scholarly journals Defining HIV-1 Envelope N-Glycan Microdomains through Site-Specific Heterogeneity Profiles

2018 ◽  
Vol 93 (1) ◽  
Author(s):  
Audra A. Hargett ◽  
Qing Wei ◽  
Barbora Knoppova ◽  
Stacy Hall ◽  
Zhi-Qiang Huang ◽  
...  
Keyword(s):  
Immune System ◽  
Immune Evasion ◽  
Immune Escape ◽  
High Resolution Mass Spectrometry ◽  
National Laboratory ◽  
Site Specific ◽  
Los Alamos National Laboratory ◽  
Glycosylation Sites ◽  
Env Protein ◽  
Hiv 1

ABSTRACT The HIV-1 envelope (Env) glycans shield the surface of Env from the immune system and form integral interactions important for a functional Env. To understand how individual N-glycosylation sites (NGS) coordinate to form a dynamic shield and evade the immune system through mutations, we tracked 20 NGS in Env from HIV-transmitted/founder (T/F) and immune escape variants and their mutants involving the N262 glycan. NGS were profiled in a site-specific manner using a high-resolution mass spectrometry (MS)-based workflow. Using this site-specific quantitative heterogeneity profiling, we empirically characterized the interdependent NGS of a microdomain in the high-mannose patch (HMP). The changes (shifts) in NGS heterogeneity between the T/F and immune escape variants defined a range of NGS that we further probed for exclusive combinations of sequons in the HMP microdomain using the Los Alamos National Laboratory HIV sequence database. The resultant sequon combinations, including the highly conserved NGS N262, N448, and N301, created an immune escape map of the conserved and variable sequons in the HMP microdomain. This report provides details on how some clustered NGS form microdomains that can be identified and tracked across Env variants. These microdomains have a limited number of N-glycan-sequon combinations that may allow the anticipation of immune escape variants. IMPORTANCE The Env protein of HIV is highly glycosylated, and the sites of glycosylation can change as the virus mutates during immune evasion. Due to these changes, the glycan location and heterogeneity of surrounding N-glycosylation sites can be altered, resulting in exposure of different glycan or proteoglycan surfaces while still producing a viable HIV variant. These changes present a need for vaccine developers to identify Env variants with epitopes most likely to induce durable protective responses. Here we describe a means of anticipating HIV-1 immune evasion by dividing Env into N-glycan microdomains that have a limited number of N-glycan sequon combinations.

Sequence Length of HIV-1 Subtype B Increases Over Time: Analyzing a Cohort of Patients with Hemophilia Over 30 Years

2021 ◽  
Author(s):  
Young-Keol Cho ◽  
Jung-eun Kim ◽  
Brian Foley
Keyword(s):  
Direct Sequencing ◽  
National Laboratory ◽  
Sequence Length ◽  
Coding Region ◽  
Los Alamos National Laboratory ◽  
Subtype B ◽  
Combined Antiretroviral Therapy ◽  
Signature Pattern ◽  
Hiv 1 ◽  
Over Time

The objective of this study is to investigate whether the sequence length of HIV-1 increases over time. A longitudinal analysis of full-length coding region sequences (FLs) in an outbreak of HIV-1 infection among patients with hemophilia and local controls identified as infected with the Korean subclade B of HIV-1 (KSB). Genes amplified by overlapping RT-PCR or nested PCR were subjected to direct sequencing. In total, 141 FLs were sequentially determined over 30 years in 62 KSB-infected patients. Non-KSB sequences were retrieved from the Los Alamos National Laboratory HIV Database. Phylogenetic analysis indicated that within KSB, 2 FLs from plasma donors O and P comprised two clusters together with 8 and 12 patients with hemophilia, respectively. Signature pattern analysis for the KSB of HIV-1 revealed signature nucleotide residues at 1.05%, compared with local controls. Additionally, in-depth FLs sequence analysis over 30 years in KSB indicates that the KSB FL significantly increases over time before combined antiretroviral therapy (cART) and decreases on cART. Furthermore, the increase in FLs over time significantly occurred in the subtypes B, C and G, but, there was no increase in the subtypes D, A, and F1. Consequently, subtypes F1 and D had the shortest sequence length. Our analysis was extended to compare HIV-1 with HIV-2 and SIVs. Essentially, the longer the sequence length (SIVsm > HIV-2 > SIVcpz > HIV-1), the longer the survival period. The increase in the length of the HIV-1 sequence over time suggests that it might be an evolutionary direction toward attenuated pathogenicity.

