Antisense Protein Research Articles

Sepsis Biomarkers: Advancements and Clinical Applications-A Narrative Review.

Sepsis is now defined as a life-threatening syndrome of organ dysfunction triggered by a dysregulated host response to infection, posing significant challenges in critical care. The main objective of this review is to evaluate the potential of emerging biomarkers for early diagnosis and accurate prognosis in sepsis management, which are pivotal for enhancing patient outcomes. Despite advances in supportive care, traditional biomarkers like C-reactive protein and procalcitonin have limitations, and recent studies have identified novel biomarkers with increased sensitivity and specificity, including circular RNAs, HOXA distal transcript antisense RNA, microRNA-486-5p, protein C, triiodothyronine, and prokineticin 2. These emerging biomarkers hold promising potential for the early detection and prognostication of sepsis. They play a crucial role not only in diagnosis but also in guiding antibiotic therapy and evaluating treatment effectiveness. The introduction of point-of-care testing technologies has brought about a paradigm shift in biomarker application, enabling swift and real-time patient evaluation. Despite these advancements, challenges persist, notably concerning biomarker variability and the lack of standardized thresholds. This review summarizes the latest advancements in sepsis biomarker research, spotlighting the progress and clinical implications. It emphasizes the significance of multi-biomarker strategies and the feasibility of personalized medicine in sepsis management. Further verification of biomarkers on a large scale and their integration into clinical practice are advocated to maximize their efficacy in future sepsis treatment.

Patterns of Diversity and Humoral Immunogenicity for HIV-1 Antisense Protein (ASP).

HIV-1 has an antisense gene overlapping env that encodes the ASP protein. ASP functions are still unknown, but it has been associated with gp120 in the viral envelope and membrane of infected cells, making it a potential target for immune response. Despite this, immune response patterns against ASP are poorly described and can be influenced by the high genetic variability of the env gene. To explore this, we analyzed 100k HIV-1 ASP sequences from the Los Alamos HIV sequence database using phylogenetic, Shannon entropy (Hs), and logo tools to study ASP variability in worldwide and Brazilian sequences from the most prevalent HIV-1 subtypes in Brazil (B, C, and F1). Data obtained in silico guided the design and synthesis of 15-mer overlapping peptides through spot synthesis on cellulose membranes. Peptide arrays were screened to assess IgG and IgM responses in pooled plasma samples from HIV controllers and individuals with acute or recent HIV infection. Excluding regions with low alignment accuracy, several sites with higher variability (Hs > 1.5) were identified among the datasets (25 for worldwide sequences, 20 for Brazilian sequences). Among sites with Hs < 1.5, sequence logos allowed the identification of 23 other sites with subtype-specific signatures. Altogether, amino acid variations with frequencies > 20% in the 48 variable sites identified were included in 92 peptides, divided into 15 sets, representing near full-length ASP. During the immune screening, the strongest responses were observed in three sets, one in the middle and one at the C-terminus of the protein. While some sets presented variations potentially associated with epitope displacement between IgG and IgM targets and subtype-specific signatures appeared to impact the level of response for some peptides, signals of cross-reactivity were observed for some sets despite the presence of B/C/F1 signatures. Our data provides a map of ASP regions preferentially targeted by IgG and IgM responses. Despite B/C/F1 subtype signatures in ASP, the amino acid variation in some areas preferentially targeted by IgM and IgG did not negatively impact the response against regions with higher immunogenicity.

