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Finding the “Switchman” Responsible for Biased Signalling

By uncovering the amino acid essential for recognising GPCR ligands, biased signalling of CCL15 can be restricted, presenting an attractive drug target for treating asthma.

Asthma is one of the most common long-term respiratory diseases in the world and its prevalence is on the rise. Epidemiological studies have revealed that the percentage of asthma ranges from one per cent to 18 per cent in different countries, making up to 400 million people living with asthma worldwide. In China, the number of asthma patients aged 20 years and above stands at 45.7 million. Yet given its prevalence, the treatment of asthma remains inadequate. There is still a great need to investigate the pathogenesis of asthma and identify new therapeutic targets.

Here, Professor Shen Huahao and Professor Ying Songmin at the Second Affiliated Hospital of the Zhejiang University School of Medicine worked with Professor Zhang Yan at the Zhejiang University School of Medicine found and characterised different N-terminal truncations of endogenous chemokine CCL15 as balanced or biased agonists targeting CCR1, laying the foundation for the development of asthma drugs targeting CCR1.

In 2003, Prof. Shen laid the groundwork by demonstrating that eosinophils could directly induce asthma. With continued effort over 10 years, professors Shen and Ying further illustrated the key role of chemokine receptor CCR1 activation and for the first time, presented CCR1 and its ligands as new targets for asthma treatment. However, clinical trials involving these receptors have remained unsuccessful.

“We have clearly identified the vital key of CCR1 activation in inflammation, but we cannot figure out a feasible way to inhibit it. It is because we fail to decipher its intrinsic mechanism. Because of that, we cannot precisely regulate it,” said Prof. Ying. “We hope that we can gain a perspicacious understanding of the interaction between CCR1 and its ligands so as to unravel this enigma.”

Chemokine receptor CCR1 is a type of G protein-coupled receptor (GPCR). GPCRs are the largest family of transmembrane proteins and are the most important drug target in the human body as they transmit extracellular stimuli across the plasma membrane and initiate cell signalling pathways. CCR1 is widely expressed in various immune cells and its suppression has been effective in controlling the maturation and migration of immune cells, thereby making it an attractive drug target for many autoimmune and allergic diseases like asthma.

Chemokine-receptor interactions play an important role in guiding leukocyte trafficking in immune surveillance and inflammation response. Just as most receptors can be recognised by various chemokines, many chemokines can also activate multiple receptors. While previously thought as redundant, this phenomenon can now be appreciated as biased agonism, allowing the fine-tuning of a chemokine-induced physiological response.

CCL15 is an endogenous ligand of CCR1 at which the N-terminal signal peptide is cleaved up on its secretion to extracellular media. In previous works, mass spectrometry of clinical samples revealed that the N-terminus of secreted CCL15 underwent further cleavage and displayed various N-terminal truncations due to proteolytic processing by activated mast cells and neutrophils. Multiple studies have put forth that the N-terminus of chemokines worked as a determinant for CCR1 activation, as well as bringing about biased agonism.

In this study, the team found that increasing the secretion of metalloenzymes in inflammation acts like a sharp knife that can cut CCL15 ligands into three subtypes with different lengths (CCL15 L, CCL15 M, and CCL15 S), leading to distinct signalling pathways.

With cryogenic-electron microscopy (cryo-EM), the density maps revealed that the side chain Tyr291 (Y291 7.43) in the binding pocket of CCR1 displayed distinct conformations with different ligands. When Y291 7.43 was mutated, the biased signalling properties of CCL15 virtually disappeared, suggesting that Y291 7.43 is the key amino acid site facilitating the biased activation of CCL15 on CCR1.

Taken into consideration with computational and pharmacological studies, the team’s work revealed how Y291 7.43 is crucial in identifying GPCR ligands and signalling.

“Y291 7.43 is like a railroad switchman that determines the direction of signals,” said Zhang. “Once we know how signalling molecules activate CCR1, we will be able to work out ways to hinder them from working. As a consequence, we may consider how to seal this ‘switchman’ in our future work.” [APBN]

Source: Shao et al. (2021). Identification and mechanism of G protein-biased ligands for chemokine receptor CCR1. Nature chemical biology, 1-8.