• RNA plasticity
• RNA structure
• VIGS
• positional entropy
• virus-induced gene silencing

• adaptive evolution
• antiviral RNA interference
• bat
• bat fly
• ectoparasites
• host–pathogen interactions
• virus–host dynamics

• Antiviral
• Gene silencing: mRNA
• siRNA

• RNA, Small Interfering/therapeutic use
• RNA, Small Interfering/genetics
• Antiviral Agents/pharmacology
• Antiviral Agents/therapeutic use
• Humans
• Animals
• Virus Diseases/drug therapy
• Viruses/drug effects
• Viruses/genetics
• Gene Silencing
• RNA Interference
• Clinical Trials as Topic

• CRISPR-Cas13d
• MT: RNA/DNA Editing
• RNA interference
• RNA structure
• SARS-CoV-2
• cleavage activity
• gene editing
• targeting efficiency

• HIGS
• MIGS
• RNAi
• RNAi inheritance
• SIGS

• 32230003 the National Science Foundation of China
• 2022DB014 Science and Technology Bureau of Xinjiang Production and Construction Corps

• Antiviral
• RNA interference
• Small interfering RNA
• Therapeutic
• Virus

• Humans
• RNA, Small Interfering/genetics
• RNA, Small Interfering/therapeutic use
• RNA, Small Interfering/metabolism
• Pandemics
• COVID-19/genetics
• COVID-19/therapy
• RNA Interference
• Viruses/genetics
• Viruses/metabolism
• Antiviral Agents/therapeutic use
• RNA, Viral

• Incheon National University

• RNA/genetics
• RNA, Catalytic/genetics
• RNA, Catalytic/chemistry
• Riboswitch
• Transcriptome
• Alternative Splicing

• Humans
• SARS-CoV-2/genetics
• SARS-CoV-2/metabolism
• Antiviral Agents/pharmacology
• COVID-19
• Gene Editing/methods
• RNA, Viral/genetics
• RNA, Viral/metabolism

• Gene construct
• Plant resistance
• Plant viruses
• RNAi
• Resistance breeding
• Silencing elements
• Toolbox
• Virus resistance

• catalytic dna
• viral infection
• synthetic biology
• antisense oligonucleotide therapy
• nucleic acids

• Humans
• SARS-CoV-2/genetics
• RNA, Viral/genetics
• Pandemics
• COVID-19
• Antiviral Agents/pharmacology
• Antiviral Agents/therapeutic use

• Wellcome Trust
• 21543/Z/19/Z Wellcome Trust
• MR/T032413/1 Medical Research Council
• RCUK | Medical Research Council (MRC)
• NHS Blood and Transplant
• CUH | Addenbrooke’s Charitable Trust, Cambridge University Hospitals (Addenbrooke’s Charitable Trust, Cambridge University Hospitals NHS Foundation Trust)
• Wellcome Trust (Wellcome)

• Plum pox virus
• PPV
• RNA silencing
• amiRNA
• potyviruses
• virus evolution
• virus resistance

• Antiviral Agents
• Genomics
• MicroRNAs/genetics
• Plant Diseases/genetics
• Plants, Genetically Modified/genetics
• Plum Pox Virus/genetics
• RNA Interference
• RNA, Viral/genetics
• Tobacco/genetics

• Ministerio de Economía, Industria y Competitividad, Gobierno de España
• Ministerio de Ciencia e Innovación

• Madin–Darby canine kidney
• Seasonal influenza viruses
• enhancing the yield
• microRNA

• 3D structure
• RNA secondary structure
• RNA structurome
• SARS-CoV-2
• decoding
• function
• genome-wide
• high-throughput techniques

• Animals
• COVID-19
• Nucleic Acid Conformation
• RNA/chemistry
• RNA/genetics
• RNA, Viral/genetics
• SARS-CoV-2/genetics
• Sequence Analysis, RNA

• BBS/E/J/000PR9788 Biotechnology and Biological Sciences Research Council

Genome-wide association studies (GWAS) aim to correlate phenotypic changes with genotypic variation. Upon transcription, single nucleotide variants (SNVs) may alter mRNA structure, with potential impacts on transcript stability, macromolecular interactions, and translation. However, plant genomes have not been assessed for the presence of these structure-altering polymorphisms or “riboSNitches.”