scholarly journals The HHV-6A Proteins U20 and U21 Target NKG2D Ligands to Escape Immune Recognition

2021 ◽  
Vol 12 ◽  
Author(s):  
Abigael Eva Chaouat ◽  
Barbara Seliger ◽  
Ofer Mandelboim ◽  
Dominik Schmiedel
Keyword(s):  
Immune System ◽  
Immune Evasion ◽  
Immune Cells ◽  
Immune Escape ◽  
Human Herpesvirus ◽  
Immune Recognition ◽  
Nkg2d Ligand ◽  
Human Immune System ◽  
Nkg2d Ligands ◽  
Infected Cells

The coevolution of the human immune system and herpesviruses led to the emergence and diversification of both cellular danger molecules recognized by immune cells on the one hand and viral countermeasures that prevent the expression of these proteins on infected cells on the other. There are eight ligands for the activating receptor NKG2D in humans – MICA, MICB, ULBP1-6. Several of them are induced and surface-expressed on herpesvirus-infected cells to serve as danger signals to activate the immune system. Therefore, these ligands are frequently targeted for suppression by viral immune evasion mechanisms. Mechanisms to downregulate NKG2D ligands and thereby escape immune recognition have been identified in all other human herpesviruses (HHV), except for HHV-6A. In this study, we identify two HHV-6A encoded immunoevasins, U20 and U21, which suppress the expression of the NKG2D ligands ULBP1 and ULBP3, respectively, during infection. Additionally, MICB is targeted by a so far unexplored viral protein. Due to the diminished NKG2D ligand surface expression on infected cells, recognition of HHV-6A infected cells by innate immune cells is impaired. Importantly, our study indicates that immune escape mechanisms between the related herpesviruses HHV-6A and HHV-6B are evolutionary conserved as the same NKG2D ligands are targeted. Our data contribute an additional piece of evidence for the importance of the NKG2D receptor – NKG2D ligand axis during human herpesvirus infections and sheds light on immune evasion mechanisms of HHV-6A.

scholarly journals Sequential Analysis of the N/O-Glycosylation of Heavily Glycosylated HIV-1 gp120 Using EThcD-sceHCD-MS/MS

2021 ◽  
Vol 12 ◽  
Author(s):  
Yong Zhang ◽  
Shanshan Zheng ◽  
Wanjun Zhao ◽  
Yonghong Mao ◽  
Wei Cao ◽  
...  
Keyword(s):  
Sequential Analysis ◽  
Viral Envelope ◽  
Viral Escape ◽  
Site Specific ◽  
Fragment Ions ◽  
Immunodeficiency Virus ◽  
Improved Performance ◽  
Env Protein ◽  
Hiv 1 ◽  
Env Glycoprotein

Deciphering the glycosylation of the viral envelope (Env) glycoprotein is critical for evaluating viral escape from the host’s immune response and developing vaccines and antiviral drugs. However, it is still challenging to precisely decode the site-specific glycosylation characteristics of the highly glycosylated Env proteins, although glycoproteomics have made significant advances in mass spectrometry techniques and data analysis tools. Here, we present a hybrid dissociation technique, EThcD-sceHCD, by combining electron transfer/higher-energy collisional dissociation (EThcD) and stepped collision energy/higher-energy collisional dissociation (sceHCD) into a sequential glycoproteomic workflow. Following this scheme, we characterized site-specific N/O-glycosylation of the human immunodeficiency virus type 1 (HIV-1) Env protein gp120. The EThcD-sceHCD method increased the number of identified glycopeptides when compared with EThcD, while producing more comprehensive fragment ions than sceHCD for site-specific glycosylation analysis, especially for accurate O-glycosite assignment. Finally, eighteen N-glycosites and five O-glycosites with attached glycans were assigned unambiguously from heavily glycosylated gp120. These results indicate that our workflow can achieve improved performance for analysis of the N/O-glycosylation of a highly glycosylated protein containing numerous potential glycosites in one process. Knowledge of the glycosylation landscape of the Env glycoprotein will be useful for understanding of HIV-1 infection and development of vaccines and drugs.

