A unified classification system for HIV-1 5' long terminal repeats.

The HIV-1 provirus mainly consists of internal coding region flanked by 1 long terminal repeats (LTRs) at each terminus. The LTRs play important roles in HIV-1 reverse transcription, integration, and transcription. However, despite of the significant study advances of the internal coding regions of HIV-1 by using definite reference classification, there are no systematic and phylogenetic classifications for HIV-1 5' LTRs, which hinders our elaboration on 5' LTR and a better understanding of the viral origin, spread and therapy. Here, by analyzing all available resources of 5' LTR sequences in public databases following 4 recognized principles for the reference classification, 83 representatives and 14 consensus sequences were identified as representatives of 2 groups, 6 subtypes, 6 sub-subtypes, and 9 CRFs. To test the reliability of the supplemented classification system, the constructed references were applied to identify the 5' LTR assignment of the 22 clinical isolates in China. The results revealed that 16 out of 22 tested strains showed a consistent subtype classification with the previous LTR-independent classification system. However, 6 strains, for which recombination events within 5' LTR were demonstrated, unexpectedly showed a different subtype classification, leading a significant change of binding sites for important transcription factors including SP1, p53, and NF-κB. The binding change of these transcriptional factors would probably affect the transcriptional activity of 5' LTR. This study supplemented a unified classification system for HIV-1 5' LTRs, which will facilitate HIV-1 characterization and be helpful for both basic and clinical research fields.

Comprehensive characterization of ERV-K (HML-8) in the chimpanzee genome revealed less genomic activity than humans.

Endogenous retroviruses (ERVs) originate from ancestral germline infections caused by exogenous retroviruses. Throughout evolution, they have become fixed within the genome of the animals into which they were integrated. As ERV elements coevolve with the host, they are normally epigenetically silenced and can become upregulated in a series of physiological and pathological processes. Generally, a detailed ERV profile in the host genome is critical for understanding the evolutionary history and functional performance of the host genome. We previously characterized and cataloged all the ERV-K subtype HML-8 loci in the human genome; however, this has not been done for the chimpanzee, the nearest living relative of humans. In this study, we aimed to catalog and characterize the integration of HML-8 in the chimpanzee genome and compare it with the integration of HML-8 in the human genome. We analyzed the integration of HML-8 and found that HML-8 pervasively invaded the chimpanzee genome. A total of 76 proviral elements were characterized on 23/24 chromosomes, including detailed elements distribution, structure, phylogeny, integration time, and their potential to regulate adjacent genes. The incomplete structure of HML-8 proviral LTRs will undoubtedly affect their activity. Moreover, the results indicated that HML-8 integration occurred before the divergence between humans and chimpanzees. Furthermore, chimpanzees include more HML-8 proviral elements (76 vs. 40) and fewer solo long terminal repeats (LTR) (0 vs. 5) than humans. These results suggested that chimpanzee genome activity is less than the human genome and that humans may have a better ability to shape and screen integrated proviral elements. Our work is informative in both an evolutionary and a functional context for ERVs.

Identification and Characterization of the HERV-K (HML-8) Group of Human Endogenous Retroviruses in the Genome.

Human endogenous retroviruses (HERVs) can be vertically transmitted in a Mendelian fashion, are stably maintained in the human genome, and are estimated to constitute ∼8% of the genome. HERVs affect human physiology and pathology through their provirus-encoded protein or long terminal repeat (LTR) element effect. Characterization of the genomic distribution is an essential step to understanding the relationships between endogenous retrovirus expression and diseases. However, the poor characterization of human MMTV-like (HML)-8 prevents a detailed understanding of the regulation of the expression of this family in humans and its impact on the host genome. In light of this, the definition of an accurate and updated HERV-K HML-8 genomic map is urgently needed. In this study, we report the results of a comprehensive analysis of HERV-K HML-8 sequence presence and distribution within the human genome and hominoids, with a detailed description of the different structural and phylogenetic aspects characterizing the group. A total of 40 proviruses and 5 solo LTR elements for human were characterized, which included a detailed description of provirus structure, integration time, potentially regulated genes, transcription factor-binding sites, and primer-binding site features. Besides, 9 chimpanzee sequences, 8 gorilla sequences, and 10 orangutan sequences belonging to the HML-8 subgroup were identified. The integration time results showed that the HML-8 elements were integrated into the primate lineage around 35 and 42 million years ago (mya), during primates evolutionary speciation. Overall, the results clarified the composition of the HML-8 groups, providing an exhaustive background for subsequent functional studies.

p53 Binding Sites in Long Terminal Repeat 5Hs (LTR5Hs) of Human Endogenous Retrovirus K Family (HML-2 Subgroup) Play Important Roles in the Regulation of LTR5Hs Transcriptional Activity.