We experimentally demonstrate the presence of riboSNitches in transcripts of two Arabidopsis genes, ZINC RIBBON 3 (ZR3) and COTTON GOLGI-RELATED 3 (CGR3), which are associated with continentality and temperature variation in the natural environment. These riboSNitches are also associated with differences in the abundance of their respective transcripts, implying a role in regulating the gene's expression in adaptation to local climate conditions. We then computationally predict riboSNitches transcriptome-wide in mRNAs of 879 naturally inbred Arabidopsis accessions. We characterize correlations between SNPs/riboSNitches in these accessions and 434 climate descriptors of their local environments, suggesting a role of these variants in local adaptation. We integrate this information in CLIMtools V2.0 and provide a new web resource, T-CLIM, that reveals associations between transcript abundance variation and local environmental variation.

We functionally validate two plant riboSNitches and, for the first time, demonstrate riboSNitch conditionality dependent on temperature, coining the term “conditional riboSNitch.” We provide the first pan-genome-wide prediction of riboSNitches in plants. We expand our previous CLIMtools web resource with riboSNitch information and with 1868 additional Arabidopsis genomes and 269 additional climate conditions, which will greatly facilitate in silico studies of natural genetic variation, its phenotypic consequences, and its role in local adaptation.

The online version contains supplementary material available at 10.1186/s13059-022-02656-4.

• Arabidopsis thaliana
• CLIMtools
• Genome-wide association study (GWAS)
• Single nucleotide variant (SNV)
• Structurome
• Transcriptome-wide association study (TWAS)
• riboSNitch

Avian leukosis virus (ALV) causes tumor diseases in poultry and is circulating all over the world, leading to significant economic losses. In addition, mixed infection of ALV with other viruses is very common and is often reported to contaminate live vaccines. At present, there is no effective method to suppress the replication of ALV in vitro, so it is very difficult to remove it in mixed infection. As a retrovirus, the replication of ALV can be limited by reverse transcriptase (RT) inhibitors like zidovudine (AZT), but it also causes nontargeted cytotoxicity. To find the optimal solution in cytotoxicity and inhibition efficiency in vitro culture system, we firstly designed a combination therapy of AZT and short hairpin RNA (shRNA) targeting ALV and then verified its efficiency by multiple biological methods. Results showed that shRNA can effectively inhibit the expression of RT and then limit the replication of ALV. The combination of AZT and shRNA can significantly improve the antiviral efficiency in viral replication, shedding, and provirus assembly under the condition of low cytotoxicity. Overall, in this study, the combination therapy of AZT and shRNA targeting ALV showed excellent antiviral performance against ALV in vitro culture system. This method can be applied to multiple scenarios, such as the removal of ALV in mixed infection or the purification of contaminated vaccine strains.

• RNAi
• avian leukosis virus
• reverse transcriptase
• shRNA
• zidovudine

• Biosensor
• CRISPR/Cas
• Diseases management
• Gene editing
• Infectious diseases
• Nanomedicine
• Personalized healthcare

