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Single‐Cell Chemical Proteomics with an Activity‐Based Probe: Identification of Low‐Copy Membrane Proteins on Primary Neurons
Author(s) -
Xu Fei,
Zhao Han,
Feng Xiaojun,
Chen Linhai,
Chen Dongjuan,
Zhang Yang,
Nan Fajun,
Liu Jianfeng,
Liu BiFeng
Publication year - 2014
Publication title -
angewandte chemie
Language(s) - English
Resource type - Journals
eISSN - 1521-3757
pISSN - 0044-8249
DOI - 10.1002/ange.201402363
Subject(s) - proteomics , chemistry , membrane protein , hek 293 cells , microbiology and biotechnology , cell , rna splicing , proteome , single cell analysis , biophysics , membrane , biology , biochemistry , receptor , gene , rna
We propose a novel single‐cell chemical proteomics (SCCP) strategy to profile low‐abundance membrane proteins in single cells. In this approach, the membrane protein GB1 and its splicing variants were targeted on cultured cell lines and primary neurons using a specifically designed activity‐based probe. The functionally labeled single cells were encapsulated in individual buffer droplets on a PDMS microwell array, and were further picked up one at a time and loaded into a capillary electrophoresis system for cell lysis, separation, and laser‐induced fluorescence detection of the targeted proteins. The results revealed the expression of GB1 splicing variants in HEK and MEF cells, which was previously only suggested at the transcriptional level. We further applied this method to investigate single primary cells and observed significant heterogeneity among individual mouse cerebellar granule neurons. Interference experiments with GB1 antagonist and agonist validated this observation.