期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 112, 期 21, 页码 6619-6624出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1505652112
关键词
Alpha-actinin; actin; kidney disease; cell mechanics; traction force
资金
- NIH [PO1GM096971, DK083592, DK59588]
- Harvard Materials Research Science and Engineering Center [DMR-0820484]
- National Science Foundation [DMR-1310266]
- Natural Sciences and Engineering Research Council (NSERC) [05843]
- Division Of Materials Research
- Direct For Mathematical & Physical Scien [1310266] Funding Source: National Science Foundation
The actin cytoskeleton is a key element of cell structure and movement whose properties are determined by a host of accessory proteins. Actin cross-linking proteins create a connected network from individual actin filaments, and though the mechanical effects of cross-linker binding affinity on actin networks have been investigated in reconstituted systems, their impact on cellular forces is unknown. Here we show that the binding affinity of the actin crosslinker a-actinin 4 (ACTN4) in cells modulates cytoplasmic mobility, cellular movement, and traction forces. Using fluorescence recovery after photobleaching, we show that an ACTN4 mutation that causes human kidney disease roughly triples the wild-type binding affinity of ACTN4 to F-actin in cells, increasing the dissociation time from 29 +/- 13 to 86 +/- 29 s. This increased affinity creates a less dynamic cytoplasm, as demonstrated by reduced intracellular microsphere movement, and an approximate halving of cell speed. Surprisingly, these less motile cells generate larger forces. Using traction force microscopy, we show that increased binding affinity of ACTN4 increases the average contractile stress (from 1.8 +/- 0.7 to 4.7 +/- 0.5 kPa), and the average strain energy (0.4 +/- 0.2 to 2.1 +/- 0.4 pJ). We speculate that these changes may be explained by an increased solid-like nature of the cytoskeleton, where myosin activity is more partitioned into tension and less is dissipated through filament sliding. These findings demonstrate the impact of cross-linker point mutations on cell dynamics and forces, and suggest mechanisms by which such physical defects lead to human disease.
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