scholarly journals An antigenic atlas of HIV-1 escape from broadly neutralizing antibodies

2018 ◽  
Author(s):  
Adam S. Dingens ◽  
Dana Arenz ◽  
Haidyn Weight ◽  
Julie Overbaugh ◽  
Jesse D. Bloom
Keyword(s):  
Neutralizing Antibodies ◽  
Immune Escape ◽  
Single Amino Acid ◽  
Structural Studies ◽  
Broadly Neutralizing Antibodies ◽  
Functional Studies ◽  
Amino Acid Mutations ◽  
Env Protein ◽  
Anti Hiv ◽  
Hiv 1

SummaryAnti-HIV broadly neutralizing antibodies (bnAbs) have revealed vaccine targets on the virus’s Env protein and are themselves promising immunotherapeutics. The efficacy of bnAb-based therapies and vaccines depends in part on how readily the virus can escape neutralization. While structural studies can define contacts between bnAbs and Env, only functional studies can define mutations that confer escape. Here we map how all single amino-acid mutations to Env affect neutralization of HIV by nine bnAbs targeting five epitopes. For most bnAbs, mutations at only a small fraction of structurally defined contact sites mediated escape, and most escape occurred at sites that are near but do not directly contact the antibody. The mutations selected by two pooled bnAbs were similar to those expected from the combination of the bnAbs’ independent action. Overall, our mutation-level antigenic atlas provides a comprehensive dataset for understanding viral immune escape and refining therapies and vaccines.

Identification and Characterization of Positively Selected Mutations in Nef of Four HIV-1 Major Subtypes from Los Alamos National Laboratory

2018 ◽  
Vol 16 (2) ◽  
pp. 130-142
Author(s):  
Elnaz Shadabi ◽  
Binhua Liang ◽  
Frank Plummer ◽  
Ma Luo
Keyword(s):  
National Laboratory ◽  
Alamos National Laboratory ◽  
Los Alamos ◽  
Evolutionary Patterns ◽  
Los Alamos National Laboratory ◽  
Immune Pressure ◽  
Subtype B ◽  
Clade B ◽  
Clade C ◽  
Hiv 1

Background:Human immunodeficiency virus-1 (HIV-1) mutates rapidly to escape host immune pressure. This results in the generation of positively selected mutations (PSM) throughout the viral genome. Escape mutations in Nef, one of the accessory proteins of HIV-1, which plays an important role in viral pathogenicity have previously been identified in several large cohort studies, but the evolution of PSMs overtime in various HIV-1 subtypes remains unknown.Methods:161 clade A1, 3093 clade B, 647 clade C and 115 clade D HIV-1 nef sequences were obtained from the HIV Database of Los Alamos National Laboratory and aligned using MEGA 6.0. The sequences from each clade were grouped based on the year of collection. Quasi analysis was used to identify PSMs and the number and locations of PSMs were compared among different subtypes.Results:PSMs for all four subtypes were distributed across the sequence of Nef, and conserved residues F90, W113, PxxPxR (a.a 72-77) remain unaltered overtime. The frequency of PSMs was stable among subtype B sequences but increased overtime for other subtypes. Phylogenetic analysis shows that sequences containing PSMs tend to cluster together at both inter and intra- subtype levels.Conclusion:Identification of PSMs and their changes overtime within various subtypes of HIV-1 is important in defining global viral evolutionary patterns that can provide insights for designing therapeutic strategies.

scholarly journals Novel gRNA design pipeline to develop broad-spectrum CRISPR/Cas9 gRNAs for safe targeting of the HIV-1 quasispecies in patients

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Neil T. Sullivan ◽  
Will Dampier ◽  
Cheng-Han Chung ◽  
Alexander G. Allen ◽  
Andrew Atkins ◽  
...  
Keyword(s):  
Broad Spectrum ◽  
In Silico ◽  
National Laboratory ◽  
Viral Gene Expression ◽  
High Sensitivity ◽  
Viral Gene ◽  
Stem Loop ◽  
Los Alamos National Laboratory ◽  
Hiv 1

