抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Recent advances in the development of genome editing technologies based on programmable nucleases have substantially improved our ability to make precise changes in the genomes of eukaryotic cells. Genome editing is already broadening our ability to elucidate the contribution of genetics to disease by facilitating the creation of more accurate cellular and animal models of pathological processes. A particularly tantalizing application of programmable nucleases is the potential to directly correct genetic mutations in affected tissues and cells to treat diseases that are refractory to traditional therapies. Here we discuss current progress toward developing programmable nuclease–based therapies as well as future prospects and challenges.

Therapeutic genome editing: prospects and challenges

The ability to manipulate the structure and function of promising nanosystems via energy input and external stimuli is emerging as an attractive paradigm for developing reconfigurable and programmable nanomaterials and multifunctional devices. Light stimulus manifestly represents a preferred external physical and chemical tool for in situ remote command of the functional attributes of nanomaterials and nanosystems due to its unique advantages of high spatial and temporal resolution and digital controllability. Photochromic moieties are known to undergo reversible photochemical transformations between different states with distinct properties, which have been extensively introduced into various functional nanosystems such as nanomachines, nanoparticles, nanoelectronics, supramolecular nanoassemblies, and biological nanosystems. The integration of photochromism into these nanosystems has endowed the resultant nanostructures or advanced materials with intriguing photoresponsive behaviors and more sophisticated functions. In this Review, we provide an account of the recent advancements in reversible photocontrol of the structures and functions of photochromic nanosystems and their applications. The important design concepts of such truly advanced materials are discussed, their fabrication methods are emphasized, and their applications are highlighted. The Review is concluded by briefly outlining the challenges that need to be addressed and the opportunities that can be tapped into. We hope that the review of the flourishing and vibrant topic with myriad possibilities would shine light on exploring the future nanoworld by encouraging and opening the windows to meaningful multidisciplinary cooperation of engineers from different backgrounds and scientists from the fields such as chemistry, physics, engineering, biology, nanotechnology and materials science.

Photochromism into nanosystems: towards lighting up the future nanoworld.

Nature uses a limited, conservative set of amino acids to synthesize proteins. The ability to genetically encode an expanded set of building blocks with new chemical and physical properties is transforming the study, manipulation and evolution of proteins, and is enabling diverse applications, including approaches to probe, image and control protein function, and to precisely engineer therapeutics. Underpinning this transformation are strategies to engineer and rewire translation. Emerging strategies aim to reprogram the genetic code so that noncanonical biopolymers can be synthesized and evolved, and to test the limits of our ability to engineer the translational machinery and systematically recode genomes.

Expanding and reprogramming the genetic code.

Although many kinds of therapies are applied in the clinic, drug-resistance is a major and unavoidable problem. Another disturbing statistic is the limited number of drug targets, which are presently only 20–25% of all protein targets that are currently being studied. Moreover, the focus of current explorations of targets are their enzymatic functions, which ignores the functions from their scaffold moiety. As a promising and appealing technology, PROteolysis TArgeting Chimeras (PROTACs) have attracted great attention both from academia and industry for finding available approaches to solve the above problems. PROTACs regulate protein function by degrading target proteins instead of inhibiting them, providing more sensitivity to drug-resistant targets and a greater chance to affect the nonenzymatic functions. PROTACs have been proven to show better selectivity compared to classic inhibitors. PROTACs can be described as a chemical knockdown approach with rapidity and reversibility, which presents new and different biology compared to other gene editing tools by avoiding misinterpretations that arise from potential genetic compensation and/or spontaneous mutations. PRTOACs have been widely explored throughout the world and have outperformed not only in cancer diseases, but also in immune disorders, viral infections and neurodegenerative diseases. Although PROTACs present a very promising and powerful approach for crossing the hurdles of present drug discovery and tool development in biology, more efforts are needed to gain to get deeper insight into the efficacy and safety of PROTACs in the clinic. More target binders and more E3 ligases applicable for developing PROTACs are waiting for exploration.

