Cellular Functions Research Articles

Critical and differential roles of eIF4A1 and eIF4A2 in B-cell development and function.

Eukaryotic initiation factor 4 A (eIF4A) plays critical roles during translation initiation of cellular mRNAs by forming the cap-binding eIF4F complex, recruiting the 40S small ribosome subunit, and scanning the 5' untranslated region (5' UTR) for the start codon. eIF4A1 and eIF4A2, two isoforms of eIF4A, are highly conserved and exchange freely within eIF4F complexes. The understanding of their biological and molecular functions remains incomplete if not fragmentary. In this study, we showed that eIF4A1 and eIF4A2 exhibit different expression patterns during B-cell development and activation. Mouse genetic analyses showed that they play critical but differential roles during B-cell development and humoral immune responses. While eIF4A1 controls global protein synthesis, eIF4A2 regulates the biogenesis of 18S ribosomal RNA and the 40S ribosome subunit. This study demonstrates the distinct cellular and molecular functions of eIF4A1 and eIF4A2 and reveals a new role of eIF4A2 in controlling 40S ribosome biogenesis.

Differential vegfc expression dictates lymphatic response during zebrafish heart development and regeneration.

Vascular endothelial growth factor C (Vegfc) is crucial for lymphatic and blood vessel development, yet its cellular sources and specific functions in heart development remain unclear. To address this, we created a vegfc reporter and an inducible overexpression line in zebrafish. We found vegfc expression in large coronary arteries, circulating thrombocytes, cardiac adipocytes, and outflow tract smooth muscle cells. Notably, while coronary lymphangiogenesis aligns with Vegfc-expressing arteries in juveniles, it occurs only after coronary artery formation. Vegfc overexpression induced ectopic lymphatics on the ventricular surface prior to arterial formation, indicating that Vegfc abundance, rather than arterial presence, drives lymphatic development. However, this overexpression did not affect coronary artery coverage, suggesting a specific role for Vegfc in lymphatic, rather than arterial, development. Thrombocytes emerged as the initial Vegfc source during inflammation following heart injuries, transitioning to endocardial and myocardial expression during regeneration. Lower Vegfc levels in the amputation model corresponded with a lack of lymphatic expansion. Importantly, Vegfc overexpression enhanced lymphatic expansion and promoted scar resolution without affecting cardiomyocyte proliferation, highlighting its role in regulating lymphangiogenesis and promoting heart regeneration.

Mitochondrial elongation impairs breast cancer metastasis.

Mitochondrial dynamics orchestrate many essential cellular functions, including metabolism, which is instrumental in promoting cancer growth and metastatic progression. However, how mitochondrial dynamics influences metastatic progression remains poorly understood. Here, we show that breast cancer cells with low metastatic potential exhibit a more fused mitochondrial network compared to highly metastatic cells. To study the impact of mitochondrial dynamics on metastasis, we promoted mitochondrial elongation in metastatic breast cancer cells by individual genetic deletion of three key regulators of mitochondrial fission (Drp1, Fis1, Mff) or by pharmacological intervention with leflunomide. Omics analyses revealed that mitochondrial elongation causes substantial alterations in metabolic pathways and processes related to cell adhesion. In vivo, enhanced mitochondrial elongation by loss of mitochondrial fission mediators or treatment with leflunomide notably reduced metastasis formation. Furthermore, the transcriptomic signature associated with elongated mitochondria correlated with improved clinical outcome in patients with breast cancer. Overall, our findings highlight mitochondrial dynamics as a potential therapeutic target in breast cancer.

Shear stress effects on epididymal epithelial cell via primary cilia mechanosensory signaling.

Shear stress, resulting from fluid flow, is a fundamental mechanical stimulus affecting various cellular functions. The epididymis, essential for sperm maturation, offers a compelling model to study the effects of shear stress on cellular behavior. This organ undergoes extensive proliferation and differentiation until puberty, achieving full functionality as spermatozoa commence their post-testicular maturation. Although the mechanical tension exerted by testicular fluid is hypothesized to drive epithelial proliferation and differentiation, the precise mechanisms remain unclear. Here we assessed whether the responsiveness of the epididymal cells to shear stress depends on functional primary cilia by combining microfluidic strategies on immortalized epididymal cells, calcium signaling assays, and high-throughput gene expression analysis. We identified 97 genes overexpressed in response to shear stress, including early growth response (Egr) 2/3, cellular communication network factor (Ccn) 1/2, and Fos proto-oncogene (Fos). While shear stress triggered a rapid increase of intracellular Ca2+, this response was abrogated following the impairment of primary ciliogenesis through pharmacological and siRNA approaches. Overall, our findings provide valuable insights into how mechanical forces influence the development of the male reproductive system, a requisite to sperm maturation.

