Biology subjects

Ye, R. D.

Publications and source records attributed to Ye, R. D..

4 recordsLinked to original sources

bioRxiv2023

Structural basis for constitutive activation and CXCL1 recognition of human herpesvirus 8-encoded G protein-coupled receptor KSHV-GPCR

Kaposis sarcoma-associated herpesvirus (KSHV) encodes a viral G protein-coupled receptor, KSHV-GPCR, that contributes to KSHV immune evasion and pathogenesis of Kaposis Sarcoma. KSHV-GPCR shares a high similarity with CXC chemokine receptors CXCR2 and can be activated by selected chemokine ligands. KSHV-GPCR is also unique for its constitutive activity by coupling to various G proteins. We investigated the structural basis of ligand-dependent as well as constitutive activity of KSHV-GPCR through cryo-EM structural determination of KSHV-GPCR-Gi signaling complexes with and without bound CXCL1 chemokine ligand. Analysis of the apo-KSHV-GPCR-Gi structure, with an overall resolution of 2.81 [A], unraveled the involvement of extracellular loop 2 in constitutive activation of the receptor. This and other structural motifs serve to stabilize the constitutively-active KSHV-GPCR. The CXCL1-bound KSHV-GPCR-Gi structure was solved to an overall resolution of 3.01 [A], and showed a two-site binding of the chemokine by the receptor. Together with functional validations, this work shed light on the structural basis for constitutive as well as CXCL1-induced activation of KSHV-GPCR. The work also demonstrates evolutionary advantage in immune evasion by KSHV through its virally encoded chemokine receptor, with potential implications in developing therapeutic strategies for KSHV infection.

immunology↗

bioRxiv2023

Cryo-EM structure of monomeric CXCL12-bound CXCR4 in active state

CXC chemokine receptor 4 (CXCR4) and its chemokine ligand CXCL12 are crucial to embryonic development, bone marrow retention of hematopoietic progenitor cells, cancer metastasis, angiogenesis and HIV-1 infection. Yet the structural basis for CXCR4 recognition of full-length CXCL12 remains unknown despite available structures of CXCR4 in complex with small molecule antagonists and a viral chemokine. Here we present a cryo-EM structure of monomeric CXCL12-CXCR4 in complex with heterotrimeric Gi proteins at a resolution of 2.65 [A]. The CXCL12-CXCR4 interaction at their N termini is stabilized by a polar interaction between the PC motif (C28NT in CXCR4) and CXC motif (the unique P10 in CXCL12). The N terminal 8 amino acids in CXCL12 insert into the transmembrane binding pocket (chemokine recognition site 2, CRS2) of CXCR4 with S4 defining an upward turn of V3, P2 and K1. Polar interactions involving C186ECL2 and D187ECL2, along with hydrogen bonding with E2887,39 and D2626,58 stabilize the N terminus of CXCL12 in CRS2. Hydrophobic interactions between side chains aligning CRS2 and P2, L5 and S6 of CXCL12 further strengthen its binding to the receptor. CXCL12 inserts deep into the binding pocket, and the 3.2[A] distance measured between V3 and the toggle switch W6.48 for G protein activation is among the shortest of all chemokine-receptor pairs. Our findings provide structural insights into the recognition mechanism of CXCR4 for its chemokine ligand CXCL12.

immunology↗

bioRxiv2023

Structural basis for EROS binding to human phagocyte NADPH oxidase NOX2

EROS (essential for reactive oxygen species) is a recently identified molecular chaperone of NOX2 (gp91phox), the catalytic subunit of phagocyte NADPH oxidase. Deficiency in NOX2 expression or function due to genetic mutations leads to chronic granulomatous disease (CGD) with recurrent bacterial and fungal infections. To delineate how EROS interacts with NOX2, we solved the cryo-EM structure of the EROS-NOX2-p22phox heterotrimeric complex. EROS binds to NOX2 in plasma membrane through its anti-parallel -helices H1 and H2, and in cytoplasm through multiple {beta}-strands that form hydrogen bonds with the C terminal fragment of NOX2. EROS binding alters the conformation of the TM2 and TM6 transmembrane helices, increases the distance between the two hemes, and causes dislocation of the binding site for flavin adenine dinucleotide (FAD). EROS colocalizes with NOX2 on cell surface of neutrophil-like HL-60 cells and forms a heterotrimer with mature NOX2-p22phox in transfected cells. Phorbol myristate acetate, an activator of NOX2, induces dissociation of EROS from NOX2 in a NanoLuc complementation assay with concurrent production of superoxide in reconstituted cells. Taken together, these findings provide a structural basis for EROS-NOX2 interaction and suggest a previously unidentified function of EROS in regulating NOX2 activation.

immunology↗

bioRxiv2023

Cryo-EM structure of the human G-protein coupled receptor 1 (GPR1) - Gi protein complex bound to the chemerin C-terminal nonapeptide

Chemerin is an adipokine with chemotactic activity to a subset of leukocytes. Chemerin mainly acts through its C-terminal nonapeptide (YFPGQFAFS, C9) that bind to three G protein-coupled receptors including chemokine-like receptor 1 (CMKLR1), G-protein coupled receptor 1 (GPR1) and C-C chemokine receptor-like 2 (CCRL2). We examined C9 signaling through GPR1 and found this receptor capable of Gi signaling but very weak {beta}-arrestin signaling. Here we report high-resolution cryo-EM structures of GPR1-Gi complexes bound to full-length chemerin and to C9, respectively. Unlike C9 that inserts directly into a transmembrane binding pocket, full-length chemerin uses its N-terminal globular core for extensive interaction with the N terminus of GPR1. Within the binding pocket, the C terminal loop containing the nonapeptide takes the same "S-shape" pose as synthetic C9 nonapeptide. These findings explain why the nonapeptide is a full agonist of GPR1, and demonstrate that chemerin uses a "two-site" model for interaction with GPR1. An analysis of the GPR1-Gi protein interface found high similarities to the CMKLR1-Gi complex, as confirmed by site-directed mutagenesis with functional verifications. Our structural analysis demonstrates shared features with chemokines in that chemerin acts as a "reverse chemokine" with switched functions of its N and C termini in the interaction with GPR1.

pharmacology and toxicology↗