Restructuring of the plasma membrane upon damage by LC3-associated macropinocytosis

The plasma membrane shapes and protects the eukaryotic cell from its surroundings and is crucial for cell life. Although initial repair mechanisms to reseal injured membranes are well established, less is known about how cells restructure damaged membranes in the aftermath to restore homeostasis. Here, we show that cells respond to plasma membrane injury by activating proteins associated with macropinocytosis specifically at the damaged membrane. Subsequent to membrane resealing, cells form large macropinosomes originating from the repair site, which eventually become positive for autophagy-related LC3B protein. This process occurs independent of ULK1, ATG13, and WIPI2 but dependent on ATG7, p62, and Rubicon. Internalized macropinosomes shrink in the cytoplasm, likely by osmotic draining, and eventually fuse with lysosomes. We propose that a form of macropinocytosis coupled to noncanonical autophagy, which we term LC3-associated macropinocytosis (LAM) functions to remove damaged material from the plasma membrane and restore membrane integrity upon injury.

Download Full-text

Exocytosis of acid sphingomyelinase by wounded cells promotes endocytosis and plasma membrane repair

The Journal of Cell Biology ◽

10.1083/jcb.201003053 ◽

2010 ◽

Vol 189 (6) ◽

pp. 1027-1038 ◽

Cited By ~ 197

Author(s):

Christina Tam ◽

Vincent Idone ◽

Cecilia Devlin ◽

Maria Cecilia Fernandes ◽

Andrew Flannery ◽

...

Keyword(s):

Plasma Membrane ◽

Lysosomal Enzyme ◽

Membrane Integrity ◽

Acid Sphingomyelinase ◽

Membrane Repair ◽

Membrane Injury ◽

Cellular Survival ◽

Plasma Membrane Repair ◽

Membrane Resealing ◽

Niemann Pick

Rapid plasma membrane resealing is essential for cellular survival. Earlier studies showed that plasma membrane repair requires Ca2+-dependent exocytosis of lysosomes and a rapid form of endocytosis that removes membrane lesions. However, the functional relationship between lysosomal exocytosis and the rapid endocytosis that follows membrane injury is unknown. In this study, we show that the lysosomal enzyme acid sphingomyelinase (ASM) is released extracellularly when cells are wounded in the presence of Ca2+. ASM-deficient cells, including human cells from Niemann-Pick type A (NPA) patients, undergo lysosomal exocytosis after wounding but are defective in injury-dependent endocytosis and plasma membrane repair. Exogenously added recombinant human ASM restores endocytosis and resealing in ASM-depleted cells, suggesting that conversion of plasma membrane sphingomyelin to ceramide by this lysosomal enzyme promotes lesion internalization. These findings reveal a molecular mechanism for restoration of plasma membrane integrity through exocytosis of lysosomes and identify defective plasma membrane repair as a possible component of the severe pathology observed in NPA patients.

Download Full-text

Plasma membrane integrity in health and disease: significance and therapeutic potential

Cell Discovery ◽

10.1038/s41421-020-00233-2 ◽

2021 ◽

Vol 7 (1) ◽

Author(s):

Catarina Dias ◽

Jesper Nylandsted

Keyword(s):

Plasma Membrane ◽

Therapeutic Potential ◽

Calcium Influx ◽

Membrane Integrity ◽

Cell Types ◽

Physical Parameters ◽

Membrane Repair ◽

Plasma Membrane Integrity ◽

Repair Mechanisms ◽

And Function

AbstractMaintenance of plasma membrane integrity is essential for normal cell viability and function. Thus, robust membrane repair mechanisms have evolved to counteract the eminent threat of a torn plasma membrane. Different repair mechanisms and the bio-physical parameters required for efficient repair are now emerging from different research groups. However, less is known about when these mechanisms come into play. This review focuses on the existence of membrane disruptions and repair mechanisms in both physiological and pathological conditions, and across multiple cell types, albeit to different degrees. Fundamentally, irrespective of the source of membrane disruption, aberrant calcium influx is the common stimulus that activates the membrane repair response. Inadequate repair responses can tip the balance between physiology and pathology, highlighting the significance of plasma membrane integrity. For example, an over-activated repair response can promote cancer invasion, while the inability to efficiently repair membrane can drive neurodegeneration and muscular dystrophies. The interdisciplinary view explored here emphasises the widespread potential of targeting plasma membrane repair mechanisms for therapeutic purposes.