Exploring the Role of Rho Kinase Enzyme in the Management of Metabolic Syndrome

Rho kinase [ROCK] enzymes are increasingly recognized for their central role in the pathogenesis of metabolic syndrome [MetS], a cluster of conditions that includes insulin resistance, hypertension, obesity, and dyslipidemia. ROCKs are serine/threonine kinases involved in the regulation of various cellular functions, including smooth muscle contraction, actin cytoskeleton organization, and gene expression. These enzymes are critically implicated in the cardiovascular and metabolic abnormalities that characterize MetS. Elevated ROCK activity has been observed in individuals with MetS, contributing to several pathogenic processes such as endothelial dysfunction, vascular inflammation, oxidative stress, and increased vascular smooth muscle contraction. These mechanisms are key drivers of hypertension and atherosclerosis, which are common complications associated with MetS. Moreover, ROCKs influence adipocyte differentiation and lipid metabolism, linking them directly to obesity and insulin resistance, two core components of the syndrome. The inhibition of ROCKs has emerged as a promising therapeutic strategy for managing MetS. Pharmacological ROCK inhibitors have shown the potential to improve insulin sensitivity, lower blood pressure, and reduce vascular inflammation and remodelling. In addition, by targeting the multiple pathways involved in the development and progression of MetS, ROCK inhibitors offer a comprehensive approach to treatment that addresses the syndrome's multifactorial nature. This therapeutic strategy not only mitigates the metabolic and cardiovascular components of the syndrome but also lowers the risk of associated complications, such as cardiovascular disease and stroke. This review concluded that interrupted Rho kinase activity contributes to the development of MetS in all its manifestations. Overall, these side effects diminish the Rho-kinase method's promise as a novel and significant treatment component.

Inhibition of SGLT2 protects podocytes in diabetic kidney disease by rebalancing mitochondria-associated endoplasmic reticulum membranes

BackgroundSodium-glucose cotransporter 2 (SGLT2) inhibitors have changed the therapeutic landscape for diabetic kidney disease (DKD) patients, but their underlying mechanisms are complicated and not fully understood. Mitochondria-associated endoplasmic reticulum membranes (MAMs), the dynamic contact sites between mitochondria and the endoplasmic reticulum (ER), serve as intracellular platforms important for regulating cellular fate and function. This study explored the roles and mechanisms of SGLT2 inhibitors in regulating MAMs formation in diabetic podocytes.MethodsWe assessed MAMs formation in podocytes from DKD patients’ renal biopsy samples and induced an increase in MAMs formation in cultured human podocytes by transfecting OMM-ER linker plasmid to investigate the effects of MAMs imbalance on podocyte injury. Empagliflozin-treated diabetic mice and podocyte-specific SGLT2 knockout diabetic mice (diabetic states were induced by streptozotocin and a high-fat diet), empagliflozin-treated podocytes, SGLT2-downregulated podocytes, and SGLT2-overexpressing podocytes were used to investigate the effects and mechanisms of SGLT2 inhibitors on MAMs formation in diabetic podocytes.ResultsMAMs were increased in podocytes and were associated with renal dysfunction in DKD patients. Increased MAMs aggravated HG-induced podocyte injury. The expression of SGLT2 was increased in diabetic podocytes. In addition, empagliflozin-treatment and podocyte-specific SGLT2 knockout attenuated MAMs formation and podocyte injury in diabetic mice. Empagliflozin treatment and SGLT2 knockdown decreased podocyte MAMs formation by activating the AMP-activated protein kinase (AMPK) pathway, while SGLT2 overexpression had the opposite effect.ConclusionsInhibition of SGLT2 attenuates MAMs imbalance in diabetic podocytes by activating the AMPK pathway. This study expands our knowledge of the roles of SGLT2 inhibitors in improving DKD podocyte injury and provides new insights into DKD treatment.

