Genetically encoded X-ray cellular imaging for nanoscale protein localization

Huating Kong; Jichao Zhang; Jiang Li; Jian Wang; Hyun-Joon Shin; Renzhong Tai; Qinglong Yan; Kai Xia; Jun Hu; Lihua Wang; Ying Zhu; Chunhai Fan

doi:10.1093/nsr/nwaa055

Genetically encoded X-ray cellular imaging for nanoscale protein localization

Natl Sci Rev. 2020 Jul;7(7):1218-1227. doi: 10.1093/nsr/nwaa055. Epub 2020 Apr 2.

Authors

Huating Kong¹, Jichao Zhang¹, Jiang Li¹, Jian Wang², Hyun-Joon Shin³, Renzhong Tai¹, Qinglong Yan⁴, Kai Xia⁴, Jun Hu¹, Lihua Wang¹, Ying Zhu¹, Chunhai Fan⁵

Affiliations

¹ Bioimaging Center, Shanghai Synchrotron Radiation Facility, Zhangjiang Laboratory, Shanghai Advanced Research Institute, Chinese Academy of Sciences, Shanghai 201210, China.
² Canadian Light Source Inc., University of Saskatchewan, Saskatoon, SK S7N 2V3, Canada.
³ Pohang Accelerator Laboratory, POSTECH, Pohang 37673, Korea.
⁴ Division of Physical Biology, CAS Key Laboratory of Interfacial Physics and Technology, Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, China.
⁵ School of Chemistry and Chemical Engineering, Frontiers Science Center for Transformative Molecules, and Shanghai Key Laboratory for Nucleic Acids Chemistry and Nanomedicine, Institute of Molecular Medicine, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200240, China.

Abstract

Spatial resolution defines the physical limit of microscopes for probing biomolecular localization and interactions in cells. Whereas synchrotron-based X-ray microscopy (XRM) represents a unique approach for imaging a whole cell with nanoscale resolution due to its intrinsic nanoscale resolution and great penetration ability, existing approaches to label biomolecules rely on the use of exogenous tags that are multi-step and error-prone. Here, we repurpose engineered peroxidases as genetically encoded X-ray-sensitive tags (GXET) for site-specific labeling of protein-of-interest in mammalian cells. We find that 3,3^'-diaminobenzidine (DAB) polymers that are in-situ catalytically formed by fusion-expressed peroxidases are visible under XRM. Using this new tag, we imaged the protein location associated with the alteration of a DNA-methylation pathway with an ultra-high resolution of 30 nanometers. Importantly, the excellent energy resolution of XRM enables multicolor imaging using different peroxidase tags. The development of GXET enlightens the way to nanoscopic imaging for biological studies.

Keywords: 3,3′-diaminobenzidine polymers; cellular imaging; genetically encoded X-ray tags; nanoscale protein localization; synchrotron-based X-ray microscopy.