Reengineering protein-phosphorylation switches

doi:10.1126/science.abj5028

GSTDTAP > 气候变化

DOI	10.1126/science.abj5028
	Reengineering protein-phosphorylation switches
	Boris N. Kholodenko; Mariko Okada
	2021-07-02
发表期刊	Science
出版年	2021
英文摘要	Toggle switches and oscillations in cellular networks have been of scientific interest since the late 1990s ([ 1 ][1]–[ 4 ][2]). Synthetic biologists have strived to build similar input-output responses by constructing DNA-RNA-protein circuits ([ 1 ][1]). Engineering genetic circuits has seen successes, including a synthetic circadian clock with oscillation periods of hours ([ 5 ][3]) and genetic bistable toggle switches operating on time scales of tens of minutes to hours ([ 6 ][4]). On page 75 of this issue, Mishra et al. ([ 7 ][5]) report the design of a faster regulatory network in yeast comprising synthetic protein phosphorylation circuits that act as logic gates. Furthermore, the authors identified similar network motifs across known endogenous signaling pathways in yeast. Phosphorylation-encoded switches and oscillators in protein networks operate quicker than genetic switches, but attempts to build these circuits synthetically have so far incorporated slower DNA logical operations ([ 8 ][6]). To attain rapid responses, optogenetics manipulations have been extensively employed. Indeed, different periods of light exposure triggered distinct enzyme activation kinetics, dynamics-dependent gene expression, and cell differentiation that are normally induced by growth factors and receptors in mammalian cells ([ 9 ][7]). However, no synthetic system has ever reproduced a logic gate that responds to signals on a time scale of seconds. In digital electronics, logical gates are combined into integrated circuits to perform chains of operations that enable computer calculations. Mishra et al. constructed phosphorylation circuits that operate as logic gates (OR, NOT, and BUFFER) used in electronics. Each gate is built with several fusion proteins. The upstream protein receives an input signal and binds a downstream protein, which is the gate output. The input protein effector domain is a protein-phosphorylating enzyme (kinase) for an OR gate or a dephosphorylating enzyme (phosphatase) for a NOT gate. In an engineered circuit, two distinct fusion proteins receive different input signals, and both proteins share the same output protein. The synthetic NOT gate has a single input protein and a single output protein. After phosphorylation of an input fusion protein that has a phosphatase domain (PTP), the initially phosphorylated output protein is dephosphorylated, which eliminates signaling. Mutual repression topology of two protein nodes or pathways in a circuit creates a toggle switch featuring two stable states—that is, either node is active when the other node is inactive ([ 10 ][8]–[ 12 ][9]). Mishra et al. built a bistable toggle switch using two paired OR and NOT gates and several BUFFER gates that formed mutual repression topology of the switch (see the figure). This occurs because a protein cascade (activated by sorbitol) phosphorylates a downstream protein that is a fusion of a PTP domain and osmotic stress sensor high osmolarity glycerol [HOG1, a mitogen-activated protein kinase (MAPK)] labeled with green fluorescent protein (GFP). Activated phospho-PTP-HOG1-GFP dephosphorylates another fusion protein called HOT-JH1 [high-osmolarity–induced transcription protein 1 (HOT1) and a catalytic domain of Janus kinase JAK 2 (JH1)]. In turn, the isopentenyl adenine (IP)–stimulated protein CRE1-HKRRSLN1 [which consists of the Arabidopsis thaliana cytokinin receptor (CRE1)–histidine kinase response regulator fused to the intracellular effector domains SLN1 (HKRRSLN1)] activates a downstream protein, MAPK kinase kinase osmolarity two-component system protein 1 (SSK1) through a chain of protein phosphorylations (BUFFER) and prevents the activation of sorbitol-stimulated protein polymyxin B resistance protein 2 (PBS2, known as MAPK kinase). The two external inputs, sorbitol and IP, activate their cognate fusion proteins through the plasma membrane osmosensor high osmolarity signaling protein 1 (SHO1) and histidine kinase receptor, respectively. Nuclear localized PTP-HOG1-GFP was measured to characterize system behaviors. These two inputs switched the system between two stable states on a time scale of seconds. Why does the engineered toggle switch have such a sophisticated design of paired OR, NOT, and BUFFER gates? Increasing the number of elements in a circuit enables rapid and robust responses. An amplifier increases the speed of a logic gate, whereas an element with a sigmoidal input-output characteristic, implemented as a BUFFER gate, filters noise and increases robustness to internal and external noise. Synthetic biochemical devices are noisier than electronic circuits because of internal noise in transcription-translation and cell heterogeneity. Future work will assess how speed and robustness of protein logic gates depend on the number and characteristics of individual elements ([ 13 ][10]). This will open possibilities for engineering gate combinations in complex integrative circuits. Mishra et al. also searched the Kyoto Encyclopedia of Genes and Genomes for all known signaling pathways in the yeast Saccharomyces cerevisiae and identified several toggled network motifs. Their analysis suggests that biological networks might inherently enable logical operations and computations. 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领域	气候变化 ; 资源环境
URL	查看原文
引用统计
文献类型	期刊论文
条目标识符	http://119.78.100.173/C666/handle/2XK7JSWQ/334173
专题	气候变化资源环境科学
推荐引用方式 GB/T 7714	Boris N. Kholodenko,Mariko Okada. Reengineering protein-phosphorylation switches[J]. Science,2021.
APA	Boris N. Kholodenko,&Mariko Okada.(2021).Reengineering protein-phosphorylation switches.Science.
MLA	Boris N. Kholodenko,et al."Reengineering protein-phosphorylation switches".Science (2021).