| 1. |
- Besse, Lenka, et al.
(author)
-
Treatment with HIV-protease inhibitor nelfinavir identifies membrane lipid composition and fluidity as a therapeutic target in advanced multiple myeloma
- 2021
-
In: Cancer Research. - 0008-5472 .- 1538-7445. ; 81, s. 4581-4593
-
Journal article (peer-reviewed)abstract
- The HIV-protease inhibitor nelfinavir has shown broad anticancer activity in various preclinical and clinical contexts.In patients with advanced, proteasome inhibitor (PI)-refractory multiple myeloma, nelfinavir-based therapy resulted in 65% partial response or better, suggesting that this may be a highly active chemotherapeutic option in this setting.The broad anticancer mechanism of action of nelfinavir implies that it interferes with fundamental aspects of cancer cell biology.We combined proteome-wide affinity-purification of nelfinavir-interacting proteins with genome-wide CRISPR/Cas9-based screening to identify protein partners that interact with nelfinavir in an activity-dependent manner alongside candidate genetic contributors affecting nelfinavir cytotoxicity.Nelfinavir had multiple activity-specific binding partners embedded in lipid bilayers of mitochondria and the endoplasmic reticulum.Nelfinavir affected the fluidity and composition of lipid-rich membranes, disrupted mitochondrial respiration, blocked vesicular transport, and affected the function of membrane-embedded drug efflux transporter ABCB1, triggering the integrated stress response.Sensitivity to nelfinavir was dependent on ADIPOR2, which maintains membrane fluidity by promoting fatty acid desaturation and incorporation into phospholipids.Supplementation with fatty acids prevented the nelfinavir-induced effect on mitochondrial metabolism, drug-efflux transporters, and stress-response activation.Conversely, depletion of fatty acids/cholesterol pools by the FDAapproved drug ezetimibe showed a synergistic anticancer activity with nelfinavir in vitro.These results identify the modification of lipid-rich membranes by nelfinavir as a novel mechanism of action to achieve broad anticancer activity, which may be suitable for the treatment of PI-refractory multiple myeloma.
|
|
| 2. |
- Bodhicharla, Rakesh, et al.
(author)
-
Membrane Fluidity Is Regulated Cell Nonautonomously by Caenorhabditis elegans PAQR-2 and Its Mammalian Homolog AdipoR2
- 2018
-
In: Genetics. - : Oxford University Press (OUP). - 0016-6731 .- 1943-2631. ; 210:1, s. 189-201
-
Journal article (peer-reviewed)abstract
- Maintenance of membrane properties is an essential aspect of cellular homeostasis of which the regulatory mechanisms remain mostly uncharacterized. In Caenorhabditis elegans, the PAQR-2 and IGLR-2 proteins act together as a plasma membrane sensor that responds to decreased fluidity by promoting fatty acid desaturation, hence restoring membrane fluidity. Here, we used mosaic analysis for paqr-2 and iglr-2, and tissue-specific paqr-2 expression, to show that membrane homeostasis is achieved cell nonautonomously. Specifically, we found that expression of paqr-2 in the hypodermis, gonad sheath cells, or intestine is sufficient to suppress systemic paqr-2 mutant phenotypes, including tail tip morphology, membrane fluidity in intestinal cells, cold and glucose intolerance, vitellogenin transport to the germline, germ cell development, and brood size. Finally, we show that the cell nonautonomous regulation of membrane homeostasis is conserved in human cells: HEK293 cells that express AdipoR2, a homolog of paqr-2, are able to normalize membrane fluidity in distant cells where AdipoR2 has been silenced. Finally, using C. elegans mutants and small interfering RNA against Δ9 stearoyl-CoA desaturase in HEK293 cells, we show that Δ9 desaturases are essential for the cell nonautonomous maintenance of membrane fluidity. We conclude that cells are able to share membrane components even when they are not in direct contact with each other, and that this contributes to the maintenance of membrane homeostasis in C. elegans and human cells.
|
|
| 3. |
- Busayavalasa, Kiran, et al.
