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Phenothiazines alter plasma membrane properties and sensitize cancer cells to injury by inhibiting annexin-mediated repair.

ABSTRACT: Repair of damaged plasma membrane in eukaryotic cells is largely dependent on the binding of annexin repair proteins to phospholipids. Changing the biophysical properties of the plasma membrane may provide means to compromise annexin-mediated repair and sensitize cells to injury. Since, cancer cells experience heightened membrane stress and are more dependent on efficient plasma membrane repair, inhibiting repair may provide approaches to sensitize cancer cells to plasma membrane damage and cell death. Here, we show that derivatives of phenothiazines, which have widespread use in the fields of psychiatry and allergy treatment, strongly sensitize cancer cells to mechanical-, chemical-, and heat-induced injury by inhibiting annexin-mediated plasma membrane repair. Using a combination of cell biology, biophysics, and computer simulations, we show that trifluoperazine acts by thinning the membrane bilayer, making it more fragile and prone to ruptures. Secondly, it decreases annexin binding by compromising the lateral diffusion of phosphatidylserine, inhibiting the ability of annexins to curve and shape membranes, which is essential for their function in plasma membrane repair. Our results reveal a novel avenue to target cancer cells by compromising plasma membrane repair in combination with noninvasive approaches that induce membrane injuries.

SUBMITTER: Heitmann ASB

PROVIDER: S-EPMC8363839 | biostudies-literature |

REPOSITORIES: biostudies-literature

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An actin-dependent annexin complex mediates plasma membrane repair in muscle.

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| S-EPMC6819108 | biostudies-literature

Mycobacterium tuberculosis evades macrophage defenses by inhibiting plasma membrane repair.

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| S-EPMC2730354 | biostudies-literature

Annexin A2 Mediates Dysferlin Accumulation and Muscle Cell Membrane Repair.

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Annexin A4 and A6 induce membrane curvature and constriction during cell membrane repair.

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| S-EPMC5696365 | biostudies-literature

Annexin-A5 assembled into two-dimensional arrays promotes cell membrane repair.

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| S-EPMC3104517 | biostudies-literature

Evidence for annexin A6-dependent plasma membrane remodelling of lipid domains.

Project description:Background and purposeAnnexin A6 (AnxA6) is a calcium-dependent phospholipid-binding protein that can be recruited to the plasma membrane to function as a scaffolding protein to regulate signal complex formation, endo- and exocytic pathways as well as distribution of cellular cholesterol. Here, we have investigated how AnxA6 influences the membrane order.Experimental approachWe used Laurdan and di-4-ANEPPDHQ staining in (i) artificial membranes; (ii) live cells to investigate membrane packing and ordered lipid phases; and (iii) a super-resolution imaging (photoactivated localization microscopy, PALM) and Ripley's K second-order point pattern analysis approach to assess how AnxA6 regulates plasma membrane order domains and protein clustering.Key resultsIn artificial membranes, purified AnxA6 induced a global increase in membrane order. However, confocal microscopy using di-4-ANEPPDHQ in live cells showed that cells expressing AnxA6, which reduces plasma membrane cholesterol levels and modifies the actin cytoskeleton meshwork, displayed a decrease in membrane order (?15 and 30% in A431 and MEF cells respectively). PALM data from Lck10 and Src15 membrane raft/non-raft markers revealed that AnxA6 expression induced clustering of both raft and non-raft markers. Altered clustering of Lck10 and Src15 in cells expressing AnxA6 was also observed after cholesterol extraction with methyl-?-cyclodextrin or actin cytoskeleton disruption with latrunculin B.Conclusions and implicationsAnxA6-induced plasma membrane remodelling indicated that elevated AnxA6 expression decreased membrane order through the regulation of cellular cholesterol homeostasis and the actin cytoskeleton. This study provides the first evidence from live cells that support current models of annexins as membrane organizers.

| S-EPMC4376448 | biostudies-literature

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| S-EPMC10719157 | biostudies-literature

Quantitative analysis of self-association and mobility of annexin A4 at the plasma membrane.

Project description:Annexins, found in most eukaryotic species, are cytosolic proteins that are able to bind negatively-charged phospholipids in a calcium-dependent manner. Annexin A4 (AnxA4) has been implicated in diverse cellular processes, including the regulation of exocytosis and ion-transport; however, its precise mechanistic role is not fully understood. AnxA4 has been shown to aggregate on lipid layers upon Ca(2+) binding in vitro, a characteristic that may be critical for its function. We have utilized advanced fluorescence microscopy to discern details on the mobility and self-assembly of AnxA4 after Ca(2+) influx at the plasma membrane in living cells. Total internal reflection microscopy in combination with Förster resonance energy transfer reveals that there is a delay between initial plasma membrane binding and the beginning of self-assembly and this process continues after the cytoplasmic pool has completely relocated. Number-and-brightness analysis suggests that the predominant membrane bound mobile form of the protein is trimeric. There also exists a pool of AnxA4 that forms highly immobile aggregates at the membrane. Fluorescence recovery after photobleaching suggests that the relative proportion of these two forms varies and is correlated with membrane morphology.

| S-EPMC3647157 | biostudies-literature

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