R01AI161363

Mechanism-Based Targeting of the RNA Processing Machinery of SARS-CoV-2 - Abstract

The massive global pandemic with high morbidity and mortality makes Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) one of the deadliest viruses in recent history. It is especially noteworthy for hijacking the normal operations of human cells. To develop effective therapies, we need a better understanding of the mechanisms that permit the virus to invade cells and evade host immune restriction.

SARS-CoV-2 encodes the non-structural protein (NSP)16/NSP10 protein complex that transfers a methyl group from S-adenosyl methionine (SAM) to 2’-OH of the first transcribing nucleotide of the viral mRNA and thus converts the cap-0 (m7GpppA) to cap-1 (m7GpppAm). The resulting viral mRNA mimics host cell’s mRNA. In this way, a cell cannot distinguish between its own RNA and that of the virus. This modification of the virally encoded mRNA not only tricks the immune system and helps the virus to take over the host translation machinery for synthesis of its own proteins for survival and propagation. Ablation of NSP16 activity should trigger an immune response to viral infection and limit pathogenesis.

Our recent paper in Nature Communications described atomic level details of the NSP16/NSP10 complex and how the enzyme is well adapted to bind the RNA cap and exert the 2’-OH methylation. We also discovered a distant pocket (located 25Å away from the catalytic center) in NSP16 that is unique to SARS-CoV-2. We also found that this pocket in NSP16 is partially composed of amino acids that are unique to SARS-CoV-2. It can bind small molecules outside of the catalytic center.

We propose to build a long-term research program aimed at deciphering the factors crucial to the maintenance of RNA genome and evasion from the host’s immune response. Our studies will reveal basic principles underlying SARS-CoV-2 RNA cap modification, the mode of nucleoprotein (NP) assembly, interplay with mRNA, and new approaches for therapeutic targeting.

In Aim 1, we will resolve a series of new structures of NSP16/NSP10 proteins captured in every step of the methyl transfer by X-ray crystallography. The structural data will be validated by detailed biochemical and biophysical studies. We will resolve the biochemical and structural determinants of the assembly of viral RNA capping machinery and identify factors underlying integrity of RNA genome.

In Aim 2, we will develop a novel molecular tool to study temporal distribution of the RNA methylation during viral infection. We will examine new models for combinatorial inhibition of viral proteins by drug repurposing or novel small molecules.

Finally, we will use our recently established reverse genetics approaches based on the use of a bacterial artificial chromosome (BAC) to generate recombinant (R)SARS-CoV2 containing mutations in NSP16 to determine their contribution in viral replication in cultured cells and pathogenesis in vivo using our recently described K18 human angiotensin converting enzyme 2 (hACE2) mouse model of SARS-CoV-2 infection and associated Coronavirus Disease 2019 (COVID-19).

The University Of Texas Health Science Center At San Antonio

TO ASSIST PUBLIC AND PRIVATE NONPROFIT INSTITUTIONS AND INDIVIDUALS TO ESTABLISH, EXPAND AND IMPROVE BIOMEDICAL RESEARCH AND RESEARCH TRAINING IN INFECTIOUS DISEASES AND RELATED AREAS, TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS. TO ASSIST PUBLIC, PRIVATE AND COMMERCIAL INSTITUTIONS TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS, TO PROVIDE RESEARCH SERVICES AS REQUIRED BY THE AGENCY FOR PROGRAMS IN INFECTIOUS DISEASES, AND CONTROLLING DISEASE CAUSED BY INFECTIOUS OR PARASITIC AGENTS, ALLERGIC AND IMMUNOLOGIC DISEASES AND RELATED AREAS. PROJECTS RANGE FROM STUDIES OF MICROBIAL PHYSIOLOGY AND ANTIGENIC STRUCTURE TO COLLABORATIVE TRIALS OF EXPERIMENTAL DRUGS AND VACCINES, MECHANISMS OF RESISTANCE TO ANTIBIOTICS AS WELL AS RESEARCH DEALING WITH EPIDEMIOLOGICAL OBSERVATIONS IN HOSPITALIZED PATIENTS OR COMMUNITY POPULATIONS AND PROGRESS IN ALLERGIC AND IMMUNOLOGIC DISEASES. BECAUSE OF THIS DUAL FOCUS, THE PROGRAM ENCOMPASSES BOTH BASIC RESEARCH AND CLINICAL RESEARCH. SMALL BUSINESS INNOVATION RESEARCH (SBIR) PROGRAM EXPANDS AND IMPROVES PRIVATE SECTOR PARTICIPATION IN BIOMEDICAL RESEARCH. THE SBIR PROGRAM INTENDS TO INCREASE AND FACILITATE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, TO INCREASE SMALL BUSINESS PARTICIPATION IN FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. THE SMALL BUSINESS TECHNOLOGY TRANSFER (STTR) PROGRAM STIMULATES AND FOSTERS SCIENTIFIC AND TECHNOLOGICAL INNOVATION THROUGH COOPERATIVE RESEARCH AND DEVELOPMENT CARRIED OUT BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO FOSTER TECHNOLOGY TRANSFER BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO INCREASE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. RESEARCH CAREER DEVELOPMENT AWARDS SUPPORT THE DEVELOPMENT OF SCIENTISTS DURING THE FORMATIVE STAGES OF THEIR CAREERS. INDIVIDUAL NATIONAL RESEARCH SERVICE AWARDS (NRSAS) ARE MADE DIRECTLY TO APPROVE APPLICANTS FOR RESEARCH TRAINING IN SPECIFIED BIOMEDICAL SHORTAGE AREAS. IN ADDITION, INSTITUTIONAL NATIONAL RESEARCH SERVICE AWARDS ARE MADE TO ENABLE INSTITUTIONS TO SELECT AND MAKE AWARDS TO INDIVIDUALS TO RECEIVE TRAINING UNDER THE AEGIS OF THEIR INSTITUTIONAL PROGRAM.

93.855 - Allergy and Infectious Diseases Research

National Institute of Allergy and Infectious Diseases (NIAID) [HHS - NIH]

San Antonio, Texas 782293904 United States

Single Zip Code

Emergency Awards: Rapid Investigation of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and Coronavirus Disease 2019 (COVID-19) (R01 Clinical Trial Not Allowed) (PAR-20-178)

COVID-19 $1,210,982 (39%) percent of this Project Grant was funded by COVID-19 emergency acts including the American Rescue Plan Act of 2021.
Amendment Since initial award the total obligations have increased 406% from $608,161 to $3,079,573.