Biology subjects

Nowak, R. B.

Publications and source records attributed to Nowak, R. B..

3 recordsLinked to original sources

bioRxiv2018

Tropomyosin 3.5 protects F-actin networks required for tissue biomechanical properties

Tropomyosins (Tpms) stabilize F-actin and regulate interactions with other actin-binding proteins. The eye lens changes shape in order to fine focus light to transmit a clear image, and thus lens organ function is tied to its biomechanical properties, presenting an opportunity to study Tpm functions in tissue mechanics. The major mouse lens Tpm is Tpm3.5 (TM5NM5), a previously unstudied isoform. Decreased levels of Tpm3.5 lead to softer and less mechanically resilient lenses that are unable to resume their original shape after compression. While cell organization and morphology appear unaffected, Tmod1 dissociates from the membrane in Tpm3.5-deficient lens fiber cells resulting in reorganization of the spectrin-F-actin and -actinin-F-actin networks at the membrane. These rearranged F-actin networks appear to be less able to support mechanical load and resilience leading to an overall change in tissue mechanical properties. This is the first in vivo evidence that Tpm is essential for cell biomechanical stability in a load-bearing non-muscle tissue and indicates that Tpm3.5 protects mechanically stable, load-bearing F-actin in vivo.\n\nSummaryTropomyosin 3.5 stabilizes F-actin in eye lens fiber cells and promotes normal tissue biomechanical properties. Tpm3.5 deficiency leads to F-actin network rearrangements and decreased lens stiffness and resilience.

cell biology↗

bioRxiv2018

The effects of mechanical strain on mouse eye lens capsule and cellular microstructure

The understanding of multiscale load transfer within complex soft tissues is incomplete. The eye lens is ideal for multiscale tissue mechanics studies as its principal function is to fine focus light at different distances onto the retina via mechanical shape changes. The biomechanical function, resiliency, and intricate microstructure of the lens make it an excellent non-connective soft tissue model. We hypothesized that compressive strain applied onto whole lens tissue leads to deformation of specific microstructures and that this deformation is reversible following removal of load. For this examination, mouse lenses were compressed by sequential application of increasing load. Using confocal microscopy and quantitative image analysis, we determined that axial strain [≥]10% reduces capsule thickness, expands epithelial cell area, and separates fiber cell tips at the anterior region of the lenses. At the equatorial region, strain [≥]6% increases fiber cell widths. The effects of strain on lens epithelial cell area, capsule thickness, and equatorial fiber cell widths are reversible following the release of lenses from strain. However, although fiber cell tip separation following the removal of low loads is reversible, the separation becomes irreversible with application of higher loads. This irreversible separation between fiber cell tips leads to incomplete bulk lens resiliency. The lens is an accessible biomechanical model system that provides new insights on multiscale transfer of loads in soft tissues.

cell biology↗

bioRxiv2017

Myosin IIA interacts with the spectrin-actin membrane skeleton to control red blood cell membrane curvature and deformability

The biconcave disc shape and deformability of mammalian red blood cells (RBCs) relies upon the membrane skeleton, a viscoelastic network of short, membrane-associated actin filaments (F-actin) cross-linked by long, flexible spectrin tetramers. Nonmuscle myosin II (NMII) motors exert force on diverse F-actin networks to control cell shapes, but a function for NMII contractility in the 2D spectrin-F-actin network in RBCs has not been tested. Here, we show that RBCs contain membrane skeleton-associated NMIIA puncta, identified as bipolar filaments by super-resolution fluorescence microscopy. NMIIA association with the membrane skeleton is ATP-dependent, consistent with NMIIA motor domains binding to membrane skeleton F-actin and contributing to membrane mechanical stability. In addition, the NMIIA heavy and light chains are phosphorylated in vivo in RBCs, indicating active regulation of NMIIA motor activity and filament assembly, while reduced heavy chain phosphorylation of membrane skeleton-associated NMIIA indicates assembly of stable filaments at the membrane. Treatment of RBCs with blebbistatin, an inhibitor of NMII motor activity, decreases the number of NMIIA filaments associated with the membrane and enhances local, nanoscale membrane oscillations, suggesting decreased membrane tension. Blebbistatin-treated RBCs also exhibit elongated shapes, loss of membrane curvature, and enhanced deformability, indicating a role for NMIIA contractility in promoting membrane stiffness and maintaining RBC biconcave disc cell shape. As structures similar to the RBC membrane skeleton are conserved in many metazoan cell types, these data demonstrate a general function for NMII in controlling specialized membrane morphology and mechanical properties through contractile interactions with short F-actin in spectrin-F-actin networks.\n\nSignificance statementThe biconcave disc shape and deformability of the mammalian RBC is vital to its circulatory function, relying upon a 2D viscoelastic spectrin-F-actin network attached to the membrane. A role for myosin II (NMII) contractility in generating tension in this network and controlling RBC shape has never been tested. We show that NMIIA forms phosphorylated bipolar filaments in RBCs, which associate with F-actin at the membrane. NMIIA motor activity is required for interactions with the spectrin-F-actin network, and regulates RBC biconcave shape and deformability. These results provide a novel mechanism for actomyosin force generation at the plasma membrane, and may be applicable to other cell types such as neurons and polarized epithelial cells with a spectrin-F-actin-based membrane skeleton.

cell biology↗