PKA regulation of neuronal function requires the dissociation of catalytic subunits from regulatory subunits

Protein kinase A (PKA) plays essential roles in diverse cellular functions. However, the spatiotemporal dynamics of endogenous PKA upon activation remain debated. The classical model predicts that PKA catalytic subunits dissociate from regulatory subunits in the presence of cAMP, whereas a second model proposes that catalytic subunits remain associated with regulatory subunits following physiological activation. Here, we report that different PKA subtypes, as defined by the regulatory subunit, exhibit distinct subcellular localization at rest in CA1 neurons of cultured hippocampal slices. Nevertheless, when all tested PKA subtypes are activated by norepinephrine, presumably via the β-adrenergic receptor, catalytic subunits translocate to dendritic spines but regulatory subunits remain unmoved. These differential spatial dynamics between the subunits indicate that at least a significant fraction of PKA dissociates. Furthermore, PKA-dependent regulation of synaptic plasticity and transmission can be supported only by wildtype, dissociable PKA, but not by inseparable PKA. These results indicate that endogenous PKA regulatory and catalytic subunits dissociate to achieve PKA function in neurons.

PKA regulation of neuronal function requires the dissociation of catalytic subunits from regulatory subunits.

Protein kinase A (PKA) plays essential roles in diverse cellular functions. However, the spatiotemporal dynamics of endogenous PKA upon activation remain debated. The classical model predicts that PKA catalytic subunits dissociate from regulatory subunits in the presence of cAMP, whereas a second model proposes that catalytic subunits remain associated with regulatory subunits following physiological activation. Here, we report that different PKA subtypes, as defined by the regulatory subunit, exhibit distinct subcellular localization at rest in CA1 neurons of cultured hippocampal slices. Nevertheless, when all tested PKA subtypes are activated by norepinephrine, presumably via the β-adrenergic receptor, catalytic subunits translocate to dendritic spines but regulatory subunits remain unmoved. These differential spatial dynamics between the subunits indicate that at least a significant fraction of PKA dissociates. Furthermore, PKA-dependent regulation of synaptic plasticity and transmission can be supported only by wildtype, dissociable PKA, but not by inseparable PKA. These results indicate that endogenous PKA regulatory and catalytic subunits dissociate to achieve PKA function in neurons.

PRDX6 contributes to selenocysteine metabolism and ferroptosis resistance.

Selenocysteine (Sec) metabolism is crucial for cellular function and ferroptosis prevention and begins with the uptake of the Sec carrier, selenoprotein P (SELENOP). Following uptake, Sec released from SELENOP is metabolized via selenocysteine lyase (SCLY), producing selenide, a substrate for selenophosphate synthetase 2 (SEPHS2), which provides the essential selenium donor, selenophosphate (H2SePO3-), for the biosynthesis of the Sec-tRNA. Here, we discovered an alternative pathway in Sec metabolism mediated by peroxiredoxin 6 (PRDX6), independent of SCLY. Mechanistically, we demonstrate that PRDX6 can readily react with selenide and interact with SEPHS2, potentially acting as a selenium delivery system. Moreover, we demonstrate the functional significance of this alternative route in human cancer cells, revealing a notable association between elevated expression of PRDX6 and human MYCN-amplified neuroblastoma subtype. Our study sheds light on a previously unrecognized aspect of Sec metabolism and its implications in ferroptosis, offering further possibilities for therapeutic exploitation.

Hydrogen Sulfide‐Triggered Artificial DNAzyme Switches for Precise Manipulation of Cellular Functions

AbstractThe development of synthetic molecular tools responsive to biological cues is crucial for advancing targeted cellular regulation. A significant challenge is the regulation of cellular processes in response to gaseous signaling molecules such as hydrogen sulfide (H2S). To address this, we present the design of Gas signaling molecule‐Responsive Artificial DNAzyme‐based Switches (GRAS) to manipulate cellular functions via H2S‐sensitive synthetic DNAzymes. By incorporating stimuli‐responsive moieties to the phosphorothioate backbone, DNAzymes are strategically designed with H2S‐responsive azide groups at cofactor binding locations within the catalytic core region. These modifications enable their activation through H2S‐reducing decaging, thereby initiating substrate cleavage activity. Our approach allows for the flexible customization of various DNAzymes to regulate distinct cellular processes in diverse scenarios. Intracellularly, the enzymatic activity of GRAS promotes H2S‐induced cleavage of specific mRNA sequences, enabling targeted gene silencing and inducing apoptosis in cancer cells. Moreover, integrating GRAS with dynamic DNA assembly allows for grafting these functional switches onto cell surface receptors, facilitating H2S‐triggered receptor dimerization. This extracellular activation transmits signals intracellularly to regulate cellular behaviors such as migration and proliferation. Collectively, synthetic switches are capable of rewiring cellular functions in response to gaseous cues, offering a promising avenue for advanced targeted cellular engineering.

