Simulation of the macrophage chemotaxis pathway facilitated by targeted proteomics

Tuesday, September 23, 2014 — Poster Session III
12:00 p.m. – 2:00 p.m.	FAES Academic Center	NIAID	SYSBIO-3

* FARE Award Winner

Authors

• N.P. Manes
• M. Koppenol-Raab
• E. An
• V.H. Sjoelund
• J. Sun
• B.R. Angermann
• M. Ishii
• M. Meier-Schellersheim
• R.N. Germain
• A. Nita-Lazar

Abstract

Chemotaxis is critical to a wide range of biological processes including bone resorption. Osteoclasts are monocyte-derived multinuclear cells that directly attach to and resorb bone. Recently it was reported that the phosphosphingolipid sphingosine-1-phosphate (S1P) regulates bone resorption by functioning as both a chemoattractant and chemorepulsant of osteoclast precursors (OPs) through two G-protein coupled receptors (S1PR1 and S1PR2, respectively), which antagonize each other in an S1P-concentration dependent manner. To develop a deeper understanding of mammalian cell chemotaxis, this investigation applied targeted proteomics, transcriptomics, and pathway simulation to investigate S1P-mediated chemotaxis of RAW 264.7 cells (model OPs). RNA-seq was used to identify expressed target proteins and their transcript abundance. Selected reaction monitoring mass spectrometry using internal peptide standards was performed to produce absolute abundance measurements of target proteins. The resulting transcript and protein abundance values correlated strongly, and, in the absence of robust measurements, the transcript abundances were used to estimate protein abundances. Pathway simulations using Simmune were then performed, and the in silico results were compared to in vivo data.

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