Regulation of lipid saturation without sensing membrane fluidity

Abstract

20Cells maintain membrane fluidity by regulating lipid saturation, but the molecular 21 mechanisms of this homeoviscous adaptation remain poorly understood. Here, we have 22 reconstituted the core machinery for sensing and regulating lipid saturation in baker's yeast 23 to directly characterize its response to defined membrane environments. Using spectroscopic 24 techniques and in vitro ubiquitylation, we uncover a unique sensitivity of the transcriptional 25 regulator Mga2 to the abundance, position, and configuration of double bonds in lipid acyl 26 chains and provide unprecedented insight into the molecular rules of membrane adaptivity. 27 Our data challenge the prevailing hypothesis that membrane viscosity serves as the 28 measured variable for regulating lipid saturation. Rather, we show that the signaling output of 29 Mga2 correlates with the size of a single sensor residue in the transmembrane helix, which 30 senses the lateral pressure and/or compressibility profile in a defined region of the 31 membrane. Our findings suggest that membrane property sensors have evolved remarkable 32 sensitivities to highly specific aspects of membrane structure and dynamics, thus paving the 33 way toward the development of genetically encoded reporters for such membrane properties 34 in the future. 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 Keywords 51 Membrane fluidity, homeoviscous response, lateral pressure profile, physicochemical 52 membrane homeostasis, lipid saturation, membrane property sensors, unsaturated fatty 53 acids, saturated fatty acids, Mga2, Spt23, Ole1, Rsp5, proteasome, ubiquitylation.54 55 Cellular membranes are complex assemblies of proteins and lipids, which collectively 56 determine physical bilayer properties such as membrane viscosity, permeability, and the 57 lateral pressure profile 1-4 . The acyl chain composition of membrane lipids is an important 58 determinant of membrane viscosity and tightly controlled in bacteria 5-7 , fungi 8,9 , worms 10,11 , 59 flies 12 , and vertebrates 13,14 . Saturated lipid acyl chains tend to form non-fluid, tightly packed 60 gel phases, while unsaturated lipid acyl chains fluidize the bilayer. Poikilothermic organisms 61 that cannot control their body temperature must adjust their lipid composition during cold 62 stress to maintain membrane functions-a phenomenon referred to as the homeoviscous 63 adaptation 15-17 . Despite recent advances in identifying candidate sensory, it remains largely 64 unknown how these sensors work on the molecular scale and how they are coordinated for 65 maintaining a physicochemical membrane homeostasis 20,21 . The fact that most, if not all, 66 membrane properties are interdependent is a key challenge for this emerging field. How do 67 cells, for example, balance the need for maintaining membrane viscosity with the need to 68 maintain organelle-specific lateral pressure profiles 22 ? In fact, perturbation of membrane 69 viscosity by genetically targeting fatty acid metabolism leads to complex changes throughout 70 the en...

Delineating the Rules for Structural Adaptation of Membrane-Associated Proteins to Evolutionary Changes in Membrane Lipidome

“…Phospholipid asymmetry and FA tail saturation both reduce membrane permeability to water, small solutes, and/or oxygen [50][51][52][53], which might provide additional advantages in the wild. Budding yeast and S. pombe utilize specialized gene expression programs executed by the membrane-sensing transcription factor Mga2 [54] to sustain FA desaturation in low-oxygen conditions [25,55]. In the future, it will be of interest to assess the wiring of this network in S. japonicus, given a markedly distinct physiology of the organism.…”

Cysteine crosslinking in native membranes establishes the transmembrane architecture of Ire1

“…2) Mutagenesis of individual TMH residues (V543-F551) including three aromatic residues lining one side of the TMH – one of which being the best-crosslinking residue F544C - causes no relevant functional defects (Figure S3A). These findings are notable because aromatic residues have been implicated in lipid/membrane sensing in other systems (37, 45, 46). 3) Our crosslinking data provide no evidence for a rotational re-organization of the TMHs during lipid bilayer stress (Figure 3C,D, Figure S3C).…”

“…scored the oligomerization propensity of IRE1α in cells via bimolecular fluorescence complementation in palmitate-treated cells and suggested that the increased lipid saturation might induce a conformational switch in the TMH region, which relies on a tryptophan (W457) as putative sensing residue and a conserved leucine zipper motif (SxxLxxx) involving serine 450 (37, 44). Intriguingly, such a rotation-based mechanism of sensing would be reminiscent of the lipid saturation sensor Mga2 from baker’s yeast controlling the expression of the essential fatty acid desaturase-encoding gene OLE1 (45, 46).…”