• Antivirals
• FMDV
• RNA interference
• Target accessibility

Viral infections lead to the death of more than a million people each year around the world, both directly and indirectly. Viruses interfere with many cell functions, particularly critical pathways for cell death, by affecting various intracellular mediators. MicroRNAs (miRNAs) are a major example of these mediators because they are involved in many (if not most) cellular mechanisms. Virus-regulated miRNAs have been implicated in three cell death pathways, namely, apoptosis, autophagy, and anoikis. Several molecules (e.g., BECN1 and B cell lymphoma 2 [BCL2] family members) are involved in both apoptosis and autophagy, while activation of anoikis leads to cell death similar to apoptosis. These mechanistic similarities suggest that common regulators, including some miRNAs (e.g., miR-21 and miR-192), are involved in different cell death pathways. Because the balance between cell proliferation and cell death is pivotal to the homeostasis of the human body, miRNAs that regulate cell death pathways have drawn much attention from researchers. miR-21 is regulated by several viruses and can affect both apoptosis and anoikis via modulating various targets, such as PDCD4, PTEN, interleukin (IL)-12, Maspin, and Fas-L. miR-34 can be downregulated by viral infection and has different effects on apoptosis, depending on the type of virus and/or host cell. The present review summarizes the existing knowledge on virus-regulated miRNAs involved in the modulation of cell death pathways. Understanding the mechanisms for virus-mediated regulation of cell death pathways could provide valuable information to improve the diagnosis and treatment of many viral diseases.

• anoikis
• apoptosis
• autophagy
• cell death
• microRNAs
• viruses

• Ago2
• Dicer
• Dicer-independent shRNA
• HDV ribozyme
• HIV
• RNAi
• miR-451

• Argonaute Proteins/genetics
• Argonaute Proteins/metabolism
• Blotting, Northern
• Cloning, Molecular/methods
• Enzyme Assays/methods
• Gene Silencing
• Genetic Vectors
• HEK293 Cells
• HIV Infections/genetics
• HIV-1/genetics
• HIV-1/metabolism
• Hepatitis Delta Virus/genetics
• Humans
• Inverted Repeat Sequences/genetics
• Luciferases/metabolism
• RNA Interference
• RNA, Catalytic/genetics
• RNA, Catalytic/metabolism
• RNA, Small Interfering/genetics
• RNA, Small Interfering/metabolism
• T-Lymphocytes/virology
• Transfection/methods

• HIV-1 latency
• HIV-1 reservoir
• antiretroviral therapy
• block-and-lock
• functional cure

• Anti-HIV Agents/pharmacology
• Anti-HIV Agents/therapeutic use
• Antiretroviral Therapy, Highly Active
• CD4-Positive T-Lymphocytes/drug effects
• CD4-Positive T-Lymphocytes/immunology
• CD4-Positive T-Lymphocytes/metabolism
• Combined Modality Therapy/methods
• Gene Expression Regulation, Viral
• Genetic Therapy/methods
• HIV Infections/drug therapy
• HIV Infections/immunology
• HIV Infections/virology
• HIV-1/drug effects
• HIV-1/physiology
• Host-Pathogen Interactions
• Humans
• Proviruses/genetics
• Transcription, Genetic
• Treatment Outcome
• Virus Activation/drug effects
• Virus Latency/drug effects
• Virus Replication/drug effects

• R01 AI097012 NIAID NIH HHS
• R01 AI118432 NIAID NIH HHS
• R33 AI116226 NIAID NIH HHS
• R61 AI140439 NIAID NIH HHS

• Cationic liposomes
• DODAB
• DODAC
• PEGylation
• monoolein
• siRNA delivery.

Human immunodeficiency virus (HIV-1) indefinitely persists, despite effective antiretroviral therapy (ART), within a small pool of latently infected cells. These cells often display markers of immunologic memory and harbor both replication-competent and -incompetent proviruses at approximately a 1:100 ratio. Although complete HIV eradication is a highly desirable goal, this likely represents a bridge too far for our current and foreseeable technologies. A more tractable goal involves engineering a sustained viral remission in the absence of ART––a “functional cure.” In this setting, HIV remains detectable during remission, but the size of the reservoir is small and the residual virus is effectively controlled by an engineered immune response or other intervention. Biological precedence for such an approach is found in the post-treatment controllers (PTCs), a rare group of HIV-infected individuals who, following ART withdrawal, do not experience viral rebound. PTCs are characterized by a small reservoir, greatly reduced inflammation, and the presence of a poorly understood immune response that limits viral rebound. Our goal is to devise a safe and effective means for replicating durable post-treatment control on a global scale. This requires devising methods to reduce the size of the reservoir and to control replication of this residual virus. In the following sections, we will review many of the approaches and tools that likely will be important for implementing such a “reduce and control” strategy and for achieving a PTC-like sustained HIV remission in the absence of ART.

