pap1, which respond by disulfide bond formation to hydrogen peroxide stress, focusing specially on the differences among the three peroxide-sensing mechanisms.

SummaryCell separation in Schizosaccharomyces pombe is achieved through the concerted action of the Eng1 endo-b-1,3-glucanase and the Agn1 endo-a-1,3-glucanase, which are transported to the septum and localize to a ring-like structure that surrounds the septum. Correct localization of these hydrolases requires the presence of both the septins and the exocyst. In this work, we show that the glucanase Eng1 contains a region at the C-terminus that acts as a carbohydrate-binding module (CBM) and that it is not present in other members of glycoside hydrolases family 81 (GH81). In vitro, the purified CBM has affinity for b-1,3-glucan chains with a minimum degree of polymerization of 30 glucose units. Deletion of the CBM results in a protein that is largely defective in complementing the separation defect of eng1D mutants. This defect is due to a reduction in the catalytic activity against insoluble substrates and to a defect in targeting of Eng1 to the septum, as the truncated protein localizes to the lateral cell wall of the cell. Thus, the targeting of Eng1 to the primary septum requires not only trans-factors (septins and the exocyst complex) but also a cis-element localized to the C-terminus of the protein.