The long terminal repeats (LTRs) of human endogenous retroviruses (HERVs) are distributed throughout the human genome and provide favorable conditions to regulate the expression of their adjacent genes. HML-2 is the most biologically active subgroup of the HERV-K family, and expression of its members has been associated with many cancer types. The LTRs of HML-2 have been classified into three subgroups (LTR5A, LTR5B, and LTR5Hs) based on phylogenetic analyses. The current study aimed to explore the LTR transcriptional activity differences among the three subtypes and further explore the underlying factors. A total of 43 LTR5A elements, 62 LTR5B elements, and 194 LTR5Hs elements were selected. A phylogenetic tree showed that the LTR5Hs group was clearly separated from the LTR5A and LTR5B groups. A luciferase reporter assay indicated that LTR5Hs had the strongest promoter activity, followed by LTR5A and LTR5B. To investigate the underlying factors, LTR5Hs was divided into 4 sections, and the homologous fragments in LTR5B were replaced successively. Replacement of the third section (-263 to 0) significantly increased LTR5B activity. Subsequent mutation experiments revealed that the increased transcriptional activity was induced by the TATA box and the two p53 binding sites within the section. Further interference with TP53 significantly decreased LTR5Hs transcriptional activity. Chromatin immunoprecipitation (ChIP) and CUT&Tag experiments finally confirmed the direct binding of the p53 protein with the two LTR5Hs p53 binding sites. Overall, the two p53 binding sites in the third section (-263 to 0) of LTR5Hs were revealed to play critical roles in the difference in transcriptional activity among the three subtypes. IMPORTANCE Human endogenous retroviruses (HERVs) were integrated into the human genome in ancient times and have been coevolving with the host. Since the Human Genome Project, HERVs have attracted increasing attention. Many studies have focused on their characterization, evolution, and biological function. In particular, the expression of HERV-K has been associated with many diseases, such as germ cell tumors, neurotoxicity, ovarian cancer, prostate cancer, and melanoma. Indeed, two HML-2-produced proteins, Np9 and Rec, are associated with certain cancers. However, their roles in these disease associations remain unclear. The current work focused on subgroup HML-2 of HERV-K, which is recognized as the most biologically active subgroup, and aimed to explore the mechanistic basis of transcriptional activity. The results revealed that p53 deeply determined the activity of HML-2 LTR5Hs. p53 is a rather important tumor suppressor protein. It can regulate the expression of genes related to cell cycle arrest, organic processes, and apoptosis in response to cellular stress and is critical for the control of homeostasis. Previous ChIP and expression studies of individual genes suggested that p53 sites in HERV LTRs may be part of the p53 transcription program and directly regulate p53 target genes in a species-specific manner. However, the exact function of p53 in the regulation of HERV LTR expression is largely elusive. Our results clearly demonstrated the interaction between LTR5Hs of HML-2 and p53. They are of great significance for the future comprehensive study of the physiological and pathological functions of LTRs of HERVs.

Comprehensive identification and characterization of the HERV-K (HML-9) group in the human genome.

BACKGROUND: Human endogenous retroviruses (HERVs) result from ancestral infections caused by exogenous retroviruses that became incorporated into the germline DNA and evolutionarily fixed in the human genome. HERVs can be transmitted vertically in a Mendelian fashion and be stably maintained in the human genome, of which they are estimated to comprise approximately 8%. HERV-K (HML1-10) transcription has been confirmed to be associated with a variety of diseases, such as breast cancer, lung cancer, prostate cancer, melanoma, rheumatoid arthritis, and amyotrophic lateral sclerosis. However, the poor characterization of HML-9 prevents a detailed understanding of the regulation of the expression of this family in humans and its impact on the host genome. In light of this, a precise and updated HERV-K HML-9 genomic map is urgently needed to better evaluate the role of these elements in human health. RESULTS: We report a comprehensive analysis of the presence and distribution of HERV-K HML-9 elements within the human genome, with a detailed characterization of the structural and phylogenetic properties of the group. A total of 23 proviruses and 47 solo LTR elements were characterized, with a detailed description of the provirus structure, integration time, potential regulated genes, transcription factor binding sites (TFBS), and primer binding site (PBS) features. The integration time results showed that the HML-9 elements found in the human genome integrated into the primate lineage between 17.5 and 48.5 million years ago (mya). CONCLUSION: The results provide a clear characterization of HML-9 and a comprehensive background for subsequent functional studies.

Identification of Potential Diagnoses Based on Immune Infiltration and Autophagy Characteristics in Major Depressive Disorder.