(author)
-
Leveraging a gain-of-function allele of Caenorhabditis elegans paqr-1 to elucidate membrane homeostasis by PAQR proteins
- 2020
-
In: Plos Genetics. - : Public Library of Science (PLoS). - 1553-7404. ; 16:8
-
Journal article (peer-reviewed)abstract
- TheC.elegansproteins PAQR-2 (a homolog of the human seven-transmembrane domain AdipoR1 and AdipoR2 proteins) and IGLR-2 (a homolog of the mammalian LRIG proteins characterized by a single transmembrane domain and the presence of immunoglobulin domains and leucine-rich repeats in their extracellular portion) form a complex that protects against plasma membrane rigidification by promoting the expression of fatty acid desaturases and the incorporation of polyunsaturated fatty acids into phospholipids, hence increasing membrane fluidity. In the present study, we leveraged a novel gain-of-function allele of PAQR-1, a PAQR-2 paralog, to carry out structure-function studies. We found that the transmembrane domains of PAQR-2 are responsible for its functional requirement for IGLR-2, that PAQR-1 does not require IGLR-2 but acts via the same pathway as PAQR-2, and that the divergent N-terminal cytoplasmic domains of the PAQR-1 and PAQR-2 proteins serve a regulatory function and may regulate access to the catalytic site of these proteins. We also show that overexpression of human AdipoR1 or AdipoR2 alone is sufficient to confer increased palmitic acid resistance in HEK293 cells, and thus act in a manner analogous to the PAQR-1 gain-of-function allele. Author summary Cells are enclosed within membranes primarily composed of fat. When membranes contain much saturated fats, they tend to become more rigid, as with butter. Conversely, when membranes are rich in unsaturated fats, they become more fluid, as with vegetable oils. Our goal is to better understand how cells monitor and adjust the composition and properties of their membranes. We focus on a small group of proteins found in all animals, and called AdipoR1 and AdipoR2 in humans, and PAQR-1 and PAQR-2 in the wormCaenorhabditis elegans. We now found a version of PAQR-1 that is more "active", and promotes increased levels of unsaturated fats in membranes. By swapping different parts of the PAQR-1 protein with those of PAQR-2, we were able to determine which protein parts played which roles. We found that it is the transmembrane domains of PAQR-2 that dictate its requirements for another protein called IGLR-2 and that the intracellular domains of PAQR-1 and PAQR-2 play a regulatory role. These studies help understand how AdipoR1 and AdipoR2 regulate membrane composition in human cells, which is a vital function for us to thrive on diets that vary greatly in the types of fats that they contain.
|
|
| 4. |
- Devkota, Ranjan, et al.
(author)
-
The adiponectin receptor AdipoR2 and its Caenorhabditis elegans homolog PAQR-2 prevent membrane rigidification by exogenous saturated fatty acids.
- 2017
-
In: PLoS genetics. - : Public Library of Science (PLoS). - 1553-7404. ; 13:9
-
Journal article (peer-reviewed)abstract
- Dietary fatty acids can be incorporated directly into phospholipids. This poses a specific challenge to cellular membranes since their composition, hence properties, could greatly vary with different diets. That vast variations in diets are tolerated therefore implies the existence of regulatory mechanisms that monitor and regulate membrane compositions. Here we show that the adiponectin receptor AdipoR2, and its C. elegans homolog PAQR-2, are essential to counter the membrane rigidifying effects of exogenously provided saturated fatty acids. In particular, we use dietary supplements or mutated E. coli as food, together with direct measurements of membrane fluidity and composition, to show that diets containing a high ratio of saturated to monounsaturated fatty acids cause membrane rigidity and lethality in the paqr-2 mutant. We also show that mammalian cells in which AdipoR2 has been knocked-down by siRNA are unable to prevent the membrane-rigidifying effects of palmitic acid. We conclude that the PAQR-2 and AdipoR2 proteins share an evolutionarily conserved function that maintains membrane fluidity in the presence of exogenous saturated fatty acids.
|
|
| 5. |
- Laudette, Marion, et al.
