Our Research

Effect of lncRNA on HIV disease. 

Less than 3% of the entire transcriptome is protein coding, signifying that non-coding RNAs represent most of the human transcriptome. Long non-coding RNA transcripts, termed lncRNAs (>200bp) have been identified in mammalian genomes by analysis of transcriptomic data, and while thousands of lncRNAs are known, not many of their functions have been identified. The lincRNAs have been implicated in the regulation of many cellular and developmental processes such as imprinting, dosage compensation, cell cycle regulation, pluripotency, retrotransposon silencing, meiotic entry, and telomere lengthening. My lab is interested in the role of lncRNAs in HIV replication and latency.

We have identified polymorphisms in the lncRNA genes that associate with HIV viral load control. These lncRNAs could potentially affect HIV pathogenesis and could represent the first example of a variation in lncRNA expression that affects disease; we are further investigating the function of inter-genic loci that mark diversity in disease outcomes.

  • Kulkarni, Viraj, Sahana Jayakumar, Mahesh Mohan, and Smita Kulkarni. (2023) 2023. “Aid or Antagonize: Nuclear Long Noncoding RNAs Regulate Host Responses and Outcomes of Viral Infections.”. Cells 12 (7). https://doi.org/10.3390/cells12070987.
    Publisher's Version

    Long noncoding RNAs (lncRNAs) are transcripts measuring >200 bp in length and devoid of protein-coding potential. LncRNAs exceed the number of protein-coding mRNAs and regulate cellular, developmental, and immune pathways through diverse molecular mechanisms. In recent years, lncRNAs have emerged as epigenetic regulators with prominent roles in health and disease. Many lncRNAs, either host or virus-encoded, have been implicated in critical cellular defense processes, such as cytokine and antiviral gene expression, the regulation of cell signaling pathways, and the activation of transcription factors. In addition, cellular and viral lncRNAs regulate virus gene expression. Viral infections and associated immune responses alter the expression of host lncRNAs regulating immune responses, host metabolism, and viral replication. The influence of lncRNAs on the pathogenesis and outcomes of viral infections is being widely explored because virus-induced lncRNAs can serve as diagnostic and therapeutic targets. Future studies should focus on thoroughly characterizing lncRNA expressions in virus-infected primary cells, investigating their role in disease prognosis, and developing biologically relevant animal or organoid models to determine their suitability for specific therapeutic targeting. Many cellular and viral lncRNAs localize in the nucleus and epigenetically modulate viral transcription, latency, and host responses to infection. In this review, we provide an overview of the role of nuclear lncRNAs in the pathogenesis and outcomes of viral infections, such as the Influenza A virus, Sendai Virus, Respiratory Syncytial Virus, Hepatitis C virus, Human Immunodeficiency Virus, and Herpes Simplex Virus. We also address significant advances and barriers in characterizing lncRNA function and explore the potential of lncRNAs as therapeutic targets.

  • Premadasa, Lakmini S, Eunhee Lee, Marina McDew-White, Xavier Alvarez, Sahana Jayakumar, Binhua Ling, Chioma M Okeoma, Siddappa N Byrareddy, Smita Kulkarni, and Mahesh Mohan. (2023) 2023. “Cannabinoid Enhancement of LncRNA MMP25-AS1/MMP25 Interaction Reduces Neutrophil Infiltration and Intestinal Epithelial Injury in HIV/SIV Infection.”. JCI Insight 8 (7). https://doi.org/10.1172/jci.insight.167903.
    Publisher's Version

    Intestinal epithelial barrier dysfunction, a hallmark of HIV/SIV infection, persists despite viral suppression by combination antiretroviral therapy (cART). Emerging evidence suggests a critical role for long noncoding RNAs (lncRNAs) in maintaining epithelial homeostasis. We simultaneously profiled lncRNA/mRNA expression exclusively in colonic epithelium (CE) of SIV-infected rhesus macaques (RMs) administered vehicle (VEH) or Δ-9-tetrahydrocannabinol (THC). Relative to controls, fewer lncRNAs were up- or downregulated in CE of THC/SIV compared with VEH/SIV RMs. Importantly, reciprocal expression of the natural antisense lncRNA MMP25-AS1 (up 2.3-fold) and its associated protein-coding gene MMP25 (attracts neutrophils by inactivating alpha-1 anti-trypsin/SERPINA1) (down 2.2-fold) was detected in CE of THC/SIV RMs. Computational analysis verified 2 perfectly matched complementary regions and an energetically stable (normalized binding free energy = -0.2626) MMP25-AS1/MMP25 duplex structure. MMP25-AS1 overexpression blocked IFN-γ-induced MMP25 mRNA and protein expression in vitro. Elevated MMP25 protein expression in CE of VEH/SIV but not THC/SIV RMs was associated with increased infiltration by myeloperoxidase/CD11b++ neutrophils (transendothelial migration) and epithelial CD47 (transepithelial migration) expression. Interestingly, THC administered in combination with cART increased MMP25-AS1 and reduced MMP25 mRNA/protein expression in jejunal epithelium of SIV-infected RMs. Our findings demonstrate that MMP25-AS1 is a potentially unique epigenetic regulator of MMP25 and that low-dose THC can reduce neutrophil infiltration and intestinal epithelial injury potentially by downregulating MMP25 expression through modulation of MMP25-AS1.

