The department is focused on the study of the lateral arrangement of biological membranes into microdomains, i.e. areas with specific shape, composition and function. We put the emphasis on their involvement in the regulation of cellular processes in response to environmental stimuli. Taking maximum advantage of the genetically accessible yeast model, we particularly investigate the role of membrane microdomains in stress perception and adaptation and in signaling and regulation of cellular metabolism. These membrane functions presuppose communication between different microdomains, both within one particular membrane and between different specialized membranes within the membrane system of a eukaryotic cell.
Glycolysis arrest is a prerequisite for binding of the evolutionarily conserved exoribonuclease Xrn1 to the plasma membrane
We show that membrane binding of Xrn1 depends on glycolytic flux rather than on the availability of fermentable carbon, is independent of TORC1 activity, and requires the essential eisosomal proteins Pil1 and Lsp1. In addition, we demonstrate that when expressed in Saccharomyces cerevisiae, the human ortholog of Xrn1 behaves identically to its yeast counterpart. We extend the current understanding of Xrn1 regulation and suggest that this regulatory mechanism is conserved from yeast to humans.
Human Xrn1 binds to eisosomes. The subcellular localisations of yeast Xrn1-GFP and its human ortholog HsXrn1-GFP were compared in post-diauxic (non-fermenting) cells before (left column) and after the addition of exogenous glucose to the culture media (right). Note the identical distribution patterns of both the proteins expressed in xrn1Δ yeast strain. Scale bar: 5 μm.
Publication:
Mondal S, Zahumensky J, Vesela P, Malinsky J. Conserved mechanism of Xrn1 regulation by glycolytic flux and protein aggregation. Heliyon 10 (19):e38786 (2024)
Lsp1 is able to assemble eisosomes under stress conditions
Eisosomes are protein complexes that stabilise plasma membrane microdomains in the cells of many organisms. They consist of BAR domain-containing Pil1 family proteins. The yeast S. cerevisiae has two such proteins, Pil1 and Lsp1. Under normal conditions, deletion of the PIL1 gene leads to loss of the eisosome. We found that under stress, Lsp1 substitutes for the Pil1 function and is enough to assemble the eisosome. Our results identify Lsp1 as an important regulator of the cellular stress response.
Lsp1 substitutes for Pil1 function in eisosome assembly under stress conditions. Freeze-fracture electron microscopy images of the plasma membrane in the wild type (A; normal condition) and pil1Δ (B,C; normal condition and salt stress, respectively) yeast cells. Note the presence of membrane invaginations (furrows) in C. (D–G) Detailed view of furrows in pil1Δ cells exposed to salt stress (F corresponds to the area in the black square in C). Scale bars: 500 nm (A–C), 200 nm (D–G).
Publication:
Vesela P, Zahumensky J, Malinsky J. Lsp1 partially substitutes for Pil1 function in eisosome assembly under stress conditions. J Cell Sci. 136 (3):jcs260554 (2023)
Two different phospholipases C, Isc1 and Pgc1, cooperate to regulate mitochondrial function
Mitochondria are constantly adapting to changes in nutrient availability and environmental stresses. We have proposed a model in which this adaptation is mediated by lipids. Specifically, we have shown that mitochondrial phospholipids regulate cellular sphingolipid biosynthesis and vice versa. In this way, the cell is able to coordinate mitochondrial structure and performance with the actual needs of the cellular metabolism. This seems to be a universally applicable principle of cellular regulation.
Mechanism for the mutual regulation of Isc1 and Pgc1 phospholipases. Two lipids, diacylglycerol (DAG) and phosphatidylglycerol (PG), inhibit the key enzymes of mitochondrial phospholipid biosynthesis: (i) the PG-phosphate synthase Pgs1, (ii) the phosphatidylserine (PS) decarboxylase Psd1, and (iii) the inositol phosphosphingolipid phospholipase C Isc1. The latter is of particular importance because ceramide, a product of Isc1-catalyzed hydrolysis, inhibits the PG-specific phospholipase C Pgc1, which degrades PG to form DAG. The entire loop ensures the efficient control of PG (cardiolipin) and phosphatidylethanolamine (PE) synthesis in the context of environmental stress stimuli affecting the sphingolipid biosynthetic pathway. OMM – outer mitochondrial membrane; IMM – inner mitochondrial membrane; IPC – inositol phosphoceramide.
Publication:
Balazova, M., Vesela, P., Babelova, L., Durisova, I., Kanovicova, P., Zahumensky, J., Malinsky, J.: (2022) Two different phospholipases C, Isc1 and Pgc1, cooperate to regulate mitochondrial function. Microbiology Spectrum. 10(6): e02489-22.
Microdomain protein Nce102 is a local sensor of plasma membrane sphingolipid balance
Sphingolipids, essential building blocks of eukaryotic membranes and important signaling molecules, are regulated in response to environmental inputs. We showed that the amount of Nce102 protein in the plasma membrane is a measure of the current need for sphingolipids in the cell, whereas its local distribution marks sites of high sphingolipid demand. Mass spectrometry analysis revealed reduced levels of hydroxylated complex sphingolipids in stress response in cells lacking Nce102.
The plasma membrane microdomain protein Nce102 contributes to the regulation of sphingolipid metabolism. (A) Synthesis of sphingolipids, important structural and signalling molecules, was inhibited by the addition of myriocin at time t = 0 min. As the yeast cells grew and divided, the punctate pattern typical for Nce102-GFP was lost in the mother cells due to lowering of sphingolipid content. In the daughter cells, Nce102-GFP remained distributed uniformly and did not localize to microdomains. Scale bar: 5 μm. (B) Deletion of the NCE102 gene resulted in changes in the level of sphingolipid classes. In the absence of Nce102 the cellular amount of long-chain bases (PHS, DHS) decreased. On the other hand, phytoceramide (PhytoCer) levels were generally increased. Some complex sphingolipids (IPC and MIPC) were increased, some decreased. LFC: log 2 fold change relative to wild type cells.
Publication:
Zahumenský, J., Fernandes, C.M., Veselá, P., Del Poeta, M., Konopka, J.B., Malínský, J.: (2022) Microdomain Protein Nce102 Is a Local Sensor of Plasma Membrane Sphingolipid Balance. Microbiology Spectrum. 10(4): e0196122.
2024
mSphere. 2024 Dec 19;9(12):e0039124. doi: 10.1128/msphere.00391-24. Epub 2024 Nov 29.
