Distinct Signaling Pathways Regulate Membrane Architecture And Drive Bone Stem Cell Differentiation

Abstract

Recent research demonstrates that membrane domains on the plasma membrane of cells and the underlying cytoskeleton regulate subsequent receptor mediated signaling. However less is known about how the three dimensional plasma membrane architecture created by proteins, lipids and cytoskeleton is regulated by the cell itself. Influencing the architecture of the plasma membrane by distinct signaling pathways may be one crucial mechanism for the cell to selectively respond to external signaling stimuli. Therefore, signaling pathways must be identified that affect its assembly.Recently, we showed that membrane receptor and domain aggregation and their dynamics drive bone stem cell differentiation toward osteoblastic lineage commitment, however we failed to identify pathways necessary for lipid, protein and cytoskeletal assembly. In order to identify these novel roles of classical pathways, we employed the Family of Image Correlation Spectroscopy, Atomic Force Microscopy followed by Fractal Analysis, roughness calculations, and molecular biology techniques to primary bone marrow stromal cells isolated from mice; C57BL/6 (B6, control) and B6C3H-1-12 (1-12, high peak bone density).Our results revealed two signaling pathways that affect membrane morphology. These signaling pathways are activated at distinct steps during osteoblast differentiation and communication with surrounding cells. While triggering the first signaling cascade altered the topography of the cell membrane by modifying the maximum valley depth, the second pathway caused a change in the perimeter and area of distinct membrane regions. Our data suggest that osteoblast precursor cells and osteoblasts undergo physical changes in the membrane dependent on their stage of differentiation. These variations in the topography were triggered by distinct signaling pathways causing a rearrangement of the plasma membrane. These changes may be crucial for the response of the cells to external stimuli received at various timeopoints during the differentiation event.