Download Full-text

Actin Cytoskeletal Dynamics in Single-Cell Wound Repair

International Journal of Molecular Sciences ◽

10.3390/ijms221910886 ◽

2021 ◽

Vol 22 (19) ◽

pp. 10886

Author(s):

Malene Laage Ebstrup ◽

Catarina Dias ◽

Anne Sofie Busk Heitmann ◽

Stine Lauritzen Sønder ◽

Jesper Nylandsted

Keyword(s):

Plasma Membrane ◽

Actin Cytoskeleton ◽

Single Cell ◽

Wound Repair ◽

Membrane Integrity ◽

Limited Attention ◽

Membrane Repair ◽

Cortical Actin ◽

Comprehensive Overview ◽

Repair Mechanisms

The plasma membrane protects the eukaryotic cell from its surroundings and is essential for cell viability; thus, it is crucial that membrane disruptions are repaired quickly to prevent immediate dyshomeostasis and cell death. Accordingly, cells have developed efficient repair mechanisms to rapidly reseal ruptures and reestablish membrane integrity. The cortical actin cytoskeleton plays an instrumental role in both plasma membrane resealing and restructuring in response to damage. Actin directly aids membrane repair or indirectly assists auxiliary repair mechanisms. Studies investigating single-cell wound repair have often focused on the recruitment and activation of specialized repair machinery, despite the undeniable need for rapid and dynamic cortical actin modulation; thus, the role of the cortical actin cytoskeleton during wound repair has received limited attention. This review aims to provide a comprehensive overview of membrane repair mechanisms directly or indirectly involving cortical actin cytoskeletal remodeling.

Download Full-text

Calciprotein particle-induced cytotoxicity via lysosomal dysfunction and altered cholesterol distribution in renal epithelial HK-2 cells

Scientific Reports ◽

10.1038/s41598-020-77308-3 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Rina Kunishige ◽

Mai Mizoguchi ◽

Asako Tsubouchi ◽

Kenjiro Hanaoka ◽

Yutaka Miura ◽

...

Keyword(s):

Plasma Membrane ◽

Serum Proteins ◽

Membrane Integrity ◽

Reactive Oxygen Species Generation ◽

Autophagic Flux ◽

Membrane Injury ◽

Renal Tubular Cells ◽

Calciprotein Particles ◽

Renal Proximal Tubular ◽

Renal Tubular

AbstractDietary phosphate overload induces chronic kidney disease (CKD), and calciprotein particles (CPPs), a form of nanoparticle comprising calcium phosphate and serum proteins, has been proposed to cause renal toxicity. However, the mechanism of CPP cytotoxicity in renal tubular cells is unknown. Here we show that in renal proximal tubular epithelial HK-2 cells, endocytosed CPPs accumulate in late endosomes/lysosomes (LELs) and increase their luminal pH by ~ 1.0 unit. This results in a decrease in lysosomal hydrolase activity and autophagic flux blockage without lysosomal rupture and reactive oxygen species generation. CPP treatment led to vulnerability to H2O2-induced oxidative stress and plasma membrane injury, probably because of autophagic flux blockage and decreased plasma membrane cholesterol, respectively. CPP-induced disruption of lysosomal homeostasis, autophagy flux and plasma membrane integrity might trigger a vicious cycle, leading to progressive nephron loss.

Download Full-text

Volume regulation and plasma membrane injury in aerobic, anaerobic, and ischemic myocardium in vitro. Effects of osmotic cell swelling on plasma membrane integrity.

Circulation Research ◽

10.1161/01.res.57.6.864 ◽

1985 ◽

Vol 57 (6) ◽

pp. 864-875 ◽

Cited By ~ 124

Author(s):

C Steenbergen ◽

M L Hill ◽

R B Jennings

Keyword(s):

Plasma Membrane ◽

Volume Regulation ◽

Membrane Integrity ◽

Ischemic Myocardium ◽

Cell Swelling ◽

Membrane Injury ◽

Plasma Membrane Integrity ◽

In Vitro Effects

Download Full-text

Cardiomyocyte damage control in heart failure and the role of the sarcolemma

Journal of Muscle Research and Cell Motility ◽

10.1007/s10974-019-09539-5 ◽

2019 ◽

Vol 40 (3-4) ◽

pp. 319-333 ◽

Cited By ~ 1

Author(s):

Ashraf Kitmitto ◽

Florence Baudoin ◽

Elizabeth J. Cartwright

Keyword(s):

