Functional and structural characterization of osteocytic MLO-Y4 cell proteins encoded by genes differentially expressed in response to mechanical signals in vitro

Lukasz Kurgan,Graeme F Murray,Henry J Donahue,Fanchi Meng

doi:10.1038/s41598-018-25113-4

Lukasz Kurgan, Graeme F Murray + Show 2 more

Open Access

https://doi.org/10.1038/s41598-018-25113-4

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Abstract

The anabolic response of bone to mechanical load is partially the result of osteocyte response to fluid flow-induced shear stress. Understanding signaling pathways activated in osteocytes exposed to fluid flow could identify novel signaling pathways involved in the response of bone to mechanical load. Bioinformatics allows for a unique perspective and provides key first steps in understanding these signaling pathways. We examined proteins encoded by genes differentially expressed in response to fluid flow in murine osteocytic MLO-Y4 cells. We considered structural and functional characteristics including putative intrinsic disorder, evolutionary conservation, interconnectedness in protein-protein interaction networks, and cellular localization. Our analysis suggests that proteins encoded by fluid flow activated genes have lower than expected conservation, are depleted in intrinsic disorder, maintain typical levels of connectivity for the murine proteome, and are found in the cytoplasm and extracellular space. Pathway analyses reveal that these proteins are associated with cellular response to stress, chemokine and cytokine activity, enzyme binding, and osteoclast differentiation. The lower than expected disorder of proteins encoded by flow activated genes suggests they are relatively specialized.

Highlights

Mechanical fluctuations in the extracellular environment, such as interstitial fluid flow-induced shear stress, induce cell perturbations including membrane deformations, cytoskeletal restructuring, conformational changes in transmembrane proteins, changes in the glycol calyx and movement of cilia
We have previously used gene microarrays, proteomics and RNA sequencing analysis (RNA-Seq) to identify increases in inflammatory C-X-C motif chemokines, including Ccl[2], and the HIF-1α, IL-17, and AMPK signaling pathways induced by fluid flow[3,4]
We comprehensively characterized several major structural and functional characteristics of osteocytic proteins encoded by genes differentially expressed due to exposure to fluid flow-induced shear stress

Summary

Introduction

Mechanical fluctuations in the extracellular environment, such as interstitial fluid flow-induced shear stress, induce cell perturbations including membrane deformations, cytoskeletal restructuring, conformational changes in transmembrane proteins, changes in the glycol calyx and movement of cilia. We have previously used gene microarrays, proteomics and RNA sequencing analysis (RNA-Seq) to identify increases in inflammatory C-X-C motif chemokines, including Ccl[2], and the HIF-1α, IL-17, and AMPK signaling pathways induced by fluid flow[3,4]. These studies were limited as they did not examine functional and structural characteristics of proteins encoded by the corresponding differentially expressed genes and have focused only on genes with the highest signal. A high level of disorder is a substantial obstacle to solving protein structure[25,26,27] and to performing rational drug design that relies on knowledge of protein structure[28,29,30]

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