An Intrinsic Link between the Metabolic and Antiviral States of the Cell

Division Of Basic Sciences - Nci

Investigators

Linked publications & trials

Abstract

Since work on this project began in February 2017, we have made a number of key advancements. Our preliminary work using rapamycin on transformed epithelial cell lines has revealed that mTOR inhibition confers a 4- to 20-fold enhancement of infection by lentiviral vectors and by Influenza A virus. Furthermore, we found that the rapamycin-dependent enhancement of infection is reversed by inhibitors of endosomal acidification (v-ATPase), revealing that the enhancement requires active degradation of cellular factors via the lysosomal pathway. Through a number of distinct approaches, we show that mTOR inhibition by multiple drugs leads to lysosomal degradation of IFITM3 in an autophage-independent manner. Instead, endocytic trafficking through multivesicular bodies is necessary to delivery of IFITM3 to lysosomes, as confirmed by a functional requirement of ESCRT member TSG101. By studying mutant IFITM3 constructs, we found that mTOR inhibition leads to clearance of IFITM2 and IFITM3 from endosomes in a manner that is dependent on endocytosis, ubiquitination, and lysosomal acidification. Furthermore, the complex including mTOR that functionally modulates IFITM3 levels is mTORC2. This work is the first instance to describe an interrelationship between mTOR, cell-intrinsic antiviral immunity, and virus entry into cells. These results have been published in 2018 (Shi et al., PNAS 115: E10069, 2018). We have now extended the results to include impacts on HIV-1 infection, which involves the study of virus entry mediated by HIV-1 Env glycoprotein. We have assessed the impact of rapamycin and other drugs on various cell types that serve as HIV-1 targets in vivo and we found that IFITM3 is also downregulated in these cells. The loss of IFITM3 from macrophages, for example, allows greater degrees of infection by multiple HIV-1 strains. _____More recently, we have compared the ability of rapalogs to downmodulate IFITM proteins and to enhance virus infections, including SARS-CoV-2. We found that some rapalogs perform these functions while others do not, laying the groundwork for a mechanistic understanding of the cellular pathways involved. We are also working on a study to resolve the relationship between the antiviral proteins IFITM and components of the PI3K/Akt/mTOR pathway. We hypothesized that IFITM3 is a positive regulator of PI3K/Akt/mTOR signaling due to the fact that, when mTOR is inhibited by rapamycin, IFITM3 is selectively degraded. Our initial observations indicate that endogenous IFITM3 in some cell types is necessary for full activation of Akt. Specifically, we found that knockdown of IFITM3 resulted in decreased phosphorylation of Akt at serine-473. These findings highlight a previously unrecognized housekeeping role for IFITM3 in cellular homeostasis. The significance of this finding is reinforced by the fact that IFITM3 is commonly upregulated in a variety of cancers. Thus, this project has provided an opportunity for my lab to explore new avenues with relevance to the basic and clinical understanding of tumorigenesis.