Heart Failure ◽

Cell Death ◽

Plasma Membrane ◽

Cardiac Function ◽

Structural Integrity ◽

Damage Control ◽

Cardiac Contraction ◽

Membrane Injury ◽

Cellular Processes ◽

Repair Mechanisms

Abstract The cardiomyocyte plasma membrane, termed the sarcolemma, is fundamental for regulating a myriad of cellular processes. For example, the structural integrity of the cardiomyocyte sarcolemma is essential for mediating cardiac contraction by forming microdomains such as the t-tubular network, caveolae and the intercalated disc. Significantly, remodelling of these sarcolemma microdomains is a key feature in the development and progression of heart failure (HF). However, despite extensive characterisation of the associated molecular and ultrastructural events there is a lack of clarity surrounding the mechanisms driving adverse morphological rearrangements. The sarcolemma also provides protection, and is the cell’s first line of defence, against external stresses such as oxygen and nutrient deprivation, inflammation and oxidative stress with a loss of sarcolemma viability shown to be a key step in cell death via necrosis. Significantly, cumulative cell death is also a feature of HF, and is linked to disease progression and loss of cardiac function. Herein, we will review the link between structural and molecular remodelling of the sarcolemma associated with the progression of HF, specifically considering the evidence for: (i) Whether intrinsic, evolutionary conserved, plasma membrane injury-repair mechanisms are in operation in the heart, and (ii) if deficits in key ‘wound-healing’ proteins (annexins, dysferlin, EHD2 and MG53) may play a yet to be fully appreciated role in triggering sarcolemma microdomain remodelling and/or necrosis. Cardiomyocytes are terminally differentiated with very limited regenerative capability and therefore preserving cell viability and cardiac function is crucially important. This review presents a novel perspective on sarcolemma remodelling by considering whether targeting proteins that regulate sarcolemma injury-repair may hold promise for developing new strategies to attenuate HF progression.

Download Full-text

Neuronal Plasma Membrane Integrity is Transiently Disturbed by Traumatic Loading

Neuroscience Insights ◽

10.1177/2633105520946090 ◽

2020 ◽

Vol 15 ◽

pp. 263310552094609

Author(s):

Gustavo R Prado ◽

Michelle C LaPlaca

Keyword(s):

Plasma Membrane ◽

Neuronal Response ◽

Membrane Integrity ◽

High Rate ◽

Membrane Disruption ◽

Neuronal Membrane ◽

Neuronal Cultures ◽

Plasma Membrane Integrity ◽

Intracellular Ca ◽

Membrane Resealing

The acute response of neurons subjected to traumatic loading involves plasma membrane disruption, yet the mechanical tolerance for membrane compromise, time course, and mechanisms for resealing are not well understood. We have used an in vitro traumatic neuronal injury model to investigate plasma membrane integrity immediately following a high-rate shear injury. Cell-impermeant fluorescent molecules were added to cortical neuronal cultures prior to insult to assess membrane integrity. The percentage of cells containing the permeability marker was dependent on the molecular size of the marker, as smaller molecules gained access to a higher percentage of cells than larger ones. Permeability increases were positively correlated with insult loading rate. Membrane disruption was transient, evidenced by a membrane resealing within the first minute after the insult. In addition, chelation of either extracellular Ca2+ or intracellular Ca2+ limited membrane resealing. However, injury following chelation of both extracellular and intracellular Ca2+ caused diminished permeability as well as a greater resealing ability compared to chelation of extracellular or intracellular Ca2+ alone. Treatment of neuronal cultures with jasplakinolide, which stabilizes filamentous actin, reduced permeability increases, while latrunculin-B, an actin depolymerizing agent, both reduced the increase in plasma membrane permeability and promoted resealing. This study gives insight into the dynamics of neuronal membrane disruption and subsequent resealing, which was found to be calcium dependent and involve actin in a role that differs from non-neuronal cells. Taken together, these data will lead to a better understanding of the acute neuronal response to traumatic loading.

Download Full-text

Short-term transcriptomic response to plasma membrane injury

Scientific Reports ◽

10.1038/s41598-021-98420-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Swantje Christin Häger ◽

Catarina Dias ◽

Stine Lauritzen Sønder ◽

André Vidas Olsen ◽

Isabelle da Piedade ◽

...

Keyword(s):

Plasma Membrane ◽

Membrane Repair ◽

Short Term ◽

Post Injury ◽

Membrane Injury ◽

Transcriptomic Response ◽

A Genome ◽

Ionic Detergent ◽

Membrane Resealing ◽

Regeneration Phase

AbstractPlasma membrane repair mechanisms are activated within seconds post-injury to promote rapid membrane resealing in eukaryotic cells and prevent cell death. However, less is known about the regeneration phase that follows and how cells respond to injury in the short-term. Here, we provide a genome-wide study into the mRNA expression profile of MCF-7 breast cancer cells exposed to injury by digitonin, a mild non-ionic detergent that permeabilizes the plasma membrane. We focused on the early transcriptional signature and found a time-dependent increase in the number of differentially expressed (> twofold, P < 0.05) genes (34, 114 and 236 genes at 20-, 40- and 60-min post-injury, respectively). Pathway analysis highlighted a robust and gradual three-part transcriptional response: (1) prompt activation of immediate-early response genes, (2) activation of specific MAPK cascades and (3) induction of inflammatory and immune pathways. Therefore, plasma membrane injury triggers a rapid and strong stress and immunogenic response. Our meta-analysis suggests that this is a conserved transcriptome response to plasma membrane injury across different cell and injury types. Taken together, our study shows that injury has profound effects on the transcriptome of wounded cells in the regeneration phase (subsequent to membrane resealing), which is likely to influence cellular status and has been previously overlooked.

Download Full-text