• HIV
• block and lock
• cure
• genome editing
• reduce and control
• shock and kill

• Animals
• Anti-Retroviral Agents/therapeutic use
• Disease Reservoirs/virology
• HIV Infections/prevention & control
• HIV Infections/therapy
• HIV Infections/virology
• HIV-1/physiology
• Humans
• Immunologic Memory
• Latent Infection
• Macaca mulatta
• Remission Induction
• Simian Acquired Immunodeficiency Syndrome/virology
• Simian Immunodeficiency Virus
• Viral Load
• Virus Activation
• Virus Latency
• Virus Replication

• P01 AI131374 NIAID NIH HHS
• P30 AI027763 NIH HHS
• R01 DA049525 NIDA NIH HHS
• R01 DA044605 NIDA NIH HHS
• P30 AI027763 NIAID NIH HHS
• P01 10018714 National Institute of Allergy and Infectious Diseases

• Arenavirus
• CRISPR
• Cas13
• Cas13-based detection
• RNA interference
• RNA viruses
• antiviral
• crRNA design
• influenza virus
• multiplexing

• A549 Cells
• Animals
• CRISPR-Associated Proteins/genetics
• CRISPR-Associated Proteins/metabolism
• CRISPR-Cas Systems
• Chlorocebus aethiops
• Clustered Regularly Interspaced Short Palindromic Repeats
• Dogs
• Escherichia coli/enzymology
• Escherichia coli/genetics
• Gene Targeting/methods
• HEK293 Cells
• Humans
• Madin Darby Canine Kidney Cells
• RNA Stability
• RNA Viruses/enzymology
• RNA Viruses/genetics
• RNA, Viral/genetics
• RNA, Viral/metabolism
• Vero Cells

• R01 MH110049 NIMH NIH HHS
• DP1 HL141201 NHLBI NIH HHS
• R01 HG009761 NHGRI NIH HHS
• U19 AI110818 NIAID NIH HHS
• RM1 HG006193 NHGRI NIH HHS

• Zika
• clustered regularly interspaced short palindromic repeats
• clustered regularly interspaced short palindromic repeats–associated protein 9
• dengue
• gene drive
• homing
• malaria

RNA interference (RNAi) can be triggered by synthetic small interfering RNAs (siRNAs) or transgene-expressed short hairpin RNAs (shRNAs). Recent evidence indicates that shRNA molecules, with a relatively short stem and small loop, are processed by Argonaute 2 protein (Ago2). We named these molecules AgoshRNA as Ago2 is involved in both the processing and the subsequent mRNA-silencing reaction. This alternative processing route yields only a single guide strand, which thus avoids potential off-target effects induced by the passenger strand of a regular shRNA. We recently described that the introduction of a 5ʹ-terminal purine (A or G) and a mismatch at the bottom of the hairpin enhances the AgoshRNA activity. The critical 5ʹ-terminal nucleotide (nt) represents the +1 position of the transcriptional promoter, which influences the transcriptional efficiency and initiation accuracy as demonstrated for the H1 RNA polymerase (Pol) III promoter. These findings highlight the necessity of considering Pol III requirements in the design of optimized AgoshRNA cassettes. In this study, we report the design and expression of potent AgoshRNAs by two other popular Pol III promoters: U6 and 7SK, which were recently reported to have a distinct transcription profile compared to the H1 promoter. We propose general rules for the design and expression of potent AgoshRNA molecules using Pol III cassettes, which should augment the application of novel AgoshRNA reagents for basic research and therapeutic purposes.