Vulnerability to APOBEC3G linked to the pathogenicity of deltaretroviruses

Human retroviruses are derived from simian ones through cross-species transmission. These retroviruses are associated with little pathogenicity in their natural hosts, but in humans, HIV causes AIDS, and human T-cell leukemia virus type 1 (HTLV-1) induces adult T-cell leukemia-lymphoma (ATL). We analyzed the proviral sequences of HTLV-1, HTLV-2, and simian T-cell leukemia virus type 1 (STLV-1) from Japanese macaques (Macaca fuscata) and found that APOBEC3G (A3G) frequently generates G-to-A mutations in the HTLV-1 provirus, whereas such mutations are rare in the HTLV-2 and STLV-1 proviruses. Therefore, we investigated the mechanism of how HTLV-2 is resistant to human A3G (hA3G). HTLV-1, HTLV-2, and STLV-1 encode the so-called antisense proteins, HTLV-1 bZIP factor (HBZ), Antisense protein of HTLV-2 (APH-2), and STLV-1 bZIP factor (SBZ), respectively. APH-2 efficiently inhibits the deaminase activity of both hA3G and simian A3G (sA3G). HBZ and SBZ strongly suppress sA3G activity but only weakly inhibit hA3G, suggesting that HTLV-1 is incompletely adapted to humans. Unexpectedly, hA3G augments the activation of the transforming growth factor (TGF)-β/Smad pathway by HBZ, and this activation is associated with ATL cell proliferation by up-regulating BATF3/IRF4 and MYC. In contrast, the combination of APH-2 and hA3G, or the combination of SBZ and sA3G, does not enhance the TGF-β/Smad pathway. Thus, HTLV-1 is vulnerable to hA3G but utilizes it to promote the proliferation of infected cells via the activation of the TGF-β/Smad pathway. Antisense factors in each virus, differently adapted to control host cellular functions through A3G, seem to dictate the pathogenesis.

FGF2 gene's antisense protein, NUDT6, plays a depressogenic role by promoting inflammation and suppressing neurogenesis without altering FGF2 signalling.

Fibroblast growth factor-2 (FGF2) is involved in the regulation of affective behaviour and shows antidepressant effects through the Akt and extracellular signal regulated kinase (ERK) 1/2 pathways. Nudix hydrolase 6 (NUDT6) protein is encoded from FGF2 gene's antisense strand and its role in the regulation of affective behaviour is unknown. Here, we overexpressed NUDT6 in the hippocampus and investigated its behavioural effects and the underlying molecular mechanisms affecting the behaviour. We showed that increasing hippocampal NUDT6 results in depression-like behaviour in rats without changing FGF2 levels or activating its downstream effectors, Akt and ERK1/2. Instead, NUDT6 acted by inducing inflammatory signalling, specifically by increasing S100 calcium binding protein A9 (S100A9) levels, activating nuclear factor-kappa B-p65 (NF-κB-p65), and elevating microglia numbers along with a reduction in neurogenesis. Our results suggest that NUDT6 could play a role in major depression by inducing a proinflammatory state. This is the first report of an antisense protein acting through a different mechanism of action than regulationof its sense protein. The opposite effects of NUDT6 and FGF2 on depression-like behaviour may serve as a mechanism to fine-tune affective behaviour. Our findings open up new venues for studying the differential regulation and functional interactions of sense and antisense proteins in neural function and behaviour, as well as in neuropsychiatric disorders. KEY POINTS: Hippocampal overexpression of nudix hydrolase 6 (NUDT6), the antisense protein of fibroblast growth factor-2 (FGF2), increases depression-like behaviour in rats. Hippocampal NUDT6 overexpression triggers a neuroinflammatory cascade by increasing S100 calcium binding proteinA9 (S100A9) expression and nuclear NF-κB-p65 translocation in neurons, in addition to microglial recruitment and activation. Hippocampal NUDT6 overexpression suppresses neurogenesis. NUDT6 exerts its actions without altering the levels or downstream signalling pathways of FGF2.

