Enhancing protein expression in single HEK 293 cells.
Recombinant proteins are routinely expressed in heterologous expression systems such as human embryonic kidney 293 (HEK 293) cells. The efficiency of the expression is critical when the expressed protein must be characterized at the single-cell level. Here we describe a simple method by which the protein expression efficiency in single HEK 293 cells is enhanced by coexpressing simian virus 40 large T antigen (TAg), a powerful oncoprotein. Using the GluR2 ionotropic glutamate receptor as an example, we found that the receptor expression in single HEK 293S cells increased approximately seven-fold. The ratio of the plasmid amount of TAg to that of the receptor was optimized at 1:10, while the receptor function was unaffected in the presence of TAg. We further used fluorescence imaging from a population of cells as an independent detection method and found a similar increase in expression of green fluorescent protein (GFP) by TAg coexpression. This method is thus applicable for enhancing the expression of both membrane and soluble proteins at the single-cell level. More importantly, the function of a protein can be studied directly in intact cells, a feature particularly useful for studying membrane proteins.
Huang
Zhen
Z
Department of Chemistry, Center for Neuroscience Research, University at Albany, SUNY, Albany, NY 12222, USA.
Li
Gang
G
Pei
Weimin
W
Sosa
Leivi A
LA
Niu
Li
L
D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D005786:Gene Expression Regulation / Q000502:physiology; D005822:Genetic Vectors / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism*; D008564:Membrane Potentials / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008967:Molecular Biology / Q000379:methods*; D009448:Neurochemistry / Q000379:methods*; D018408:Patch-Clamp Techniques; D010957:Plasmids / Q000502:physiology; D018091:Receptors, AMPA / Q000096:biosynthesis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D015854:Up-Regulation / Q000502:physiology
D000957:Antigens, Viral, Tumor; D008565:Membrane Proteins; D018091:Receptors, AMPA; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; C104722:glutamate receptor ionotropic, AMPA 2