• 5’-terminal nucleotide
• AgoshRNA
• RNA polymerase III promoter
• shRNA
• transcription

Short hairpin RNAs (shRNAs) can induce gene silencing via the RNA interference (RNAi) mechanism. We designed an alternative shRNA molecule with a relatively short base-paired stem that bypasses Dicer and instead is processed by the Argonaute 2 (Ago2) protein into a single guide RNA strand that effectively induces RNAi. We called these molecules AgoshRNAs. Active anti-HIV AgoshRNAs were developed, but their RNAi activity was generally reduced compared with the matching shRNAs. In an attempt to further optimize the AgoshRNA design, we inserted several self-cleaving ribozymes at the 3′ terminus of the transcribed AgoshRNA and evaluated the impact on AgoshRNA processing and activity. The hepatitis delta virus (HDV) ribozyme is efficiently removed from the transcribed AgoshRNAs and generates a uniform 3′ overhang, which translates into the enhanced antiviral activity of these molecules.

• Ago2
• Dicer
• Dicer-independent shRNA
• HIV
• RNAi
• miR-451
• ribozyme

• HIV-1
• biomarker
• disease progression
• immune system
• microRNA

• host–virus interaction
• nicotiana tabacum, potyvirus
• response to infection
• systems biology
• tev
• transcriptomics
• virus evolution
• viral fitness

• Gene Expression
• Genetic Fitness
• Host-Pathogen Interactions
• Microarray Analysis
• Models, Biological
• Plant Diseases
• Potyvirus/genetics
• Real-Time Polymerase Chain Reaction
• Nicotiana/metabolism
• Nicotiana/virology
• Transcriptome

• Alternative pre-mRNA splicing
• Exon definition
• HIV-1 dependency factors (HDF), restriction factors, novel antiretroviral therapy
• HIV-1 infection
• Splicing regulatory elements

HIV-1 has already claimed over 35 million human lives globally. No curative treatments are currently available, and the only treatment option for over 36 million people currently living with HIV/AIDS are antiretroviral drugs that disrupt the function of virus-encoded proteins. However, such virus-targeted therapeutic strategies are constrained by the ability of the virus to develop drug-resistance. Despite major advances in HIV/AIDS research over the years, substantial knowledge gaps exist in many aspects of HIV-1 replication, especially its interaction with the host. Hence, understanding the mechanistic details of virus–host interactions may lead to novel therapeutic strategies for the prevention and/or management of HIV/AIDS. Notably, unprecedented progress in deciphering host gene silencing processes mediated by several classes of cellular small non-coding RNAs (sncRNA) presents a promising and timely opportunity for developing non-traditional antiviral therapeutic strategies. Cellular microRNAs (miRNA) belong to one such important class of sncRNAs that regulate protein synthesis. Evidence is mounting that cellular miRNAs play important roles in viral replication, either usurped by the virus to promote its replication or employed by the host to control viral infection by directly targeting the viral genome or by targeting cellular proteins required for productive virus replication. In this review, we summarize the findings to date on the role of miRNAs in HIV-1 biology.

• HIV
• Post-transcriptional regulation
• miRNA

• CRISPR-Cas
• Combination therapy
• HIV-1
• Latency
• NHEJ
• Virus escape
• dCas9

• Bacterial Proteins/genetics
• Bacterial Proteins/metabolism
• CRISPR-Associated Protein 9
• CRISPR-Cas Systems
• Clustered Regularly Interspaced Short Palindromic Repeats
• DNA Cleavage
• DNA, Viral/genetics
• DNA, Viral/metabolism
• Endonucleases/genetics
• Endonucleases/metabolism
• HIV Infections/therapy
• HIV Infections/virology
• HIV Long Terminal Repeat
• HIV-1/genetics
• HIV-1/growth & development
• HIV-1/metabolism
• Humans
• Molecular Targeted Therapy/methods
• RNA, Guide, CRISPR-Cas Systems/genetics
• RNA, Guide, CRISPR-Cas Systems/metabolism
• Tumor Necrosis Factor-alpha/pharmacology
• Virus Latency/drug effects
• Virus Replication/drug effects

