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Abstract
BackgroundThe cytoskeleton is a key component of the system responsible for transmitting mechanical cues from the cellular environment to the nucleus, where they trigger downstream responses. This communication is particularly relevant in embryonic stem (ES) cells since forces can regulate cell fate and guide developmental processes. However, little is known regarding cytoskeleton organization in ES cells, and thus, relevant aspects of nuclear-cytoskeletal interactions remain elusive.ResultsWe explored the three-dimensional distribution of the cytoskeleton in live ES cells and show that these filaments affect the shape of the nucleus. Next, we evaluated if cytoskeletal components indirectly modulate the binding of the pluripotency transcription factor OCT4 to chromatin targets. We show that actin depolymerization triggers OCT4 binding to chromatin sites whereas vimentin disruption produces the opposite effect. In contrast to actin, vimentin contributes to the preservation of OCT4-chromatin interactions and, consequently, may have a pro-stemness role.ConclusionsOur results suggest roles of components of the cytoskeleton in shaping the nucleus of ES cells, influencing the interactions of the transcription factor OCT4 with the chromatin and potentially affecting pluripotency and cell fate.
Highlights
The cytoskeleton is a key component of the system responsible for transmitting mechanical cues from the cellular environment to the nucleus, where they trigger downstream responses
We evaluate if the different cytoskeleton components modulate the nuclear shape and use fluorescence correlation spectroscopy (FCS) to test if these networks affect the dynamical organization of OCT4, a key pluripotency transcription factor (TF)
Three-dimensional organization of the cytoskeleton of mouse embryonic stem (ES) cells In order to examine the 3D organization of the cytoskeleton in naïve ES cells, we acquired confocal z-stacks of live cells co-expressing cytoskeleton-related proteins fused to green fluorescent proteins (GFP or Enhanced green fluorescent protein (EGFP)) and the histone H2B fused to the red fluorescent protein Monomeric red fluorescent protein (mCherry) (H2B-mCherry) to visualize the cell nucleus simultaneously
Summary
The cytoskeleton is a key component of the system responsible for transmitting mechanical cues from the cellular environment to the nucleus, where they trigger downstream responses This communication is relevant in embryonic stem (ES) cells since forces can regulate cell fate and guide developmental processes. Forces applied to cells may affect the shape and position of the nucleus [3, 4] and modulate diverse aspects of its function including chromatin organization and gene expression programs [3, 5] This relation is relevant in stem cells since forces can regulate cell fate and guide developmental processes [6, 7]. Disruption or alterations of cytoskeleton components as actin [10, 11] or vimentin intermediate filaments [12] affect cell fate decisions emphasizing the necessity of a three-dimensional (3D) description of the cytoskeleton organization in live ES cells
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