Yanchang Bian

Heme is essential for numerous cellular processes and is dynamically trafficked within cells, yet its labile form is a potent pro-oxidant and signaling molecule whose concentration must be held within a narrow physiological range. How cells achieve this control remains poorly understood. Here we identify the conserved endoplasmic reticulum (ER) membrane protein TMEM161B as a key regulator of the labile heme pool. We report cryo-EM structures of human TMEM161B in monomeric and dimeric states that further assemble into extended filamentous arrays through two distinct oligomerization interfaces. The cytosolic loop of each dimer forms a pseudo-C2-symmetric loop clamp that coordinates up to 8–9 heme molecules, representing a previously undescribed mode of membrane-associated heme binding. Using a genetically encoded cytosolic heme sensor, we show that TMEM161B overexpression reduces the labile heme pool whereas knockdown elevates it, accompanied by corresponding heme-responsive transcriptional changes. In C. elegans, loss of the TMEM161 ortholog strongly suppresses the hrg-1p::GFP reporter, consistent with an elevated available heme pool, demonstrating that this buffering function is conserved from nematodes to humans. Together, these findings define TMEM161B as a membrane-embedded heme buffer that assembles into linear arrays on the ER, and raise the possibility that the labile heme pool is spatially compartmentalized across organelle membranes to a greater extent than currently recognized.

Oxidative protein folding in the endoplasmic reticulum (ER) is essential for eukaryotic cells yet generates hydrogen peroxide (H2O2), a reactive oxygen species. The ER-transmembrane protein that supports ER proteostasis and guards the cytosol for antioxidant defense remains unidentified. Here, we combine AlphaFold2 and functional screens in C. elegans to discover a previously uncharacterized and evolutionarily conserved protein ERGU-1 that fulfills these roles. Deleting ERGU-1 upregulates H2O2 and NRF2/SKN-1-dependent gene expression. ERGU-1 deficiency also impairs organismal reproduction and behavioral responses to H2O2. Both C. elegans ERGU-1 and human homolog TMEM161B localize to ER membranes, forming reticular networks. Human and Drosophila homologs of ERGU-1 rescue C. elegans mutant phenotypes, demonstrating ancient and conserved functions. In addition, purified ERGU-1 and TMEM161B exhibit redox-modulated oligomeric states. Together, our results reveal an ER-membrane-specific machinery, suggesting a conserved mechanism for maintaining ER redox homeostasis and proteostasis in animal cells.

Although biosensors for specific cellular ions are widely available, real-time monitoring of overall ionic strength in living organisms remains challenging. Here, we present a genetically encoded nuclear translocation ionic sensor (GENTIS) that enables direct visualization of ionic stress in vivo. Using this sensor alongside longitudinal tracking via an automated microfluidic platform, we find that Caenorhabditis elegans larvae experience highly synchronized, rhythmic elevations in intestinal ionic strength during the molt, a stage during which developmentally timed sleep occurs. Cytosolic proton accumulation through inhibition of vacuolar-type adenosine triphosphatases (V-ATPases) triggers GENTIS nuclear translocation and evokes behavioral quiescence, characterized by reduced feeding, locomotion, and activation of sleep-active neurons. Apical membrane V-ATPases naturally undergo disassembly during molting and stress, conditions that cause proton accumulation and sleep. Notably, this proton-linked sleep is suppressed by proton buffering with ammonium. Together, these findings establish GENTIS as a powerful tool for tracking ionic strength dynamics in vivo and reveal that proton ionic rhythms contribute to the regulation of sleep.