Abstract

Previously, we reported that the mitochondrial translocator protein (TSPO) induces HIV-1 envelope (Env) degradation via the endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway, but the mechanism was not clear. Here we investigated how the four ER-associated glycoside hydrolase family 47 (GH47) α-mannosidases, ERManI, and ER-degradation enhancing α-mannosidase-like (EDEM) proteins 1, 2, and 3, are involved in the Env degradation process. Ectopic expression of these four α-mannosidases uncovers that only ERManI inhibits HIV-1 Env expression in a dose-dependent manner. In addition, genetic knock-out of the ERManI gene MAN1B1 using CRISPR/Cas9 technology disrupts the TSPO-mediated Env degradation. Biochemical studies show that HIV-1 Env interacts with ERManI, and between the ERManI cytoplasmic, transmembrane, lumenal stem, and lumenal catalytic domains, the catalytic domain plays a critical role in the Env-ERManI interaction. In addition, functional studies show that inactivation of the catalytic sites by site-directed mutagenesis disrupts the ERManI activity. These studies identify ERManI as a critical GH47 α-mannosidase in the ER-associated protein degradation pathway that initiates the Env degradation and suggests that its catalytic domain and enzymatic activity play an important role in this process.

Highlights

  • HIV-1 envelope (Env) glycoprotein is targeted to endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway for degradation after infecting cells

  • Treatment of these infected cells with an ERAD inhibitor kifunensine (KIF) significantly increased the Env expression in N2-NP cells (Fig. 2, C and D); KIF increased HIV-1 replication in N2-NP cells but not in N5-P cells (Fig. 2E). These results suggest that Env is degraded via ERAD, which is responsible for HIV-1 inhibition in N2-NP cells

  • In this report we studied the molecular mechanism of TSPOinduced HIV-1 Env degradation via ERAD and identified ER class I ␣-mannosidase (ERManI) as a critical initiator for the degradation

Read more

Summary

Introduction

HIV-1 envelope (Env) glycoprotein is targeted to endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway for degradation after infecting cells. We investigated how the four ER-associated glycoside hydrolase family 47 (GH47) ␣-mannosidases, ERManI, and ER-degradation enhancing ␣-mannosidase-like (EDEM) proteins 1, 2, and 3, are involved in the Env degradation process. Ectopic expression of these four ␣-mannosidases uncovers that only ERManI inhibits HIV-1 Env expression in a dose-dependent manner. Functional studies show that inactivation of the catalytic sites by site-directed mutagenesis disrupts the ERManI activity These studies identify ERManI as a critical GH47 ␣-mannosidase in the ER-associated protein degradation pathway that initiates the Env degradation and suggests that its catalytic domain and enzymatic activity play an important role in this process

Objectives
Results
Conclusion
Full Text
Published version (Free)

Talk to us

Join us for a 30 min session where you can share your feedback and ask us any queries you have

Schedule a call