Research Interests
Over the years, Prof. Rizvi’s laboratory has focused on studying the molecular steps involved in the replication of retroviruses, including genomic RNA (gRNA) packaging, dimerization, and export using a number of different retroviruses, including human, simian, and feline immunodeficiency viruses (HIV, SIV, & FIV), Mason-Pfizer monkey virus (MPMV), and mouse mammary tumor virus (MMTV). The overall goal of these efforts has been: 1) enhancement of our understanding of retrovirus replication & regulation of gene expression, 2) improvements in the design and development of retroviral/lentiviral vectors and packaging cells lines for safe and efficient human gene therapy, 3) testing of novel and classical vaccines and antivirals against HIV/AIDS, and finally, 4) development of innovative methods to detect, quantify, & identify viruses and other biological materials based on electrical parameters. Post COVID-19 pandemic, his laboratory started applying the expertise learned from retroviruses to determine how one can expedite the study of SARS-CoV-2 replication, the etiological agent of COVID-19, with the hope of identifying therapeutic agents for this devastating pandemic. A summary of the research undertaken over the years is provided below.
Structural basis of retroviral Genomic RNA (gRNA) export, dimerization, and packaging
Genomic RNA (gRNA) packaging is a hallmark of retroviral life cycle where two copies of the gRNA, most likely in their dimeric form, are encapsidated into the virus particles. The specificity towards gRNA packaging is conferred by the recognition of specific cis-acting sequences, the packaging signal, present at the 5’ end of the viral genome (that assumes a higher order structure), which interacts with the Gag protein. It has also been proposed that dimerization leads to conformational changes, exposing single-stranded Gag binding motifs (generally purine-rich sequence), facilitating Gag binding during recruitment of the gRNA for packaging. Using a combination of in vivo genetic complementation assays, in vitro biochemical probing/mapping-SHAPE, and structural prediction/phylogenetic approaches, Prof. Rizvi’s laboratory has mapped the structural determinants of gRNA packaging and dimerization in SIV, FIV, MMTV, and MPMV as well as cross/co-packaging among retroviruses. They have taken this work to the next level by identifying Gag binding sites on the gRNA for FIV, MMTV, and MPMV, while work is in progress for SIV. This has been accomplished by expressing full-length Gag proteins from these retroviruses and testing their RNA binding potential combined with footprinting assays based on SHAPE. Expression of full-length retroviral Gag is a feat in itself that so far has only been achieved for HIV-1 due to the toxic nature of these proteins. Their ongoing research efforts are directed towards delineating Gag-gRNA interactions that distinguish gRNA substrates for dimerization and subsequent RNA packaging from those for Gag/Pol translation. Recent studies are directed towards identifying the MMTV Rem binding sites on the Rem Responsive Element (RmRE), an RNA-protein interaction that is critical for the nuclear export of unspliced gRNA.
Molecular mechanism(s) of SARS-CoV-2 gRNA packaging and SARS-CoV-2 host interactions
SARS-CoV-2 caused the COVID-19 pandemic; therefore, understanding the replication process of SARS-CoV-2 is critical to develop novel therapeutics and better vaccines against COVID-19. How SARS-CoV-2 “recognizes” and “incorporates” its RNA genome (gRNA) into the newly assembling/forming viral particles is crucial for understanding and thereby controlling virus transmission. Viral particle formation involves recognition and interaction of sequences constituting the “packaging signal” (PS) on the gRNA with the nucleocapsid (N) and membrane (M) proteins of the virus, resulting in gRNA packaging/incorporation into the virus particles. However, not much is known about how SARS-CoV-2) packages its gRNA, a crucial aspect to continue its life cycle. Prof. Rizvi’s laboratory is working towards defining the initial recognition of gRNA by N & M proteins employing a combination of in vitro biochemical, structural, and in vivo genetic approaches. These approaches should map the SARS-CoV-2 packaging signal, define and validate its higher order structure, and identify specific nucleotides that interact with N & M protein during gRNA packaging. Furthermore, these studies should pave the way for high resolution structural analysis of the N & M protein-RNA interaction(s), which can be used for rational drug design as therapeutic interventions against SARS-CoV-2. Finally, Prof. Rizvi’s laboratory has identified miRNAs induced in COVID-19 patients during SARS-CoV-2 infection, diseased stages, and those that target the viral genome.
Development of retroviral vectors for human gene therapy
One practical application of studying retroviral replication has been the development of retroviral vectors for human gene therapy. Thus, Prof. Rizvi has been successful in creating split-genome viral replication assays not only for HIV-1 and SIV, but also FIV, MMTV, and MPMV. These assays have allowed them to not only study retrovirus replication in a highly sensitive and specific manner but also allow transduction of human cells with marker genes successfully without the need for biosafety level 3 (BSL-3) facilities. This work has been extended to MMTV to develop safer self-inactivating (SIN) vectors for human gene therapy. These vectors have been tested both in cell culture and the mouse model system for gene delivery.
Vaccine/antiviral strategies against HIV/AIDS
The Rizvi laboratory has been involved in facilitating the development of vaccine and novel antiviral strategies against HIV/AIDS. These strategies have used either the more novel DNA-based approach, in collaboration with Prof. H. L. Robinson (Emory University), the guru of DNA vaccines, or the more classical passive immunization approach, in collaboration with Prof. R. M. Ruprecht, (Harvard University). In both cases, the simian-human (SHIV)-monkey model system was used for these studies. The passive immunization approach was used in neonatal monkeys (macaques) using human monoclonal antibodies to study protection against mother-to-child transmission and oral challenge, while the DNA vaccines were tested in combination with protein boost to study the development of protective neutralizing antibody response against challenge with live virus. These studies have led to seminal contributions towards the development of vaccines as potential therapeutics against HIV/AIDS. In addition, his laboratory has also been involved in the development of novel antiviral agents from the virus itself by exploiting its replicative biology.
Detection, quantification, and identification of viruses and other biological materials based on electrical parameters
Most of the existing techniques for viral screening and quantification suffer from limitations due to the need for extensive sample preparation and labeling, which is fairly costly, requiring a great deal of time. Therefore, in collaboration with Dr. Mahmoud Al Ahmad (Department of Electrical Engineering, UAE University), Prof. Rizvi’s laboratory has been working towards developing novel label-free virus screening and quantification techniques based on electrical parameters. These studies have demonstrated that viruses can in fact be detected, quantified, and identified within minutes in tissue culture medium without the need for any protein labeling or amplification strategies and have resulted in several pivotal publications in this area. More recently, the scope of these studies has been expanded by characterizing other biological molecules, such as nucleotides (A, C, T, G) and DNA, virus-expressing cells, cancer cells, as well as blood cells in urine based on their electrical properties. It is anticipated that the electrical approach is just a starting point towards establishing the foundation for the electrical-based identification and quantification of unlimited number of biological materials, and nano-sized particles, including SARS-CoV-2.