(author)
-
Cardiomyocyte-specific PCSK9 deficiency compromises mitochondrial bioenergetics and heart function
- 2023
-
In: Cardiovascular Research. - : Oxford University Press (OUP). - 0008-6363 .- 1755-3245. ; 119:7, s. 1537-1552
-
Journal article (peer-reviewed)abstract
- Aims Pro-protein convertase subtilisin-kexin type 9 (PCSK9), which is expressed mainly in the liver and at low levels in the heart, regulates cholesterol levels by directing low-density lipoprotein receptors to degradation. Studies to determine the role of PCSK9 in the heart are complicated by the close link between cardiac function and systemic lipid metabolism. Here, we sought to elucidate the function of PCSK9 specifically in the heart by generating and analysing mice with cardiomyocyte-specific Pcsk9 deficiency (CMPcsk9−/− mice) and by silencing Pcsk9 acutely in a cell culture model of adult cardiomyocyte-like cells. Methods and results Mice with cardiomyocyte-specific deletion of Pcsk9 had reduced contractile capacity, impaired cardiac function, and left ventricular dilatation at 28 weeks of age and died prematurely. Transcriptomic analyses revealed alterations of signalling pathways linked to cardiomyopathy and energy metabolism in hearts from CM-Pcsk9−/− mice vs. wild-type littermates. In agreement, levels of genes and proteins involved in mitochondrial metabolism were reduced in CM-Pcsk9−/− hearts. By using a Seahorse flux analyser, we showed that mitochondrial but not glycolytic function was impaired in cardiomyocytes from CM-Pcsk9−/− mice. We further showed that assembly and activity of electron transport chain (ETC) complexes were altered in isolated mitochondria from CM-Pcsk9−/− mice. Circulating lipid levels were unchanged in CM-Pcsk9−/− mice, but the lipid composition of mitochondrial membranes was altered. In addition, cardiomyocytes from CM-Pcsk9−/− mice had an increased number of mitochondria–endoplasmic reticulum contacts and alterations in the morphology of cristae, the physical location of the ETC complexes. We also showed that acute Pcsk9 silencing in adult cardiomyocyte-like cells reduced the activity of ETC complexes and impaired mitochondrial metabolism. Conclusion PCSK9, despite its low expression in cardiomyocytes, contributes to cardiac metabolic function, and PCSK9 deficiency in cardiomyocytes is linked to cardiomyopathy, impaired heart function, and compromised energy production.
|
|
| 6. |
- Palmgren, Henrik, et al.
(author)
-
Elevated Adipocyte Membrane Phospholipid Saturation Does Not Compromise Insulin Signaling
- 2023
-
In: DIABETES. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 72:10, s. 1350-1363
-
Journal article (peer-reviewed)abstract
- Increased saturated fatty acid (SFA) levels in membrane phospholipids have been implicated in the development of metabolic disease. Here, we tested the hypothesis that increased SFA content in cell membranes negatively impacts adipocyte insulin signaling. Preadipocyte cell models with elevated SFA levels in phospholipids were generated by disrupting the ADIPOR2 locus, which resulted in a striking twofold increase in SFA-containing phosphatidylcholines and phosphatidylethanolamines, which persisted in differentiated adipocytes. Similar changes in phospholipid composition were observed in white adipose tissues isolated from the ADIPOR2-knockout mice. The SFA levels in phospholipids could be further increased by treating ADIPOR2-deficient cells with palmitic acid and resulted in reduced membrane fluidity and endoplasmic reticulum stress in mouse and human preadipocytes. Strikingly, increased SFA levels in differentiated adipocyte phospholipids had no effect on adipocyte gene expression or insulin signaling in vitro. Similarly, increased adipocyte phospholipid saturation did not impair white adipose tissue function in vivo, even in mice fed a high-saturated fat diet at thermoneutrality. We conclude that increasing SFA levels in adipocyte phospholipids is well tolerated and does not affect adipocyte insulin signaling in vitro and in vivo.
|
|
| 7. |
- Pilon, Marc, 1966, et al.
(author)
-
PAQR proteins and the evolution of a superpower: Eating all kinds of fats Animals rely on evolutionarily conserved membrane homeostasis proteins to compensate for dietary variation
- 2023
-
In: Bioessays. - 0265-9247. ; 45:9
-
Journal article (peer-reviewed)abstract
- Recently published work showed that members of the PAQR protein family are activated by cell membrane rigidity and contribute to our ability to eat a wide variety of diets. Cell membranes are primarily composed of phospholipids containing dietarily obtained fatty acids, which poses a challenge to membrane properties because diets can vary greatly in their fatty acid composition and could impart opposite properties to the cellular membranes. In particular, saturated fatty acids (SFAs) can pack tightly and form rigid membranes (like butter at room temperature) while unsaturated fatty acids (UFAs) form more fluid membranes (like vegetable oils). Proteins of the PAQR protein family, characterized by the presence of seven transmembrane domains and a cytosolic N-terminus, contribute to membrane homeostasis in bacteria, yeasts, and animals. These proteins respond to membrane rigidity by stimulating fatty acid desaturation and incorporation of UFAs into phospholipids and explain the ability of animals to thrive on diets with widely varied fat composition.
|
|
| 8. |
- Ruiz, Mario, 1984, et al.