Impact of Chronic Moderate Psychological Stress on Skin Aging: Exploratory Clinical Study and Cellular Functioning.

Skin is continuously exposed to environmental external and internal factors, including psychological stress (PS). PS has been reported to trigger different dermatoses such as psoriasis, atopic dermatitis, vitiligo, alopecia areata, and acne through the release of cortisol and epinephrine. To clinically explore PS-induced measurable skin aging signs in subjects with moderate versus mild chronic PS, and to investigate the effect of chronic PS on DNA damage at cellular level. In vitro stress tests with cortisol and epinephrine, and with cortisol only on extracellular matrix (ECM) synthesis, as well as on normal human skin fibroblast and keratinocyte functioning, including skin barrier and wound healing were performed. Moderately stressed subjects in the context of the clinical study had a significantly decreased antioxidant potential and impacted skin barrier integrity, as well as significantly increased signs of microrelief alterations (skin texture and fine lines) reaching an increased severity of about 32.9%. At a cellular level, DNA integrity, ECM synthesis, wound healing, and skin barrier parameters were impacted by increased stress hormone levels. The clinical exploratory studies presented herewith, as well as the study of cell functioning under stress, have provided evidence that chronic PS significantly affects skin homeostasis and triggers skin aging.

Mechanisms and perspectives of B vitamins associated one carbon metabolism on colorectal cancer risk

Colorectal cancer (CRC) represents a significant global health challenge as a common malignancy of the digestive tract. The involvement of B vitamins-specifically folic acid (B9), riboflavin (B2), pyridoxine (B6), and cobalamin (B12)-is crucial in metabolic processes by mediating the transfer of one-carbon (1C) units, which plays a fundamental role in cellular functions and tumor growth. 1C metabolism is involved in synthesis of proteins, lipids, nucleic acids, and other cofactors. 1C metabolism, intertwined with the metabolism of other nutrients, forms complex pathways where B vitamins act as precursors or coenzymes, influencing the production of various intermediates. These vitamins, as essential nutrients, are implicated to varying the pathogenesis and progression of colorectal cancer such as epigenetics. Furthermore, 1C metabolism affects tumor cell fate through multiple aspects including nucleotide synthesis, redox homeostasis, and the interaction with gut microbiota. Given these roles, understanding and monitoring B vitamin levels and their metabolic pathways are essential for colorectal cancer prevention and management. This approach not only helps in reducing tumor-related mortality but also opens new avenues for research into CRC mechanisms and potential therapeutic strategies.

A PSA SNP associates with cellular function and clinical outcome in men with prostate cancer

Genetic variation at the 19q13.3 KLK locus is linked with prostate cancer susceptibility in men. The non-synonymous KLK3 single nucleotide polymorphism (SNP), rs17632542 (c.536 T > C; Ile163Thr-substitution in PSA) is associated with reduced prostate cancer risk, however, the functional relevance is unknown. Here, we identify that the SNP variant-induced change in PSA biochemical activity mediates prostate cancer pathogenesis. The ‘Thr’ PSA variant leads to small subcutaneous tumours, supporting reduced prostate cancer risk. However, ‘Thr’ PSA also displays higher metastatic potential with pronounced osteolytic activity in an experimental metastasis in-vivo model. Biochemical characterisation of this PSA variant demonstrates markedly reduced proteolytic activity that correlates with differences in in-vivo tumour burden. The SNP is associated with increased risk for aggressive disease and prostate cancer-specific mortality in three independent cohorts, highlighting its critical function in mediating metastasis. Carriers of this SNP allele have reduced serum total PSA and a higher free/total PSA ratio that could contribute to late biopsy decisions and delay in diagnosis. Our results provide a molecular explanation for the prominent 19q13.3 KLK locus, rs17632542 SNP, association with a spectrum of prostate cancer clinical outcomes.