Background: Major depressive disorder (MDD) is a serious mental illness characterized by mood changes and high suicide rates. However, no studies are available to support a blood test method for MDD diagnosis. The objective of this research was to identify potential peripheral blood biomarkers for MDD and characterize the novel pathophysiology. Methods: We accessed whole blood microarray sequencing data for MDD and control samples from public databases. Biological functions were analysed by GO and KEGG pathway enrichment analyses using the clusterprofile R package. Infiltrated immune cell (IIC) proportions were identified using the CIBERSORT algorithm. Clustering was performed using the ConsensusClusterPlus R package. Protein-protein interactions (PPI) were assessed by constructing a PPI network using STRING and visualized using Cytoscape software. Rats were exposed to chronic unpredictable mild stress (CUMS) for 6 weeks to induce stress behaviour. Stress behaviour was evaluated by open field experiments and forced swimming tests. Flow cytometry was used to analyse the proportion of CD8+ T cells. The expression of the corresponding key genes was detected by qRT-PCR. Results: We divided MDD patients into CD8H and CD8L clusters. The functional enrichment of marker genes in the CD8H cluster indicated that autophagy-related terms and pathways were significantly enriched. Furthermore, we obtained 110 autophagy-related marker genes (ARMGs) in the CD8H cluster through intersection analysis. GO and KEGG analyses further showed that these ARMGs may regulate a variety of autophagy processes and be involved in the onset and advancement of MDD. Finally, 10 key ARMGs were identified through PPI analysis: RAB1A, GNAI3, VAMP7, RAB33B, MYC, LAMP2, RAB11A, HIF1A, KIF5B, and PTEN. In the CUMS model, flow cytometric analysis confirmed the above findings. qRT-PCR revealed significant decreases in the mRNA levels of Gnai3, Rab33b, Lamp2, and Kif5b in the CUMS groups. Conclusion: In this study, MDD was divided into two subtypes. We combined immune infiltrating CD8+ T cells with autophagy-related genes and screened a total of 10 ARMG genes. In particular, RAB1A, GNAI3, RAB33B, LAMP2, and KIF5B were first reported in MDD. These genes may offer new hope for the clinical diagnosis of MDD.

The involvement of ADAR1 in antidepressant action by regulating BDNF via miR-432.

Brain-derived neurotrophic factor (BDNF) is a biomarker of depression. Recent studies have found adenosine deaminase acting on RNA1 (ADAR1) is a novel target being sensitive to stress at epigenetic level. The epigenetic regulation mechanism of stress-related depression is still unclear so far. To explore the potential regulating mechanism of ADAR1 on BDNF, over and low expression of ADAR1 in PC12 and SH-SY5Y cell lines are prepared. In the meanwhile, chronic unpredictable stress (CUS) mice are treated with ADAR1 inducer (interferon-γ, IFN-γ). ADAR1 regulates BDNF expression, which is proven by that over and low expressions of ADAR1 increase and decrease BDNF mRNA and protein respectively in vitro. Additionally, ADAR1 inducer alleviates the depressive-like behavior of CUS mice by recovering the decreased BDNF protein in brain and serum. Moreover, over and low expressions of ADAR1 reduce and enhance microRNA-432 (miR-432) expression respectively in vitro. Furtherly, over and low miR-432 expressions lead to decreased and increased BDNF and ADAR1 mRNA, protein and immunoreactivity respectively in vitro. The above results demonstrate that ADAR1 is involved in antidepressant action by regulating BDNF via miR-432. Those novel findings can provide a new idea for the study of epigenetic regulation mechanism, early diagnosis, and effective treatment of stress-related depression.

L-lactate preconditioning promotes plasticity-related proteins expression and reduces neurological deficits by potentiating GPR81 signaling in rat traumatic brain injury model.

Currently, there is no efficacious pharmacological treatment for traumatic brain injury (TBI). Previous studies revealed that L-lactate preconditioning has shown rich neuroprotective effects against cerebral ischemia, and therefore has the potential to improve neurological outcomes after TBI. L-lactate played a neuroprotective role by activating GPR81 in diseases of the central nervous system (CNS) such as TBI and cerebral ischemia. In this study we investigated the effects of L-lactate preconditioning on TBI and explored the underlying mechanisms. In this study, the mNSS test revealed that L-lactate preconditioning alleviates the neurological deficit caused by TBI in rats. L-lactate preconditioning significantly increased the expression of GPR81, PSD95, GAP43, BDNF, and MCT2 24 h after TBI in the cortex and hippocampus compared with the sham group. Taken together, these data suggested that L-lactate preconditioning is an effective method with which to recover neurological function after TBI. This reveals the mechanism of L-lactate preconditioning on TBI and provides a potential therapeutic method for TBI in humans.

Coexistence of Nerve Enlargement and Neuratrophy Detected by Ultrasonography in Leprosy Patients.

The purpose of this study was to evaluate peripheral neural impairment in leprosy patients by ultrasonography (US). The cross-sectional areas (CSAs) of the median (M), ulnar (U) and common fibular (CF) nerves were compared in 71 leprosy patients and 29 healthy controls, and the data were analyzed between the leprosy, multibacillary (MB)/paucibacillary (PB), reaction (R)/no reaction (NR), disability (D)/no disability (ND), and longer/shorter duration groups after treatment. We found that for the nerves located in upper limbs, the CSAs were significantly increased in the leprosy patients vs the controls; the PB group vs the MB group; the R group vs the NR group; the ND group vs the D group; and the longer duration group vs the shorter duration group at some positions of the M nerve and U nerve. In contrast, for the nerves located in lower limbs, the CSAs were significantly reduced in the leprosy patients vs the controls and in the longer duration group vs the shorter duration group at some positions of the CF nerve. This result indicated that nerve enlargement and neuratrophy coexist in leprosy patients.