• gene therapy
• miRNA cassettes
• vector tropism
• viral vectors

Although a limited percentage of the genome produces proteins, approximately 90% is transcribed, indicating important roles for noncoding RNA (ncRNA). It is now known that these ncRNAs have a multitude of cellular functions ranging from the regulation of gene expression to roles as structural elements in ribonucleoprotein complexes. ncRNA is also represented at nearly every step of viral life cycles. This chapter will focus on ncRNAs of both host and viral origin and their roles in retroviral life cycles. Cellular ncRNA represents a significant portion of material packaged into retroviral virions and includes transfer RNAs, 7SL RNA, U RNA, and vault RNA. Initially thought to be random packaging events, these host RNAs are now proposed to contribute to viral assembly and infectivity. Within the cell, long ncRNA and endogenous retroviruses have been found to regulate aspects of the retroviral life cycle in diverse ways. Additionally, the HIV-1 transactivating response element RNA is thought to impact viral infection beyond the well-characterized role as a transcription activator. RNA interference, thought to be an early version of the innate immune response to viral infection, can still be observed in plants and invertebrates today. The ability of retroviral infection to manipulate the host RNAi pathway is described here. Finally, RNA-based therapies, including gene editing approaches, are being explored as antiretroviral treatments and are discussed.

Purpose: Human telomerase reverse transcriptase (hTERT) plays a crucial role in tumorigenesis and progression of cancers. Gene silencing of hTERT by short interfering RNA (siRNA) is considered as a promising strategy for cancer gene therapy. Various algorithms have been devised for designing a high efficient siRNA which is a significant issue in the clinical usage. Thereby, in the present study, the relation of siRNA designing criteria and the gene silencing efficiency was evaluated.

Methods: The siRNA sequences were designed and characterized by using on line soft wares. Cationic co-polymer (polyethylene glycol-g-polyethylene imine (PEG-g-PEI)) was used for the construction of polyelectrolyte complexes (PECs) containing siRNAs. The cellular uptake of the PECs was evaluated. The gene silencing efficiency of different siRNA sequences was investigated and the effect of observing the rational designing on the functionality of siRNAs was assessed.

Results: The size of PEG-g-PEI siRNA with N/P (Nitrogen/Phosphate) ratio of 2.5 was 114 ± 0.645 nm. The transfection efficiency of PECs was desirable (95.5% ± 2.4%.). The results of Real-Time PCR showed that main sequence (MS) reduced the hTERT expression up to 90% and control positive sequence (CPS) up to 63%. These findings demonstrated that the accessibility to the target site has priority than the other criteria such as sequence preferences and thermodynamic features.

Conclusion: siRNA opens a hopeful window in cancer therapy which provides a convenient and tolerable therapeutic approach. Thereby, using the set of criteria and rational algorithms in the designing of siRNA remarkably affect the gene silencing efficiency.

• Gene silencing
• RNA Interference
• hTERT

• HIV-1
• RNA interference
• cell transplant
• functional cure
• micro RNA
• small/short hairpin RNA
• small/short interfering RNA

• Anti-HIV Agents/therapeutic use
• Gene Expression Regulation, Viral
• HIV Infections/therapy
• HIV Infections/virology
• HIV-1/drug effects
• HIV-1/genetics
• Humans
• RNA Interference/physiology
• RNA, Small Interfering/therapeutic use
• RNAi Therapeutics
• Virus Replication