Generative adversarial network model to classify human induced pluripotent stem cell-cardiomyocytes based on maturation level

Our study develops a generative adversarial network (GAN)-based method that generates faithful synthetic image data of human cardiomyocytes at varying stages in their maturation process, as a tool to significantly enhance the classification accuracy of cells and ultimately assist the throughput of computational analysis of cellular structure and functions. Human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) were cultured on micropatterned collagen coated hydrogels of physiological stiffnesses to facilitate maturation and optical measurements were performed for their structural and functional analyses. Control groups were cultured on collagen coated glass well plates. These image recordings were used as the real data to train the GAN model. The results show the GAN approach is able to replicate true features from the real data, and inclusion of such synthetic data significantly improves the classification accuracy compared to usage of only real experimental data that is often limited in scale and diversity. The proposed model outperformed four conventional machine learning algorithms with respect to improved data generalization ability and data classification by incorporating synthetic data. This work demonstrates the importance of integrating synthetic data in situations where there are limited sample sizes and thus, effectively addresses the challenges imposed by data availability.

The unique role of vimentin in the intermediate filament proteins family

Intermediate filaments (IFs), being traditionally the least studied component of the cytoskeleton, have begun to receive more attention in recent years. IFs are found in different cell types and are specific to them. Accumulated data have shifted the paradigm about the role of IFs as structures that merely provide mechanical strength to the cell. In addition to this role, IFs have been shown to participate in maintaining cell shape and strengthening cell adhesion. The data have also been obtained that point out to the role of IFs in a number of other biological processes, including organization of microtubules and microfilaments, regulation of nuclear structure and activity, cell cycle control, and regulation of signal transduction pathways. They are also actively involved in the regulation of several aspects of intracellular transport. Among the intermediate filament proteins, vimentin is of particular interest for researchers. Vimentin has been shown to be associated with a range of diseases, including cancer, cataracts, Crohn’s disease, rheumatoid arthritis, and HIV. In this review, we focus almost exclusively on vimentin and the currently known functions of vimentin intermediate filaments (VIFs). This is due to the structural features of vimentin, biological functions of its domains, and its involvement in the regulation of a wide range of basic cellular functions, and its role in the development of human diseases. Particular attention in the review will be paid to comparing the role of VIFs with the role of intermediate filaments consisting of other proteins in cell physiology.

Copy number variation heterogeneity reveals biological inconsistency in hierarchical cancer classifications

Cancers are heterogeneous diseases with unifying features of abnormal and consuming cell growth, where the deregulation of normal cellular functions is initiated by the accumulation of genomic mutations in cells of - potentially - any organ. At diagnosis malignancies typically present with patterns of somatic genome variants on diverse levels of heterogeneity. Among the different types of genomic alterations, copy number variants (CNV) represent a distinct, near-ubiquitous class of structural variants. Cancer classifications are foundational for patient care and oncology research. Terminologies such as the National Cancer Institute Thesaurus provide large sets of hierarchical cancer classification vocabularies and promote data interoperability and ontology-driven computational analysis. To find out how categorical classifications correspond to genomic observations, we conducted a meta-analysis of inter-sample genomic heterogeneity for classification hierarchies on CNV profiles from 97,142 individual samples across 512 cancer entities, and evaluated recurring CNV signatures across diagnostic subsets. Our results highlight specific biological mechanisms across cancer entities with the potential for improvement of patient stratification and future enhancement of cancer classification systems and provide some indications for cooperative genomic events across distinct clinical entities.

Effect of Different Zn Concentrations on Crop Growth and Root Nodulation in Yard-Long Bean (Vigna unguiculata L.)

An experiment was conducted to study the effect of Zinc (Zn) as a micronutrient on growth and nodulation in Vigna unguiculata L. The experiment was carried out as a pot experiment in the Crop Farm, Faculty of Agriculture, Palachcholai, Eastern University, Sri Lanka from August to November 2023. The experiment was laid out in Complete Randomized Design with six treatments and four replicates. The treatments are T1 (Control), T2 (50 mg ZnSO4/kg soil), T3 (100 mg ZnSO4/kg soil), T4 (150 mg ZnSO4/kg soil), T5 (200 mg ZnSO4/kg soil) & T6 (250 mg ZnSO4/kg soil). The experiment used source of Zinc in the form of Zinc Sulphate (ZnSO4,5H2O) which yard-long bean plants can grow. The experiment results showed that, T2 treatment, the soil treated with 50 mg ZnSO4/kg soil significantly increase the Plant height, Chlorophyll content, Leaf area, Fresh weight of shoot & root, Dry weight of shoot & root, Number of nodules, Effective nodule percentage, as well as Soil respiration compared to the other treatments. However, beyond the level of 200 mg ZnSO4/kg soil, reduces the growth & nodulation of Vigna unguiculata L. and this shows that the level 200 mg ZnSO4/kg soil and beyond that level were the toxic level to the Vigna unguiculata L. Accordingly, the treatment T2 (50 mg ZnSO4/kg soil) was at the micronutrient level of Zn concentration which involved in many key cellular functions and show the best positive effect on growth and nodulation in Vigna unguiculata L. The results clearly indicate that, Zn can be use as a micronutrient up to a trace level in inducing plant growth and nodulation. However, at higher levels of Zn may act as heavy metal and cause phyto-toxicity in plants.

