Measurement of the unfolded protein response (UPR) in monocytes.
In mammalian cells, the primary function of the endoplasmic reticulum (ER) is to synthesize and assemble membrane and secreted proteins. As the main site of protein folding and posttranslational modification in the cell, the ER operates a highly conserved quality control system to ensure only correctly assembled proteins exit the ER and misfolded and unfolded proteins are retained for disposal. Any disruption in the equilibrium of the ER engages a multifaceted intracellular signaling pathway termed the unfolded protein response (UPR) to restore normal conditions in the cell. A variety of pathological conditions can induce activation of the UPR, including neurodegenerative disorders such as Parkinson's disease, metabolic disorders such as atherosclerosis, and conformational disorders such as cystic fibrosis. Conformational disorders are characterized by mutations that modify the final structure of a protein and any cells that express abnormal protein risk functional impairment. The monocyte is an important and long-lived immune cell and acts as a key immunological orchestrator, dictating the intensity and duration of the host immune response. Monocytes expressing misfolded or unfolded protein may exhibit UPR activation and this can compromise the host immune system. Here, we describe in detail methods and protocols for the examination of UPR activation in peripheral blood monocytes. This guide should provide new investigators to the field with a broad understanding of the tools required to investigate the UPR in the monocyte.
Department of Medicine, Royal College of Surgeons in Ireland, Education and Research Centre, Beaumont Hospital, Dublin, Ireland.
Tomás P
TP
Carroll
Catherine M
CM
Greene
Noel G
NG
McElvaney
D004268:DNA-Binding Proteins / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D016209:Interleukin-8 / Q000032:analysis; D009000:Monocytes / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors / Q000235:genetics; D056811:Unfolded Protein Response
D004268:DNA-Binding Proteins; D016209:Interleukin-8; D012333:RNA, Messenger; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors