A neuronal coping mechanism linking stress-induced anxiety to motivation for reward.

Stress coping involves innate and active motivational behaviors that reduce anxiety under stressful situations. However, the neuronal bases directly linking stress, anxiety, and motivation are largely unknown. Here, we show that acute stressors activate mouse GABAergic neurons in the interpeduncular nucleus (IPN). Stress-coping behavior including self-grooming and reward behavior including sucrose consumption inherently reduced IPN GABAergic neuron activity. Optogenetic silencing of IPN GABAergic neuron activation during acute stress episodes mimicked coping strategies and alleviated anxiety-like behavior. In a mouse model of stress-enhanced motivation for sucrose seeking, photoinhibition of IPN GABAergic neurons reduced stress-induced motivation for sucrose, whereas photoactivation of IPN GABAergic neurons or excitatory inputs from medial habenula potentiated sucrose seeking. Single-cell sequencing, fiber photometry, and optogenetic experiments revealed that stress-activated IPN GABAergic neurons that drive motivated sucrose seeking express somatostatin. Together, these data suggest that stress induces innate behaviors and motivates reward seeking to oppose IPN neuronal activation as an anxiolytic stress-coping mechanism.

Retro Is Cool: Structure of the Versatile Retromer Complex.

In this issue of Structure, Kendall et al. (2020) reveal the cryo-EM structure of the mammalian retromer complex, which is essential in sorting membrane proteins in endosomes. The retromer heterotrimer can oligomerize in multiple conformations; this versatility is promoted by a flexible interface of electrostatic residues on the VPS35 subunit.

Off the wall: The rhyme and reason of Neurospora crassa hyphal morphogenesis.

The fungal cell wall building processes are the ultimate determinants of hyphal shape. In Neurospora crassa the main cell wall components, ß-1,3-glucan and chitin, are synthesized by enzymes conveyed by specialized vesicles to the hyphal tip. These vesicles follow different secretory routes, which are delicately coordinated by cargo-specific Rab GTPases until their accumulation at the Spitzenkörper. From there, the exocyst mediates the docking of secretory vesicles to the plasma membrane, where they ultimately get fused. Although significant progress has been done on the cellular mechanisms that carry cell wall synthesizing enzymes from the endoplasmic reticulum to hyphal tips, a lot of information is still missing. Here, the current knowledge on N. crassa cell wall composition and biosynthesis is presented with an emphasis on the underlying molecular and cellular secretory processes.

Exposing the Elusive Exocyst Structure.

A major challenge for a molecular understanding of membrane trafficking has been the elucidation of high-resolution structures of large, multisubunit tethering complexes that spatially and temporally control intracellular membrane fusion. Exocyst is a large hetero-octameric protein complex proposed to tether secretory vesicles at the plasma membrane to provide quality control of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-mediated membrane fusion. Breakthroughs in methodologies, including sample preparation, biochemical characterization, fluorescence microscopy, and single-particle cryoelectron microscopy, are providing critical insights into the structure and function of the exocyst. These studies now pose more questions than answers for understanding fundamental functional mechanisms, and they open wide the door for future studies to elucidate interactions with protein and membrane partners, potential conformational changes, and molecular insights into tethering reactions.

Hyphal ontogeny in Neurospora crassa: a model organism for all seasons.

Filamentous fungi have proven to be a better-suited model system than unicellular yeasts in analyses of cellular processes such as polarized growth, exocytosis, endocytosis, and cytoskeleton-based organelle traffic. For example, the filamentous fungus Neurospora crassa develops a variety of cellular forms. Studying the molecular basis of these forms has led to a better, yet incipient, understanding of polarized growth. Polarity factors as well as Rho GTPases, septins, and a localized delivery of vesicles are the central elements described so far that participate in the shift from isotropic to polarized growth. The growth of the cell wall by apical biosynthesis and remodeling of polysaccharide components is a key process in hyphal morphogenesis. The coordinated action of motor proteins and Rab GTPases mediates the vesicular journey along the hyphae toward the apex, where the exocyst mediates vesicle fusion with the plasma membrane. Cytoplasmic microtubules and actin microfilaments serve as tracks for the transport of vesicular carriers as well as organelles in the tubular cell, contributing to polarization. In addition to exocytosis, endocytosis is required to set and maintain the apical polarity of the cell. Here, we summarize some of the most recent breakthroughs in hyphal morphogenesis and apical growth in N. crassa and the emerging questions that we believe should be addressed.

Role of BGT-1 and BGT-2, two predicted GPI-anchored glycoside hydrolases/glycosyltransferases, in cell wall remodeling in Neurospora crassa.

Neurospora crassa BGT-1 (NCU06381) and BGT-2 (NCU09175) are two putative glycoside hydrolases (GHs) with additional predicted glycosyltransferase activity and binding sites for a glycosyl phosphatidyl inositol (GPI) anchor that would facilitate their attachment to the plasma membrane (PM). To discern their role in key morphogenetic events during vegetative development of N. crassa, BGT-1 and BGT-2 were labeled with the green fluorescent protein (GFP). The gfp was inserted immediately after the signal peptide sequence, within the bgt-1 encoding sequence, or directly before the GPI-binding site in the case of bgt-2. Both BGT-1-GFP and BGT-2-GFP were observed at the PM of the hyphal apical dome, excluding the foremost apical region and the Spitzenkörper (Spk), where chitin and ß-1,3-glucan synthases have been previously found. These and previous studies suggest a division of labor of the cell wall synthesizing machinery at the hyphal dome: at the very tip, glucans are synthesized by enzymes that accumulate at the Spk, before getting incorporated into the PM, whereas at the subtending zone below the apex, glucans are presumably hydrolyzed, producing amenable ends for further branching and crosslinking with other cell wall polymers. Additionally, BGT-1-GFP and BGT-2-GFP were observed at the leading edge of new developing septa, at unreleased interconidial junctions, at conidial poles, at germling and hyphal fusion sites, and at sites of branch emergence, all of them processes that seemingly involve cell wall remodeling. Even though single and double mutant strains for the corresponding genes did not show a drastic reduction of growth rate, bgt-2Δ and bgt-1Δ::bgt-2Δ strains exhibited an increased resistance to the cell wall stressors calcofluor white (CW) and congo red (CR) than the reference strain, which suggests they present significant architectural changes in their cell wall. Furthermore, the conidiation defects observed in the mutants indicate a significant role of BGT-1 and BGT-2 on the re-arrangement of glucans needed at the conidiophore cell wall to allow conidial separation.