Endothelial cell-expressed long non-coding RNAs in cardiac regeneration

Abstract Endothelial cells (ECs) are the most abundant non-myocyte cell types in the heart. Upon cardiac injury, revascularization can occur through proliferation and migration of ECs from existing blood vessels in the surrounding myocardium. In animal models, revascularization is crucial to stimulate cardiomyocyte survival and regeneration. In the human heart, establishing blood flow could improve outcomes of tissue engineering or cell based therapy to regenerate the heart after injury. In addition to establishing blood flow, EC also secrete factors in a paracrine manner which can act on neighbouring cardiomyocytes. We investigate a role for non-coding RNA in the regulation of endothelial cell function and their potential role in neovascularization and cardiac regeneration. A large portion of the genome is transcribed into non-coding RNA, which does not encode protein. Many long non-coding RNAs (lncRNAs) have been shown to be involved in important regulatory processes such as genomic imprinting and chromatin modification. To identify lncRNAs which could be involved in cardiac regeneration, we isolated ECs from neonatal mice at postnatal day 1 and 5.5. The potential for cardiac regeneration was observed in mice at postnatal day 1, but this capacity is lost by day 5.5. Differential expression of lncRNAs in ECs was assessed by RNA sequencing. Three potential target lncRNAs, which also have a human homologue, were selected. NR2F2-AS1 and HAND2-AS1 were upregulated at postnatal day 1, whereas FOXD2-AS1 was upregulated at day 5.5. These genes were selected for further validation in human cardiac microvascular endothelial cells (CMEC). Knockdown of NR2F2-AS1 resulted in a decrease in proliferation but did not affect angiogenic sprouting. Loss of HAND2-AS1 resulted in a trend to impaired sprouting but did not affect proliferation. FOXD2-AS1 knockdown resulted in a significant decrease in angiogenic sprouting. We observed no change in the rate of migration after knockdown of any of the three genes. Knockdown of the lncRNAs did not affect expression of the antisense protein coding transcripts, therefore the effects we observe are likely mediated by the lncRNAs. In conclusion, lncRNAs NR2F2-AS1, HAND2-AS1 and FOXD2-AS1 regulate different aspects of EC function such as proliferation and sprouting. Modulation of expression of these transcripts therefore may be considered as a therapeutic tool to promote cardiac regeneration.

PCSK9 Targeting in the Management of Hypercholesterolaemia

It has long been established that elevated plasma concentrations of low-density lipoprotein cholesterol (LDL-C) are among the prominent contributors leading to the development of atherosclerotic plaques and, ultimately, cardiovascular disease. In the current era of optimal risk factor modification, proprotein convertase subtilisin/kexin type 9 (PCSK9) targeting has emerged as a potent therapeutic approach in the management of hypercholesterolaemia, addressing several substantial, unmet clinical needs. PCSK9 monoclonal antibodies, evolocumab and alirocumab, as well as inclisiran, which is a small interfering RNA that halts the transcription of PCSK9 mRNA, are being increasingly used in current clinical practice, as they induce intensive LDL-C reductions without any significant safety and tolerability concerns. Based on the success of these agents, the concept of PCSK9 targeting with novel agents with enhanced biological properties, or via different administration routes, has received considerable attention. In this regard, numerous antisense oligonucleotides, peptides, and proteins are currently under evaluation in randomised controlled trials, yielding propitious results up to date; they may enter clinical use in the coming years. Meanwhile, a PCSK9 vaccine, as well as genome editing via clustered regularly interspaced palindromic repeats/Cas9, hold great promise to eradicate LDL-C altogether as a cardiovascular risk factor. This review aims to present and discuss the current clinical and scientific evidence pertaining to the field of medications that exert their biological effect by targeting PCSK9, which are either in use in clinical practice, or are currently being evaluated in pre-clinical or clinical studies, and may prove beneficial in the near future.

Structural analysis and molecular dynamics simulation studies of HIV-1 antisense protein predict its potential role in HIV replication and pathogenesis.

The functional significance of the HIV-1 Antisense Protein (ASP) has been a paradox since its discovery. The expression of this protein in HIV-1-infected cells and its involvement in autophagy, transcriptional regulation, and viral latency have sporadically been reported in various studies. Yet, the definite role of this protein in HIV-1 infection remains unclear. Deciphering the 3D structure of HIV-1 ASP would throw light on its potential role in HIV lifecycle and host-virus interaction. Hence, using extensive molecular modeling and dynamics simulation for 200 ns, we predicted the plausible 3D-structures of ASP from two reference strains of HIV-1 namely, Indie-C1 (subtype-C) and NL4-3 (subtype-B) so as to derive its functional implication through structural domain analysis. In spite of sequence and structural differences in subtype B and C ASP, both structures appear to share common domains like the Von Willebrand Factor Domain-A (VWFA), Integrin subunit alpha-X (ITGSX), and ETV6-Transcriptional repressor, thereby reiterating the potential role of HIV-1 ASP in transcriptional repression and autophagy, as reported in earlier studies. Gromos-based cluster analysis of the centroid structures also reassured the accuracy of the prediction. This is the first study to elucidate a highly plausible structure for HIV-1 ASP which could serve as a feeder for further experimental validation studies.

Novel perspectives on antisense transcription in HIV-1, HTLV-1, and HTLV-2.