  • Kulkarni, Smita, Janice J Endsley, Zhao Lai, Todd Bradley, and Riti Sharan. (2023) 2023. “Single-Cell Transcriptomics of Mtb/HIV Co-Infection.”. Cells 12 (18). https://doi.org/10.3390/cells12182295.
    Publisher's Version

    Tuberculosis (TB) and Human Immunodeficiency Virus (HIV) co-infection continues to pose a significant healthcare burden. HIV co-infection during TB predisposes the host to the reactivation of latent TB infection (LTBI), worsening disease conditions and mortality. There is a lack of biomarkers of LTBI reactivation and/or immune-related transcriptional signatures to distinguish active TB from LTBI and predict TB reactivation upon HIV co-infection. Characterizing individual cells using next-generation sequencing-based technologies has facilitated novel biological discoveries about infectious diseases, including TB and HIV pathogenesis. Compared to the more conventional sequencing techniques that provide a bulk assessment, single-cell RNA sequencing (scRNA-seq) can reveal complex and new cell types and identify more high-resolution cellular heterogeneity. This review will summarize the progress made in defining the immune atlas of TB and HIV infections using scRNA-seq, including host-pathogen interactions, heterogeneity in HIV pathogenesis, and the animal models employed to model disease. This review will also address the tools needed to bridge the gap between disease outcomes in single infection vs. co-infection. Finally, it will elaborate on the translational benefits of single-cell sequencing in TB/HIV diagnosis in humans.

Role of alternative 3’UTRs in gene regulation and diseases. 

Several human genes utilize alternative polyadenylation to generate transcript isoforms with varying lengths of 3’untranslated regions (3’UTR). 3’UTRs encode docking sites for regulatory RNA binding proteins and microRNAs, and thus are major determinants of post-transcriptional gene regulation. Alternative 3’UTR usage is extensively modulated in development, differentiation, proliferation, and neuron activation; alternative 3’UTRs could contribute to both gene expression and protein diversity.

Mutations in polyadenylation signals and other poly(A) cis-elements that lead to changes in gene expression can contribute to the development of human genetic diseases. A dysregulation of alternative 3’UTRs plays a role in cardiac hypertrophy and tumor progression. Manipulation of the length of 3’UTR can alter gene regulation and is considered promising for future therapeutic blocking of angiogenic signals in tumors.

Even though over 70% of human genes contain multiple poly(A) sites and could potentially be subjected to alternative 3’UTR regulation, it is not known what percentage is actually regulated by alternative 3’UTR and whether this results in any biological effects. We are studying alternative 3’UTRs of immune genes that may have an effect on immune response diversity and subsequent outcomes of infectious and autoimmune diseases.

  • Kulkarni, Viraj, Sahana Jayakumar, Mahesh Mohan, and Smita Kulkarni. (2023) 2023. “Aid or Antagonize: Nuclear Long Noncoding RNAs Regulate Host Responses and Outcomes of Viral Infections.”. Cells 12 (7). https://doi.org/10.3390/cells12070987.
    Publisher's Version

    Long noncoding RNAs (lncRNAs) are transcripts measuring >200 bp in length and devoid of protein-coding potential. LncRNAs exceed the number of protein-coding mRNAs and regulate cellular, developmental, and immune pathways through diverse molecular mechanisms. In recent years, lncRNAs have emerged as epigenetic regulators with prominent roles in health and disease. Many lncRNAs, either host or virus-encoded, have been implicated in critical cellular defense processes, such as cytokine and antiviral gene expression, the regulation of cell signaling pathways, and the activation of transcription factors. In addition, cellular and viral lncRNAs regulate virus gene expression. Viral infections and associated immune responses alter the expression of host lncRNAs regulating immune responses, host metabolism, and viral replication. The influence of lncRNAs on the pathogenesis and outcomes of viral infections is being widely explored because virus-induced lncRNAs can serve as diagnostic and therapeutic targets. Future studies should focus on thoroughly characterizing lncRNA expressions in virus-infected primary cells, investigating their role in disease prognosis, and developing biologically relevant animal or organoid models to determine their suitability for specific therapeutic targeting. Many cellular and viral lncRNAs localize in the nucleus and epigenetically modulate viral transcription, latency, and host responses to infection. In this review, we provide an overview of the role of nuclear lncRNAs in the pathogenesis and outcomes of viral infections, such as the Influenza A virus, Sendai Virus, Respiratory Syncytial Virus, Hepatitis C virus, Human Immunodeficiency Virus, and Herpes Simplex Virus. We also address significant advances and barriers in characterizing lncRNA function and explore the potential of lncRNAs as therapeutic targets.