(author)
-
A small molecule screen for paqr-2 suppressors identifies Tyloxapol as a membrane fluidizer for C. elegans and mammalian cells
- 2022
-
In: Biochimica Et Biophysica Acta-Biomembranes. - : Elsevier BV. - 0005-2736. ; 1864:9
-
Journal article (peer-reviewed)abstract
- Defects in cell membrane homeostasis are implicated in numerous disorders, including cancer, neurodegeneration and diabetes. There is therefore a need for a powerful model to study membrane homeostasis and to identify eventual therapeutic routes. The C. elegans gene paqr-2 encodes a homolog of the mammalian AdipoR1 and AdipoR2 proteins that, when mutated, causes a membrane homeostasis defect accompanied by multiple phenotypes such as intolerance to dietary saturated fatty acids, intolerance to cold and a characteristic tail tip morphology defect. We screened a compound library to identify molecules that can suppress the paqr-2 phenotypes. A single positive hit, Tyloxapol, was found that very effectively suppresses multiple paqr-2 phenotypes. Tyloxapol is a non-ionic detergent currently in use clinically as an expectorant. Importantly, we examined the potential of Tyloxapol as a fluidizer in human cells and found that it improves the viability and membrane fluidity of AdipoR2-deficient human cells challenged with palmitic acid, a membrane-rigidifying saturated fatty acid.
|
|
| 9. |
- Ruiz, Mario, 1984, et al.
(author)
-
AdipoR1 and AdipoR2 maintain membrane fluidity in most human cell types and independently of adiponectin
- 2019
-
In: Journal of Lipid Research. - 0022-2275. ; 60:5, s. 995-1004
-
Journal article (peer-reviewed)abstract
- The FA composition of phospholipids must be tightly regulated to maintain optimal cell membrane properties and compensate for a highly variable supply of dietary FAs. Previous studies have shown that AdipoR2 and its homologue PAQR-2 are important regulators of phospholipid FA composition in HEK293 cells and Caenorhabditiselegans, respectively. Here we show that both AdipoR1 and AdipoR2 are essential for sustaining desaturase expression and high levels of unsaturated FAs in membrane phospholipids of many human cell types, including primary human umbilical vein endothelial cells, and for preventing membrane rigidification in cells challenged with exogenous palmitate, a saturated FA. Three independent methods confirm the role of the AdipoRs as regulators of membrane composition and fluidity: fluorescence recovery after photobleaching, measurements of Laurdan dye generalized polarization, and mass spectrometry to determine the FA composition of phospholipids. Furthermore, we show that the AdipoRs can prevent lipotoxicity in the complete absence of adiponectin, their putative ligand. We propose that the primary cellular function of AdipoR1 and AdipoR2 is to maintain membrane fluidity in most human cell types and that adiponectin is not required for this function.
|
|
| 10. |
- Ruiz, Mario, 1984, et al.
(author)
-
AdipoR2 recruits protein interactors to promote fatty acid elongation and membrane fluidity
- 2023
-
In: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 299:6
-
Journal article (peer-reviewed)abstract
- The human AdipoR2 and its Caenorhabditis elegans homolog PAQR-2 are multipass plasma membrane proteins that protect cells against membrane rigidification. However, how AdipoR2 promotes membrane fluidity mechanistically is not clear. Using 13C-labeled fatty acids, we show that AdipoR2 can promote the elongation and incorporation of membrane-fluidizing polyunsaturated fatty acids into phospholipids. To elucidate the molecular basis of these activities, we performed immunoprecipitations of tagged AdipoR2 and PAQR-2 expressed in HEK293 cells or whole C. elegans, respectively, and identified coimmunoprecipitated proteins using mass spectrometry. We found that several of the evolutionarily conserved AdipoR2/PAQR-2 interactors are important for fatty acid elongation and incorporation into phospholipids. We experimentally verified some of these interactions, namely, with the dehydratase HACD3 that is essential for the third of four steps in long-chain fatty acid elongation and ACSL4 that is important for activation of unsaturated fatty acids and their channeling into phospholipids. We conclude that AdipoR2 and PAQR-2 can recruit protein interactors to promote the production and incorporation of unsaturated fatty acids into phospholipids.
|
|