• PPP-133377 CIHR
• HBF-143163 CIHR
• PJT-148704 CIHR

Short hairpin RNAs (shRNAs) are widely used for gene silencing by the RNA interference (RNAi) mechanism. The shRNA precursor is processed by the Dicer enzyme into active small interfering RNAs (siRNAs) that subsequently target a complementary mRNA for cleavage by the Argonaute 2 (Ago2) complex. Recent evidence indicates that shRNAs with a relatively short basepaired stem bypass Dicer and are instead processed by Ago2. We termed these molecules AgoshRNAs as both processing and silencing steps are mediated by Ago2 and proposed rules for the design of effective AgoshRNA molecules. Active and non-cytotoxic AgoshRNAs against HIV-1 RNA were generated, but their silencing activity was generally reduced compared with the matching shRNAs. Thus, further optimization of the AgoshRNA design is needed. In this study, we evaluated the importance of the single-stranded loop, in particular its size and nucleotide sequence, in AgoshRNA-mediated silencing. We document that the pyrimidine/purine content is important for AgoshRNA-mediated silencing activity.

• Ago2
• Dicer
• Dicer-independent shRNA
• HIV-1
• RNAi
• miR-451
• shRNA design

• HIV
• RNAi
• escaping mutation
• gene therapy
• long-term inhibition

Recent evidence indicates that shRNAs with a relatively short basepaired stem do not require Dicer processing, but instead are processed by the Argonaute 2 protein (Ago2). We named these molecules AgoshRNAs as both their processing and silencing function are mediated by Ago2. This alternative processing yields only a single RNA guide strand, which can avoid off-target effects induced by the passenger strand of regular shRNAs. It is important to understand this alternative processing route in mechanistic detail such that one can design improved RNA reagents. We verified that AgoshRNAs trigger site-specific cleavage of a complementary mRNA. Second, we document the importance of the identity of the 5΄-terminal nucleotide and its basepairing status for AgoshRNA activity. AgoshRNA activity is significantly reduced or even abrogated with C or U at the 5΄-terminal and is enhanced by introduction of a bottom mismatch and 5΄-terminal nucleotide A or G. The 5΄-terminal RNA nucleotide also represents the +1 position of the transcriptional promoter in the DNA, thus further complicating the analysis. Indeed, we report that +1 modification affects the transcriptional efficiency and accuracy of start site selection, with A or G as optimal nucleotide. These combined results allow us to propose general rules for the design and expression of potent AgoshRNA molecules.

MicroRNAs (miRNAs) are powerful regulators of gene expression and fine-tuning genes in all tissues. Cellular miRNAs can control 100s of biologic processes (e.g., morphogenesis of embryonic structures, differentiation of tissue-specific cells, and metabolic control in specific cell types) and have been involved in the regulation of nearly all cellular pathways. Inherently to their involvement in different physiologic processes, miRNAs deregulation has been associated with several diseases. Moreover, several viruses have been described as either, avoid and block cellular miRNAs or synthesize their own miRNA to facilitate infection and pathogenesis. Adenoviruses genome encodes two non-coding RNAs, known as viral-associated (VA) RNA_I and VA RNA_II, which seem to play an important role either by blocking important proteins from miRNA pathway, such as Exportin-5 and Dicer, or by targeting relevant cellular factors. Drastic changes in cellular miRNA expression profile are also noticeable and several cellular functions are affected by these changes. This review focuses on the mechanisms underlying the biogenesis and molecular interactions of miRNAs providing basic concepts of their functions as well as in the interplay between miRNAs and human adenoviruses.