The genome of retroviruses contains two promoter elements (called long terminal repeat or LTR) at the 5' and 3' end of their genome. Although the expression of retroviral genes generally depends on the promoter located in the 5' LTR, the 3' LTR also has promoter activity responsible for producing antisense transcripts. These natural antisense transcripts (NATs) are a class of RNA molecules transcribed from the opposite strand of a protein-coding gene. NATs have been identified in many prokaryotic and eukaryotic systems, as well as in human retroviruses such as human immunodeficiency virus type 1 (HIV-1) and HTLV-1/2 (human T-cell leukemia virus type 1/2). The antisense transcripts of HIV-1, HTLV-1, and HTLV-2 have been briefly characterized over the past several years. However, a complete appreciation of the role these transcripts play in the virus lifecycle and the cellular factors which regulate their transcription is still lacking. This review provides an overview of antisense transcription in human retroviruses with a specific focus on the MEF-2 family of transcription factors, the function(s) of the antisense protein products, and the application of antisense transcription models in therapeutics against HIV-1 and HTLV-1 in the context of co-infection.

A newly identified interaction between nucleolar NPM1/B23 and the HTLV-I basic leucine zipper factor in HTLV-1 infected cells.

Human T-cell leukemia virus type 1 is the causative agent of HTLV-1-associated myelopathy/tropical spastic paraparesis and adult T-cell leukemia-lymphoma (ATL). The HTLV-1 basic leucine zipper factor (HBZ) has been associated to the cancer-inducing properties of this virus, although the exact mechanism is unknown. In this study, we identified nucleophosmin (NPM1/B23) as a new interaction partner of HBZ. We show that sHBZ and the less abundant uHBZ isoform interact with nucleolar NPM1/B23 in infected cells and HTLV-1 positive patient cells, unlike equivalent antisense proteins of related non-leukemogenic HTLV-2, -3 and-4 viruses. We further demonstrate that sHBZ association to NPM1/B23 is sensitive to RNase. Interestingly, sHBZ was shown to interact with its own RNA. Through siRNA and overexpression experiments, we further provide evidence that NPM1/B23 acts negatively on viral gene expression with potential impact on cell transformation. Our results hence provide a new insight over HBZ-binding partners in relation to cellular localization and potential function on cell proliferation and should lead to a better understanding of the link between HBZ and ATL development.

Antisense Protein: A Novel HIV-1 Gene Requiring Attention

Antisense Protein: A Novel HIV-1 Gene Requiring Attention

The Regulatory Role of Both MBNL1 and MBNL1-AS1 in Several Common Cancers.

Muscle blind-like-proteins (MBNL) are a class of tissue-specific RNA metabolism regulators that control pre-messenger RNA-splicing. Inactivation of MBNL can lead to myotonic dystrophy in adults. MBNL is mainly expressed in skeletal muscle, neuron tissue, thymus, liver, and kidney and plays an important role in the ultimate differentiation of muscle cells and neurons. MBNL1 is a member of the MBNL protein family. The inactivation of MBNL1 protein is particularly important in the development of myotonic dystrophy and can lead to cataract formation, abnormal muscle relaxation, cardiac and neurological dysfunction, etc. The induction of MBNL1 in tumors is known to significantly inhibit tumor progression and thus significantly prolong survival. MBNL1 antisense protein MBNL1-AS1 also plays an important role in tumor migration and development. This review reveals the role of MBNL1 and MBNL1-AS1 in the complex pathogenesis of many tumors, which provide a new target for the treatment of tumors. Correlated research are systematically retrieved via PubMed. In this review, the role of MBNL1 and MBNL1- AS1 were analyzed. MBNL1 is down-regulated in breast cancer, leukemia, stomach cancer, esophageal cancer, glioma, and Huntington's disease. The function of inhibiting tumor cell metastasis decreased. It is up-regulated in cervical cancer and colorectal cancer, which can promote the development of tumor cells. Antisense protein MBNL1- AS1 can inhibit tumor cell proliferation and metastasis in colorectal cancer, non-small cell lung cancer, and gastric cancer. MBNL1 is an important regulator of tumor metastasis and growth, which exhibits a promising therapeutic target and can be further explored.

