ZFYVE21 is a complement-induced Rab5 effector that activates non-canonical NF-\u03baB via phosphoinosotide remodeling of endosomes

Caodi Fang,Guangxin Li,Whitney Fu,Jordan S Pober,Manal Patel,Kevin Liu,Lingfeng Qin,Jonathan Merola,Thomas D Manes,Peter A Nigrovic,Nancy C Kirkiles-Smith,Zuzana Tobiasova,Lufang Liu,Dan Jane-Wit,George Tellides,Catherine Xie,Gregory T Tietjen,Rebecca Liu

doi:10.1038/s41467-019-10041-2

Abstract

Complement promotes vascular inflammation in transplant organ rejection and connective tissue diseases. Here we identify ZFYVE21 as a complement-induced Rab5 effector that induces non-canonical NF-κB in endothelial cells (EC). In response to membrane attack complexes (MAC), ZFYVE21 is post-translationally stabilized on MAC+Rab5+ endosomes in a Rab5- and PI(3)P-dependent manner. ZFYVE21 promotes SMURF2-mediated polyubiquitinylation and proteasome-dependent degradation of endosome-associated PTEN to induce vesicular enrichment of PI(3,4,5)P3 and sequential recruitment of activated Akt and NF-κB-inducing kinase (NIK). Pharmacologic alteration of cellular phosphoinositide content with miltefosine reduces ZFYVE21 induction, EC activation, and allograft vasculopathy in a humanized mouse model. ZFYVE21 induction distinctly occurs in response to MAC and is detected in human renal and synovial tissues. Our data identifies ZFYVE21 as a Rab5 effector, defines a Rab5-ZFYVE21-SMURF2-pAkt axis by which it mediates EC activation, and demonstrates a role for this pathway in complement-mediated conditions.

Highlights

Complement promotes vascular inflammation in transplant organ rejection and connective tissue diseases
We previously showed that complement membrane attack complexes (MACs) trigger endothelial cells (EC) activation[1,2]
Human umbilical vein endothelial cells (HUVECs) were stably transduced with a Rab5-GFP fusion protein and treated with “high” panel reactive antibody (PRA) sera supplemented with AF647-labeled C9, the most abundant complement component in MAC

Summary

Introduction

Complement promotes vascular inflammation in transplant organ rejection and connective tissue diseases. We used sera from allo-sensitized transplant candidates containing high-titer panel reactive antibody (PRA) to induce alloantibodymediated MACs on human EC in vitro[1,2] and in vivo[5] In these models, MAC and not IgG binding or anaphylatoxins elicited EC activation by inducing via posttranslational stabilization of nuclear factor (NF)-κB-inducing kinase (NIK)[2], to activate noncanonical NF-κB. Informed by a genome-wide small interfering RNA (siRNA) screen[1], MAC were shown to initiate an alternative, TRAF3-independent mechanism of NIK stabilization in ~30 min In this alternative mechanism, membrane-bound MAC are internalized via clathrin-mediated endocytosis and transferred to Rab5+ endosomes to form a MAC +Rab5+ compartment. ZFYVE21 is identified as a posttranslationally induced Rab[5] effector that triggers phosphoinositide (PI) remodeling of early endosome membranes to mediate non-canonical NF-κB signal activation and tissue inflammation. ZFYVE21 is further explored both as a therapeutic target to modulate EC activation in vivo and as a specific biomarker for complement-mediated signaling in human tissues

Methods

Results

Conclusion