Divergent Molecular Responses to Heavy Water in Arabidopsis thaliana Compared to Bacteria and Yeast

Heavy water (D2O) is scarce in nature, and despite its physical similarity to water, D2O disrupts cellular function due to the isotope effect. While microbes can survive in nearly pure D2O, eukaryotes such as Arabidopsis thaliana are more sensitive and are unable to survive higher concentrations of D2O. To explore the underlying molecular mechanisms for these differences, we conducted a comparative proteomic analysis of E. coli, S. cerevisiae, and Arabidopsis after 180 min of growth in a D2O-supplemented media. Shared adaptive mechanisms across these species were identified, including changes in ribosomal protein abundances, accumulation of chaperones, and altered metabolism of polyamines and amino acids. However, Arabidopsis exhibited unique vulnerabilities, such as a muted stress response, lack of rapid activation of reactive oxygen species metabolism, and depletion of stress phytohormone abscisic acid signaling components. Experiments with mutants show that modulating the HSP70 pool composition may promote D2O resilience. Additionally, Arabidopsis rapidly incorporated deuterium into sucrose, indicating that photosynthesis facilitates deuterium intake. These findings provide valuable insights into the molecular mechanisms that dictate differential tolerance to D2O across species and lay the groundwork for further studies on the biological effects of uncommon isotopes, with potential implications for biotechnology and environmental science.

KRAS inhibitors in drug resistance and potential for combination therapy.

Kirsten Rat Sarcoma (KRAS) is a potent target for cancer therapy because it acts as a signaling hub, engaging in various signaling pathways and regulating a number of cellular functions like cell differentiation, proliferation, and survival. Recently, an emergency approval from the US-FDA has been issued for KRASG12C inhibitors (sotorasib and adagrasib) for metastatic lung cancer treatment. However, clinical studies on covalent KRASG12C inhibitors have rapidly confronted resistance in patients. Many methods are being assessed to overcome this resistance, along with various combinatorial clinical studies that are in process. Moreover, because KRASG12D and KRASG12V are more common than KRASG12C, focus must be placed on the therapeutic strategies for this type of patient, along with sustained efforts in research on these targets. In the present review, we try to focus on various strategies to overcome rapid resistance through the use of combinational treatments to improve the activity of KRASG12C inhibitors.

The transmembrane domain of the desmosomal cadherin desmoglein-1 governs lipid raft association to promote desmosome adhesive strength.

Cholesterol- and sphingolipid-enriched domains called lipid rafts are hypothesized to selectively coordinate protein complex assembly within the plasma membrane to regulate cellular functions. Desmosomes are mechanically resilient adhesive junctions that associate with lipid raft membrane domains, yet the mechanisms directing raft association of the desmosomal proteins, particularly the transmembrane desmosomal cadherins, are poorly understood. We identified the desmoglein-1 (DSG1) transmembrane domain (TMD) as a key determinant of desmoglein lipid raft association and designed a panel of DSG1TMD variants to assess the contribution of TMD physicochemical properties (length, bulkiness, and palmitoylation) to DSG1 lipid raft association. Sucrose gradient fractionations revealed that TMD length and bulkiness, but not palmitoylation, govern DSG1 lipid raft association. Further, DSG1 raft association determines plakoglobin recruitment to raft domains. Super-resolution imaging and functional assays uncovered a strong relationship between the efficiency of DSG1TMD lipid raft association and the formation of morphologically and functionally robust desmosomes. Lipid raft association regulated both desmosome assembly dynamics and DSG1 cell surface stability, indicating that DSG1 lipid raft association is required for both desmosome formation and maintenance. These studies identify the biophysical properties of desmoglein transmembrane domains as key determinants of lipid raft association and desmosome adhesive function.