Research

Exploring host-pathogen interactions in HEV and SARS-CoV-2 infected cells

Host-pathogen interactions determine the survival or extinction of the host and the pathogen. This eternal conflict between the host and the pathogen is a major driving force in their evolution. Therefore, a thorough understanding of the molecular details of the host-pathogen conflicts are crucial for decoding their survival skills and developing specific therapeutics against the pathogen.

Establishment and analysis of HEV-host protein interaction network

Toward understanding the molecular details of HEV life cycle, we screened human liver and fetal brain cDNA libraries to identify the host interaction partners of proteins encoded by genotype 1 HEV and constructed the virus-host PPI network. Analysis of the network indicated a role of HEV proteins in modulating multiple host biological processes such as stress and immune responses, the ubiquitin-proteasome system, energy and iron metabolism, and protein translation. Further investigations revealed the presence of multiple host translation regulatory factors in the viral translation/ replication complex. Depletion of host translation factors such as eIF4A2, eIF3A, and RACK1 signiﬁcantly reduced the viral replication, whereas eIF2AK4 depletion had no effect. These ﬁndings highlight the ingenuity of the pathogen in manipulating the host machinery to its own beneﬁt, a clear understanding of which is essential for the identiﬁcation of strategic targets and development of speciﬁc antivirals against HEV.

Deciphering the antiviral role of host Lamp2a protein against SARS-CoV-2

Replication of a positive-strand RNA virus involves an RNA-protein complex consisting of viral genomic RNA, host RNA(s), virus-encoded proteins, and host proteins. Dissecting out individual components of the replication complex will help decode the mechanism of viral replication. 5'- and 3'-UTRs in positive-strand RNA viruses play essential regulatory roles in virus replication. Using an RNA-protein interaction detection (RaPID) assay coupled to liquid chromatography with tandem mass spectrometry, we identiﬁed host interaction partners of SARS-CoV-2 5'- and 3'-UTRs and generated an RNA-protein interaction network. By combining these data with the previously known protein-protein interaction data proposed to be involved in virus replication, we generated the RNA-proteinprotein interaction (RPPI) network, likely to be essential for controlling SARS-CoV-2 replication. Notably, bioinformatics analysis of the RPPI network revealed the enrichment of factors involved in translation initiation and RNA metabolism. Lysosome-associated membrane protein-2a (Lamp2a), the receptor for chaperone-mediated autophagy, is one of the host proteins that interact with the 5'-UTR. Further studies showed that the Lamp2 level is upregulated in SARS-CoV-2-infected cells and that the absence of the Lamp2a isoform enhanced the viral RNA level whereas its overexpression signiﬁcantly reduced the viral RNA level. Lamp2a and viral RNA colocalize in the infected cells, and there is an increased autophagic ﬂux in infected cells, although there is no change in the formation of autophagolysosomes. In summary, our study provides a useful resource of SARS-CoV-2 5'- and 3'-UTR binding proteins and reveals the antiviral role of Lamp2a protein during SARS-CoV-2 infection.

Deciphering the antiviral role of HERV-R -envelope against SARS-CoV-2

Human endogenous retroviruses (HERVs) represent retroviral elements that were integrated into the ancestral human genome. HERVs are important in embryonic development as well as in the manifestation of diseases, including cancer, inflammation, and viral infections. We observed down regulation of HERV-R envelope in cell-based models and PBMCs of COVID-19 patients. Overexpression of HERV-R inhibits SARS-CoV-2 replication, suggesting its antiviral activity. Further analyses demonstrate the role of the extracellular signal-regulated kinase (ERK) in regulating HERV-R antiviral activity. Lastly, we demonstrate that the crosstalk between ERK and p38 MAPK controls the synthesis of the HERV-R envelope protein, which in turn modulates SARSCoV-2 replication. These findings suggest the role of the HERV-R envelope as a prosurvival host factor against SARS-CoV-2 and illustrate a possible advantage of integration and evolutionary maintenance of retroviral elements in the human genome.

Viral vaccines

Viral hepatitis is a major public health concern and WHO aims at eliminating it by 2030. Hepatitis E virus (HEV) is a major cause of acute viral hepatitis (AVH) and community level outbreaks. It may also cause chronic and fulminant hepatitis. The disease worsens during pregnancy, with a 20-25% mortality rate. Recent reports suggest chronic HEV infection in ~60% of liver transplantation patients. My laboratory is engaged in developing a recombinant virus-like particle (VLP)-based vaccine against the HEV.

We have also established an mRNA vaccine development platform to address the requirement of vaccines against pathogens of human medical importance. The platform utilizes the sequence information of the potential vaccine antigens of any pathogen to produce a series of lipid nanoparticle (LNP) encapsulated-mRNA vaccine formulations, which are evaluated in cell line-based and small animal model systems to identify the best vaccine antigen. Proof of function of the mRNA vaccine development platform was shown in our laboratory by producing and evaluating vaccine candidates against different variants of SARS-CoV-2. Our experimental conditions produced circular mRNA (cRNA)-LNP vaccine formulations encoding RBD of Wuhan, Delta and Omicron (BA.4/5) variants of SARS-CoV-2, with an average particle size of 136nm, PDI of 0.05 and 96% cRNA encapsulation efficiency. The average end point IgG (anti-RBD) and Neutralization (by PRNT assay) titer of the different RBD variants was in the range of 104-105 and 103-104, respectively. Moreover, single variant vaccine formulations as well as a bivalent cRNA vaccine formulation [encoding RBD of Delta and Omicron (BA.4/5) variants] protected against corresponding SARS-CoV-2 variant challenges in mouse model of COVID-19, thus confirming the functional efficacy of the vaccine formulations.