LncRNA ZFAS1 regulates the proliferation, oxidative stress, fibrosis, and inflammation of high glucose-induced human mesangial cells via the miR-588/ROCK1 axis

BackgroundDiabetic nephropathy (DN) is a critical and the most common microvascular complication and its pathogenesis is still faintly understood. Thus, this study was performed to examine the long non-coding RNA ZNFX1 Antisense Gene Protein 1 (lncRNA ZFAS1) biological function and mechanism of regulation in DN.MethodHuman glomerular mesangial cells (HGMC) were induced with high glucose (HG, 25 mM) to establish HG-induced cell viability, pro-inflammation observed in DN. After, target miRNA and mRNA were predicted through Lncbase and Targetscan. Subsequently, the expression of ZFAS1, miR-588, and ROCK1 in DN clinical samples and cell-model was examined through qRT-PCR and western blot analysis. We upheld the targeted interaction between miR-588 and ZFAS1 or ROCK1 through a dual-luciferase reporter assay. The proliferation of the cell was also examined through CCK-8 assay, while the level of HG-induced oxidative stress was established by measuring reactive oxygen species (ROS) level, and also the activities of antioxidant enzymes in the cell. Lastly, the level of accumulated extracellular matrix (ECM) protein-fibronectin and collagen type IV, and inflammatory cytokines produced by the cell was analyzed through western blot analysis and ELISA.ResultsZFAS1 was significantly upregulated in the DN blood samples and HG-induced HGMC. Prediction result revealed that the ZFAS1 endogenously targets the miR-588 seed sequence while miR-588 plays a role in post-transcriptional regulation of ROCK1 mRNA. Moreover, we found that miR-588 expression was significantly downregulated in DN blood samples and negatively correlates with ZFAS1 expression. Further results show that silencing ZFAS1 had a protective effect on HG-induced proliferation, oxidative stress, fibrosis, and inflammation in HGMC while miR-588 inhibition and ROCK1 overexpression reversed this effect.ConclusionsAltogether, our data suggest that ZFAS1 regulates the proliferation, oxidative stress, fibrosis, and inflammation of high glucose-induced diabetic nephropathy through the miR-588/ROCK1 axis.

Extending the Coding Potential of Viral Genomes with Overlapping Antisense ORFs: A Case for the De Novo Creation of the Gene Encoding the Antisense Protein ASP of HIV-1.

Gene overprinting occurs when point mutations within a genomic region with an existing coding sequence create a new one in another reading frame. This process is quite frequent in viral genomes either to maximize the amount of information that they encode or in response to strong selective pressure. The most frequent scenario involves two different reading frames in the same DNA strand (sense overlap). Much less frequent are cases of overlapping genes that are encoded on opposite DNA strands (antisense overlap). One such example is the antisense ORF, asp in the minus strand of the HIV-1 genome overlapping the env gene. The asp gene is highly conserved in pandemic HIV-1 strains of group M, and it is absent in non-pandemic HIV-1 groups, HIV-2, and lentiviruses infecting non-human primates, suggesting that the ~190-amino acid protein that is expressed from this gene (ASP) may play a role in virus spread. While the function of ASP in the virus life cycle remains to be elucidated, mounting evidence from several research groups indicates that ASP is expressed in vivo. There are two alternative hypotheses that could be envisioned to explain the origin of the asp ORF. On one hand, asp may have originally been present in the ancestor of contemporary lentiviruses, and subsequently lost in all descendants except for most HIV-1 strains of group M due to selective advantage. Alternatively, the asp ORF may have originated very recently with the emergence of group M HIV-1 strains from SIVcpz. Here, we used a combination of computational and statistical approaches to study the genomic region of env in primate lentiviruses to shed light on the origin, structure, and sequence evolution of the asp ORF. The results emerging from our studies support the hypothesis of a recent de novo addition of the antisense ORF to the HIV-1 genome through a process that entailed progressive removal of existing internal stop codons from SIV strains to HIV-1 strains of group M, and fine tuning of the codon sequence in env that reduced the chances of new stop codons occurring in asp. Altogether, the study supports the notion that the HIV-1 asp gene encodes an accessory protein, providing a selective advantage to the virus.

Retroviral Antisense Transcripts and Genes: 33 Years after First Predicted, a Silent Retroviral Revolution?