PROTACs: great opportunities for academia and industry.

Biological processes are naturally regulated with high spatial and temporal control, as is perhaps most evident in metazoan embryogenesis Chemical tools have been extensively utilized in cell and developmental biology to investigate cellular processes, and conditional control methods have expanded applications of these technologies toward resolving complex biological questions Light represents an excellent external trigger since it can be controlled with very high spatial and temporal precision To this end, several optically regulated tools have been developed and applied to living systems In this review we discuss recent developments of optochemical tools, including small molecules, peptides, proteins, and nucleic acids that can be irreversibly or reversibly controlled through light irradiation, with a focus on applications in cells and animals

Optochemical Control of Biological Processes in Cells and Animals.

The site-specific incorporation of three new coumarin lysine analogues into proteins was achieved in bacterial and mammalian cells using an engineered pyrrolysyl-tRNA synthetase system. The genetically encoded coumarin lysines were successfully applied as fluorescent cellular probes for protein localization and for the optical activation of protein function. As a proof-of-principle, photoregulation of firefly luciferase was achieved in live cells by caging a key lysine residue, and excellent OFF to ON light-switching ratios were observed. Furthermore, two-photon and single-photon optochemical control of EGFP maturation was demonstrated, enabling the use of different, potentially orthogonal excitation wavelengths (365, 405, and 760 nm) for the sequential activation of protein function in live cells. These results demonstrate that coumarin lysines are a new and valuable class of optical probes that can be used for the investigation and regulation of protein structure, dynamics, function, and localization in live cells. The small size of coumarin, the site-specific incorporation, the application as both a light-activated caging group and as a fluorescent probe, and the broad range of excitation wavelengths are advantageous over other genetically encoded photocontrol systems and provide a precise and multifunctional tool for cellular biology.

Genetically encoded optochemical probes for simultaneous fluorescence reporting and light activation of protein function with two-photon excitation.

As an emerging approach to protein perturbation, small molecule-induced protein degradation has gained significant attention as both a chemical tool and a potential therapeutic. To enable discrete control over its function, we have developed a broadly applicable approach for the optical activation of small molecule-induced protein degradation. By installing two different photolabile protecting groups, so-called caging groups, onto two different ligands recruiting Von Hippel-Lindau (VHL) and cereblon (CRBN) E3 ubiquitin ligases, our strategy enables light-triggered protein degradation for any small molecule warhead.

Optical Control of Small Molecule-Induced Protein Degradation.

Optochemische Steuerung biologischer Vorgänge in Zellen und Tieren

We report the genetic incorporation of caged cysteine and caged homocysteine into proteins in bacterial and mammalian cells. The genetic code of these cells was expanded with an engineered pyrrolysine tRNA/tRNA synthetase pair that accepts both light-activatable amino acids as substrates. Incorporation was validated by reporter assays, western blots, and mass spectrometry, and differences in incorporation efficiency were explained by molecular modeling of synthetase-amino acid interactions. As a proof-of-principle application, the genetic replacement of an active-site cysteine residue with a caged cysteine residue in Renilla luciferase led to a complete loss of enzyme activity; however, upon brief exposure to UV light, a >150-fold increase in enzymatic activity was observed, thus showcasing the applicability of the caged cysteine in live human cells. A simultaneously conducted genetic replacement with homocysteine yielded an enzyme with greatly reduced activity, thereby demonstrating the precise probing of a protein active site. These discoveries provide a new tool for the optochemical control of protein function in mammalian cells and expand the set of genetically encoded unnatural amino acids.

Yuta Naro

Papers

Optochemical Control of Biological Processes in Cells and Animals.

Genetically encoded optochemical probes for simultaneous fluorescence reporting and light activation of protein function with two-photon excitation.

Optical Control of Small Molecule-Induced Protein Degradation.

Optochemische Steuerung biologischer Vorgänge in Zellen und Tieren

Genetic encoding of caged cysteine and caged homocysteine in bacterial and mammalian cells.