  • Premadasa, Lakmini S, Eunhee Lee, Marina McDew-White, Xavier Alvarez, Sahana Jayakumar, Binhua Ling, Chioma M Okeoma, Siddappa N Byrareddy, Smita Kulkarni, and Mahesh Mohan. (2023) 2023. “Cannabinoid Enhancement of LncRNA MMP25-AS1/MMP25 Interaction Reduces Neutrophil Infiltration and Intestinal Epithelial Injury in HIV/SIV Infection.”. JCI Insight 8 (7). https://doi.org/10.1172/jci.insight.167903.
    Publisher's Version

    Intestinal epithelial barrier dysfunction, a hallmark of HIV/SIV infection, persists despite viral suppression by combination antiretroviral therapy (cART). Emerging evidence suggests a critical role for long noncoding RNAs (lncRNAs) in maintaining epithelial homeostasis. We simultaneously profiled lncRNA/mRNA expression exclusively in colonic epithelium (CE) of SIV-infected rhesus macaques (RMs) administered vehicle (VEH) or Δ-9-tetrahydrocannabinol (THC). Relative to controls, fewer lncRNAs were up- or downregulated in CE of THC/SIV compared with VEH/SIV RMs. Importantly, reciprocal expression of the natural antisense lncRNA MMP25-AS1 (up 2.3-fold) and its associated protein-coding gene MMP25 (attracts neutrophils by inactivating alpha-1 anti-trypsin/SERPINA1) (down 2.2-fold) was detected in CE of THC/SIV RMs. Computational analysis verified 2 perfectly matched complementary regions and an energetically stable (normalized binding free energy = -0.2626) MMP25-AS1/MMP25 duplex structure. MMP25-AS1 overexpression blocked IFN-γ-induced MMP25 mRNA and protein expression in vitro. Elevated MMP25 protein expression in CE of VEH/SIV but not THC/SIV RMs was associated with increased infiltration by myeloperoxidase/CD11b++ neutrophils (transendothelial migration) and epithelial CD47 (transepithelial migration) expression. Interestingly, THC administered in combination with cART increased MMP25-AS1 and reduced MMP25 mRNA/protein expression in jejunal epithelium of SIV-infected RMs. Our findings demonstrate that MMP25-AS1 is a potentially unique epigenetic regulator of MMP25 and that low-dose THC can reduce neutrophil infiltration and intestinal epithelial injury potentially by downregulating MMP25 expression through modulation of MMP25-AS1.

  • Kulkarni, Smita, Janice J Endsley, Zhao Lai, Todd Bradley, and Riti Sharan. (2023) 2023. “Single-Cell Transcriptomics of Mtb/HIV Co-Infection.”. Cells 12 (18). https://doi.org/10.3390/cells12182295.
    Publisher's Version

    Tuberculosis (TB) and Human Immunodeficiency Virus (HIV) co-infection continues to pose a significant healthcare burden. HIV co-infection during TB predisposes the host to the reactivation of latent TB infection (LTBI), worsening disease conditions and mortality. There is a lack of biomarkers of LTBI reactivation and/or immune-related transcriptional signatures to distinguish active TB from LTBI and predict TB reactivation upon HIV co-infection. Characterizing individual cells using next-generation sequencing-based technologies has facilitated novel biological discoveries about infectious diseases, including TB and HIV pathogenesis. Compared to the more conventional sequencing techniques that provide a bulk assessment, single-cell RNA sequencing (scRNA-seq) can reveal complex and new cell types and identify more high-resolution cellular heterogeneity. This review will summarize the progress made in defining the immune atlas of TB and HIV infections using scRNA-seq, including host-pathogen interactions, heterogeneity in HIV pathogenesis, and the animal models employed to model disease. This review will also address the tools needed to bridge the gap between disease outcomes in single infection vs. co-infection. Finally, it will elaborate on the translational benefits of single-cell sequencing in TB/HIV diagnosis in humans.

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