Paradigm shifts throughout the history of microbiology have typically been ignored, or met with skepticism and resistance, by the scientific community. This has been especially true in the field of virology, where the discovery of a “contagium vivum fluidum”, or infectious fluid remaining after excluding bacteria by filtration, was initially ignored because it did not coincide with the established view of microorganisms. Subsequent studies on such infectious agents, eventually termed “viruses”, were met with skepticism. However, after an abundance of proof accumulated, viruses were eventually acknowledged as defined microbiological entities. Next, the proposed role of viruses in oncogenesis in animals was disputed, as was the unique mechanism of genome replication by reverse transcription of RNA by the retroviruses. This same pattern of skepticism holds true for the prediction of the existence of retroviral “antisense” transcripts and genes. From the time of their discovery, it was thought that retroviruses encoded proteins on only one strand of proviral DNA. However, in 1988, it was predicted that human immunodeficiency virus type 1 (HIV-1), and other retroviruses, express an antisense protein encoded on the DNA strand opposite that encoding the known viral proteins. Confirmation came quickly with the characterization of the antisense protein, HBZ, of the human T-cell leukemia virus type 1 (HTLV-1), and the finding that both the protein and its antisense mRNA transcript play key roles in viral replication and pathogenesis. However, acceptance of the existence, and potential importance, of a corresponding antisense transcript and protein (ASP) in HIV-1 infection and pathogenesis has lagged, despite gradually accumulating theoretical and experimental evidence. The most striking theoretical evidence is the finding that asp is highly conserved in group M viruses and correlates exclusively with subtypes, or clades, responsible for the AIDS pandemic. This review outlines the history of the major shifts in thought pertaining to the nature and characteristics of viruses, and in particular retroviruses, and details the development of the hypothesis that retroviral antisense transcripts and genes exist. We conclude that there is a need to accelerate studies on ASP, and its transcript(s), with the view that both may be important, and overlooked, targets in anti-HIV therapeutic and vaccine strategies.

MYOTONIC DYSTROPHY: EP.235 MyoScreen, a robust muscle-based assay platform validated for screening pharmacologicals against myotonic dystrophy

Development of a predictive in vitro model for deciphering the mechanisms involved in Myotonic Dystrophy type 1 (DM1) and accelerating the identification of new therapies remains a crucial unmet need. Our objective was to provide such an in vitro DM1 model capitalizing on the use of MyoScreen™, a well-established high-throughput screening platform integrating human primary skeletal myotubes that demonstrate mature sarcomeric organization and expected responses to chemical/electrical stimulation and pharmacologically relevant drugs. Primary myoblasts from three DM1 patients (blood CTG repeats from 300-1300) and one healthy donor were obtained, amplified and differentiated into aligned myotubes. Main hallmarks of the disease, such as DMPK mRNA foci colocalizing with MBNL1 protein aggregates, were present in the DM1 myotubes and quantified using high-content analysis. No direct relationship between the CTG expansion size and foci number was noted. The splicing profile of several genes involved in the Excitation-Contraction (EC) coupling machinery are altered in DM1. Thus, calcium flux upon chemical induction of EC coupling was investigated and defects in calcium transients were detected in two DM1 donors. To further characterize DM1 myotubes, Cell Profiling was applied to extract over 300 MBNL1 expression features from the generated images. We found this to be a powerful method to discriminate between DM1 and healthy myotube populations. A corresponding machine learning model was built to measure the efficacy of a (CAG)7 ASO (antisense oligonucleotide) to reverse the DM1-associated characteristics. A dose-dependent rescue by up to 60% was observed in DM1 donors. Overall, the DM1 MyoScreen platform will be useful for predictive preclinical testing to rapidly identify new therapeutic targets and select candidate therapeutics targeting skeletal muscle including ASO conjugates, compounds, siRNAs and AAVs (adenovirus associated vectors) producing antisense RNAs and proteins.

HTLV-2 Encoded Antisense Protein APH-2 Suppresses HIV-1 Replication.

