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16 March 2025

New Study Reveals Role Of Gamma-Actin In Epithelial Cells

Research uncovers how gamma-actin modulates beta-actin levels and junction dynamics, impacting cell membrane mechanics.

A recent study sheds light on the intriguing regulatory roles of γ-actin and β-actin isoforms within epithelial cells, particularly focusing on how the depletion of γ-actin can significantly influence the expression of β-actin and the dynamics of cellular junctions. The findings reveal the complexity of biomechanical couplings linking these actin isoforms with non-muscle myosin 2A, which are pivotal to maintaining the integrity and function of epithelial tissues.

Epithelial tissues, known for their protective and barrier functions, rely heavily on their structural organization, largely governed by the actomyosin cytoskeleton. This network comprises various actin isoforms, each with distinct biochemical properties and functionalities. While β-actin is traditionally associated with cell stretching and motility, γ-actin is more concentrated at the apical junctions of epithelial cells, where it plays unique roles. Understanding how these two isoforms interact and compensate for each other could provide insights with significant clinical relevance, especially concerning pathologies arising from epithelial dysfunction.

Employing CRISPR/Cas9 gene editing technology, researchers created γ-actin knockout (KO) clones from Madin-Darby Canine Kidney (MDCK) cell lines. Their investigations unveiled compelling results: the knockout of γ-actin led to marked increases in β-actin expression at cell-cell junctions as well as within the cytoplasm. Immunofluorescence assays confirm these increases, corroborated by biochemical analyses showing heightened β-actin protein and mRNA levels. Importantly, this accumulation correlates with evidence of increased junctional dynamics, indicating potential regulatory feedback circuits within the actomyosin network.

One of the key discoveries of the research revolves around the effect of γ-actin deficiency on tight junctions (TJs) and apical membrane mechanics. The study states, "the KO of γ-actin promotes increased TJ membrane tortuosity...," highlighting significant alterations to junctional properties. Through comprehensive analysis, the scientists reported heightened tight junction tortuosity attributed to the upregulation of non-muscle myosin 2A alongside β-actin, which was also found to modulate junctional morphology. Given the important barrier functions of tight junctions, these findings suggest significant biological consequences of altered actin dynamics and may indicate broader relevance for both developmental biology and disease contexts.

To assess the mechanics of the apical membrane, atomic force microscopy (AFM) was employed. Measurements indicated reduced stiffness of the apical membrane cortex upon γ-actin knockout, establishing γ-actin as instrumental for maintaining membrane integrity and resistance to deformation. It was observed ;without the presence of γ-actin, the elastic properties of the cell membrane are compromised, underlying possible links to disease states where epithelial stiffness is altered.

Despite these changes, the researchers noted no significant disruption to the overall organization of tight junction proteins or their functionality. They found, "the KO of γ-actin does not alter the TEER (transepithelial electrical resistance)," indicating intact junctional barrier function even when cytoskeletal dynamics change. This provides valuable insight, as previous investigations suggested defects upon γ-actin depletion could contribute to increased permeability and loss of barrier function.

Interestingly, the dynamic exchange of junction-associated proteins, including ZO-1 and cingulin, was altered significantly. The study states, "the KO of γ-actin results in increased dynamic exchange of cingulin and ZO-1," indicating changes to junctional interactions and communities affecting how epithelial cells respond to mechanical stressors. Enhanced dynamics of these proteins may open potential avenues for therapeutic interventions, targeting the mechanical responses of epithelial tissues during inflammation or other diseases.

Through these insights, the researchers have laid the groundwork for future explorations, particularly concerning how mechanical and biochemical cues converge to regulate epithelial cell behavior and health. Overall, this work significantly enhances our comprehension of the biomechanical circuitry governing epithelial structure, advocating for future research probing the specific pathways facilitating these regulatory mechanisms and their impact on cell and tissue physiology.

These findings, elucidated through rigorous scientific investigations, underline the importance of cytoskeletal dynamics and their regulation, contributing to the broader narrative of cancer, tissue regeneration, and epithelial barrier integrity.