|ZFIN ID: ZDB-PUB-150107-6|
Imaging complex protein metabolism in live organisms by stimulated Raman scattering microscopy with isotope labeling
Wei, L., Shen, Y., Xu, F., Hu, F., Harrington, J.K., Targoff, K.L., Min, W.
|Source:||ACS Chemical Biology 10(3): 901-8 (Journal)|
|Registered Authors:||Targoff, Kimara, Xu, Fang|
|PubMed:||25560305 Full text @ ACS Chem. Biol.|
Wei, L., Shen, Y., Xu, F., Hu, F., Harrington, J.K., Targoff, K.L., Min, W. (2015) Imaging complex protein metabolism in live organisms by stimulated Raman scattering microscopy with isotope labeling. ACS Chemical Biology. 10(3):901-8.
ABSTRACTProtein metabolism consisting of both synthesis and degradation is highly complex, playing an indispensable regulatory role throughout physiological and pathological processes. To study protein metabolism, extensive efforts, such as autoradiography, mass spectrometry and fluorescence microscopy, have been devoted over the past decades. However, noninvasive and global visualization of protein metabolism has been proven to be highly challenging especially in live systems. Recently, stimulated Raman scattering (SRS) microscopy coupled with metabolic labeling of deuterated amino acids (D-AAs) was demonstrated for imaging newly synthesized proteins in cultured cell lines. Herein we significantly generalize this notion to develop a comprehensive labeling and imaging platform for live visualization of complex protein metabolism including synthesis, degradation and pulse-chase analysis of two temporally defined populations. First, the deuterium labeling efficiency was optimized, allowing time-lapse imaging of protein synthesis dynamics within individual live cells with high spatial-temporal resolution. Second, by tracking the methyl group (CH3) distribution attributed to pre-existing proteins, this platform also enables us to map protein degradation inside live cells. Third, using two sub-sets of structurally and spectroscopically distinct D-AAs, we achieved two-color pulse-chase imaging, as demonstrated by observing aggregate formation of mutant hungtingtin proteins. Finally, going beyond simple cell lines, we demonstrated imaging ability of protein synthesis in brain tissues, zebrafish and mice in vivo. Hence, the presented labeling and imaging platform would be a valuable tool to study complex protein metabolism with high sensitivity, resolution and biocompatibility for a broad spectrum of systems ranging from cells to model animals, and possibly to humans.
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