Antisense protein of Human T-cell Leukemia Virus Type 2 (HTLV-2), also called APH-2, negatively regulates the HTLV-2 and helps the virus to maintain latency via scheming the transcription. Despite the remarkable occurrence of HTLV-2/HIV-1 co-infection, the role of APH-2 influencing HIV-1 replication kinetics is poorly understood and needs investigation. In this study, we investigated the plausible role of APH-2 regulating HIV-1 replication. Herein, we report that the overexpression of APH-2 not only hampered the release of HIV-1 pNL4.3 from 293T cells in a dose-dependent manner but also affected the cellular gag expression. A similar and consistent effect of APH-2 overexpression was also observed in case of HIV-1 gag expression vector HXB2 pGag-EGFP. APH-2 overexpression also inhibited the ability of HIV-1 Tat to transactivate the HIV-1 LTR-driven expression of luciferase. Furthermore, the introduction of mutations in the IXXLL motif at the N-terminal domain of APH-2 reverted the inhibitory effect on HIV-1 Tat-mediated transcription, suggesting the possible role of this motif towards the downregulation of Tat-mediated transactivation. Overall, these findings indicate that the HTLV-2 APH-2 may affect the HIV-1 replication at multiple levels by (a) inhibiting the Tat-mediated transactivation and (b) hampering the virus release by affecting the cellular gag expression.

Antisense Oligonucleotide-Induced Amyloid Precursor Protein Splicing Modulation as a Therapeutic Approach for Dutch-Type Cerebral Amyloid Angiopathy.

Dutch-type cerebral amyloid angiopathy (D-CAA) is a monogenic form of cerebral amyloid angiopathy and is inherited in an autosomal dominant manner. The disease is caused by a point mutation in exon 17 of the amyloid precursor protein (APP) gene that leads to an amino acid substitution at codon 693. The mutation is located within the amyloid beta (Aβ) domain of APP, and leads to accumulation of toxic Aβ peptide in and around the cerebral vasculature. We have designed an antisense oligonucleotide (AON) approach that results in skipping of exon 17, generating a shorter APP isoform that lacks part of the Aβ domain and the D-CAA mutation. We demonstrate efficient AON-induced skipping of exon 17 at RNA level and the occurrence of a shorter APP protein isoform in three different cell types. This resulted in a reduction of Aβ40 in neuronally differentiated, patient-derived induced pluripotent stem cells. AON-treated wild-type mice showed successful exon skipping on RNA and protein levels throughout the brain. These results illustrate APP splice modulation as a promising therapeutic approach for D-CAA.

The HIV-1 Antisense Gene ASP: The New Kid on the Block.

Viruses have developed incredibly creative ways of making a virtue out of necessity, including taking full advantage of their small genomes. Indeed, viruses often encode multiple proteins within the same genomic region by using two or more reading frames in both orientations through a process called overprinting. Complex retroviruses provide compelling examples of that. The human immunodeficiency virus type 1 (HIV-1) genome expresses sixteen proteins from nine genes that are encoded in the three positive-sense reading frames. In addition, the genome of some HIV-1 strains contains a tenth gene in one of the negative-sense reading frames. The so-called Antisense Protein (ASP) gene overlaps the HIV-1 Rev Response Element (RRE) and the envelope glycoprotein gene, and encodes a highly hydrophobic protein of ~190 amino acids. Despite being identified over thirty years ago, relatively few studies have investigated the role that ASP may play in the virus lifecycle, and its expression in vivo is still questioned. Here we review the current knowledge about ASP, and we discuss some of the many unanswered questions.

Antiviral Activity of Nanocomplexes of Antisense Oligonucleotides Targeting VP72 Protein in Vero Cells Infected by African Swine Fever Virus

Antiviral Activity of Nanocomplexes of Antisense Oligonucleotides Targeting VP72 Protein in Vero Cells Infected by African Swine Fever Virus

Antisense Transcripts and Antisense Protein: A New Perspective on Human Immunodeficiency Virus Type 1.

It was first predicted in 1988 that there may be an Open Reading Frame (ORF) on the negative strand of the Human Immunodeficiency Virus type 1 (HIV-1) genome that could encode a protein named AntiSense Protein (ASP). In spite of some controversy, reports began to emerge some years later describing the detection of HIV-1 antisense transcripts, the presence of ASP in transfected and infected cells, and the existence of an immune response targeting ASP. Recently, it was established that the asp gene is exclusively conserved within the pandemic group M of HIV-1. In this review, we summarize the latest findings on HIV-1 antisense transcripts and ASP, and we discuss their potential functions in HIV-1 infection together with the role played by antisense transcripts and ASPs in some other viruses. Finally, we suggest pathways raised by the study of antisense transcripts and ASPs that may warrant exploration in the future.