Translocation of interleukin-1β into a vesicle intermediate in autophagy-mediated secretion.

Min Zhang,Randy Schekman,Ke Xu,Samuel J Kenny,Liang Ge

doi:10.7554/elife.11205

Abstract

Recent evidence suggests that autophagy facilitates the unconventional secretion of the pro-inflammatory cytokine interleukin 1β (IL-1β). Here, we reconstituted an autophagy-regulated secretion of mature IL-1β (m-IL-1β) in non-macrophage cells. We found that cytoplasmic IL-1β associates with the autophagosome and m-IL-1β enters into the lumen of a vesicle intermediate but not into the cytoplasmic interior formed by engulfment of the autophagic membrane. In advance of secretion, m-IL-1β appears to be translocated across a membrane in an event that may require m-IL-1β to be unfolded or remain conformationally flexible and is dependent on two KFERQ-like motifs essential for the association of IL-1β with HSP90. A vesicle, possibly a precursor of the phagophore, contains translocated m-IL-1β and later turns into an autophagosome in which m-IL-1β resides within the intermembrane space of the double-membrane structure. Completion of IL-1β secretion requires Golgi reassembly and stacking proteins (GRASPs) and multi-vesicular body (MVB) formation.

Highlights

Most eukaryotic secretory proteins with an N-terminal signal peptide are delivered through the classical secretion pathway involving an endoplasmic reticulum (ER)-to-Golgi apparatus itinerary (Lee et al, 2004; Schatz and Dobberstein, 1996)
Little cell lysis occurred during the treatment we used to induce interleukin 1b (IL-1b) secretion: Much less precursor than mature IL-1b and little cytoplasmic tubulin was detected released into the cell supernatant during the 2 hr incubation in starvation medium (Figure 1A)
Inhibition of autophagy by the phosphatidylinositol 3-kinase (PI3K) inhibitors 3-methyladenine (3-MA) or wortmannin (Wtm) blocked IL-1b secretion activated by starvation and caused the accumulation of mature IL-1b in the cell (Figure 1B)

Summary

Introduction

Most eukaryotic secretory proteins with an N-terminal signal peptide are delivered through the classical secretion pathway involving an endoplasmic reticulum (ER)-to-Golgi apparatus itinerary (Lee et al, 2004; Schatz and Dobberstein, 1996). A substantial number of secretory proteins lack a classical signal peptide, called leaderless cargoes, and are released by unconventional means of secretion (Nickel and Rabouille, 2009; Nickel and Seedorf, 2008). Unlike a unified route for classical protein secretion, leaderless cargoes undergoing unconventional secretion employ multiple means of protein delivery, the details of which are largely unknown (Ding et al, 2012; Nickel, 2010; Rabouille et al, 2012; Zhang and Schekman, 2013). Pro-IL-1b is subsequently converted into the active form, the 17 kDa mature IL-1b, by the pro-inflammatory protease caspase-1 which is activated, in response to extracellular stimuli, after its recruitment to a multi-protein complex called the inflammasome (Burns et al, 2003; Cerretti et al, 1992; Rathinam et al, 2012; Thornberry et al, 1992). Many reports including two recent publications make the case for cell lysis

Methods

Results

Conclusion