• adenoviruses
• gene regulation
• mRNA translation
• microRNA
• oncogenesis
• pathogenesis

• CRISPR-Cas Systems/genetics
• Gene Editing/methods
• Genetic Engineering/methods
• Genetic Therapy/methods
• Genome, Viral/genetics
• HIV Infections/therapy
• HIV-1/genetics
• Humans
• Natronobacterium/enzymology
• RNA, Small Interfering/genetics
• RNA, Small Interfering/therapeutic use
• Transcription Activator-Like Effector Nucleases/therapeutic use
• Zinc Finger Nucleases/therapeutic use

• HIV-1
• N6-methyladenosine
• Posttranscriptional gene regulation
• RNA modification
• mRNA function
• mRNA stability

Analysis of the incorporation of cellular microRNAs (miRNAs) into highly purified HIV-1 virions revealed that this largely, but not entirely, mirrored the level of miRNA expression in the producer CD4⁺ T cells. Specifically, of the 58 cellular miRNAs detected at significant levels in the producer cells, only 5 were found in virions at a level 2- to 4-fold higher than that predicted on the basis of random cytoplasmic sampling. Of note, these included two miRNAs, miR-155 and miR-92a, that were reported previously to at least weakly bind HIV-1 transcripts. To test whether miRNA binding to the HIV-1 genome can induce virion incorporation, artificial miRNA target sites were introduced into the viral genome and a 10- to 40-fold increase in the packaging of the cognate miRNAs into virions was then observed, leading to the recruitment of up to 1.6 miRNA copies per virion. Importantly, this high level of incorporation significantly inhibited HIV-1 virion infectivity. These results suggest that target sites for cellular miRNAs can inhibit RNA virus replication at two distinct steps, i.e., during infection and during viral gene expression, thus explaining why a range of different RNA viruses appear to have evolved to avoid cellular miRNA binding to their genome.

The genomes of RNA viruses have the potential to interact with cellular miRNAs, which could lead to their incorporation into virions, with unknown effects on virion function. Here, it is demonstrated that wild-type HIV-1 virions essentially randomly incorporate low levels of the miRNAs expressed by infected cells. However, the specific incorporation of high levels of individual cellular miRNAs can be induced by insertion of cognate target sites into the viral genome. Of note, this results in a modest but significant inhibition of virion infectivity. These data imply that cellular miRNAs have the potential to inhibit viral replication by interfering with not only viral mRNA function but also virion infectivity.

Because of a high mutation rate, HIV exists as a viral swarm of many sequence variants evolving under various selective pressures from the human immune system. Although the Nef gene codes for the most immunogenic of HIV accessory proteins, which alone makes it of great interest to HIV research, it also encodes an RNA structure, whose contribution to HIV virulence has been largely unexplored. Nef RNA helps HIV escape RNA interference (RNAi) through nucleotide changes and alternative folding. This study examines Historic and Modern Datasets of patient HIV-1 Nef sequences during the evolution of the North American epidemic for local changes in RNA plasticity. By definition, RNA plasticity refers to an RNA molecule’s ability to take alternative folds (i.e., alternative conformations). Our most important finding is that an evolutionarily conserved region of the HIV-1 Nef gene, which we denote by R2, recently underwent a statistically significant increase in its RNA plasticity. Thus, our results indicate that Modern Nef R2 typically accommodates an alternative fold more readily than Historic Nef R2. Moreover, the increase in RNA plasticity resides mostly in synonymous nucleotide changes, which cannot be a response to selective pressures on the Nef protein. R2 may therefore be of interest in the development of antiviral RNAi therapies.

• Intramural Research Program of the NIH, National Library of Medicine

HIV-1 can persist in a latent form in resting memory CD4+ cells and macrophages carrying an integrated copy of the HIV genome. Because of the presence of these stable reservoir cells, eradication by antiretroviral therapy is unlikely and in order to achieve eradication, alternative treatment options are required. Stem cell transplantation has been considered previously to effect the clinical course of HIV-infection but in practice eradication or virus control was not achievable. However, modifications of stem cell transplantation using natural or artificial resistant cell sources, combination with new techniques of gene editing or generating cytotoxic anti HIV effector cells have stimulated this field of HIV cell therapy substantially. Here, we look back on 30 years of stem cell therapy in HIV patients and discuss most recent developments in this direction.

• Adoptive cell transfer
• Cell therapy
• Gene therapy
• HIV-1
• Stem cell transplantation