AbstractThe CRISPR/Cas9 system has been proposed as a cure strategy for HIV. However, few published guide RNAs (gRNAs) are predicted to cleave the majority of HIV-1 viral quasispecies (vQS) observed within and among patients. We report the design of a novel pipeline to identify gRNAs that target HIV across a large number of infected individuals. Next generation sequencing (NGS) of LTRs from 269 HIV-1-infected samples in the Drexel CARES Cohort was used to select gRNAs with predicted broad-spectrum activity. In silico, D-LTR-P4-227913 (package of the top 4 gRNAs) accounted for all detectable genetic variation within the vQS of the 269 samples and the Los Alamos National Laboratory HIV database. In silico secondary structure analyses from NGS indicated extensive TAR stem-loop malformations predicted to inactivate proviral transcription, which was confirmed by reduced viral gene expression in TZM-bl or P4R5 cells. Similarly, a high sensitivity in vitro CRISPR/Cas9 cleavage assay showed that the top-ranked gRNA was the most effective at cleaving patient-derived HIV-1 LTRs from five patients. Furthermore, the D-LTR-P4-227913 was predicted to cleave a median of 96.1% of patient-derived sequences from other HIV subtypes. These results demonstrate that the gRNAs possess broad-spectrum cutting activity and could contribute to an HIV cure.

HIV-1 POPULATION DYNAMICS IN VIVO: IMPLICATIONS FOR PATHOGENESIS, EFFECT OF CYTOKINE AND THERAPY STRATEGY

2004 ◽  
Vol 12 (03) ◽  
pp. 315-333
Author(s):  
JIE LOU ◽  
ZHIEN MA ◽  
MEIZHI LOU ◽  
YIMING SHAO
Keyword(s):  
Immune Response ◽  
T Cells ◽  
Immune System ◽  
Immune Escape ◽  
Interleukin 2 ◽  
Human Immune System ◽  
Therapy Strategy ◽  
The Impact ◽  
Hiv 1

In this paper, we present a dynamical model to study the spread of HIV-1 in vivo. Our goal is to better understand the interaction between HIV-1 and the human immune system. Making use of the Hill function, we describe two kinds of processions occurring in the immune response: the activation interactions or inhibitory interactions occurring between different components in the immune response, and the autocatalytic maintenance of the CD4+ T cells and CD8+ T cells populations. We also consider the impact of the cytokine interleukin-2 (IL-2) and the CD8 antiviral factor (CAF) on HIV-1 infection. Through numerical simulations we get several results. First, we find that the effects of IL-2 and CAF in the treatment for the infected are limiting. Namely, the curative effect will not always increase along with the dose of IL-2 or CAF or both. The increasing trend will stagnate at a certain dose that we used. Second, we find some possible reasons for the collapse of the lymph system in HIV-1 infection — the loss of these restraining functions, and/or the genetic variability of the virus due to immune escape that enhances the virulence, which then bring the collapse of the immune system. In some conditions the system will produce a Hopf bifurcation. We also simulate the theoretical warrant of the feasibility of the combined chemotherapy strategies for the HIV-1 infected patient.

scholarly journals Glycan Microheterogeneity at the PGT135 Antibody Recognition Site on HIV-1 gp120 Reveals a Molecular Mechanism for Neutralization Resistance

2015 ◽  
Vol 89 (13) ◽  
pp. 6952-6959 ◽  
Author(s):  
Laura K. Pritchard ◽  
Daniel I. R. Spencer ◽  
Louise Royle ◽  
Snezana Vasiljevic ◽  
Stefanie A. Krumm ◽  
...  
Keyword(s):  
Molecular Mechanism ◽  
Neutralizing Antibodies ◽  
Vaccine Design ◽  
Broadly Neutralizing Antibodies ◽  
Site Specific ◽  
Antibody Recognition ◽  
Glycosylation Sites ◽  
Minor Population ◽  
A Minor ◽  
Hiv 1

Broadly neutralizing antibodies have been isolated that bind the glycan shield of the HIV-1 envelope spike. One such antibody, PGT135, contacts the intrinsic mannose patch of gp120 at the Asn332, Asn392, and Asn386 glycosylation sites. Here, site-specific glycosylation analysis of recombinant gp120 revealed glycan microheterogeneity sufficient to explain the existence of a minor population of virions resistant to PGT135 neutralization. Target microheterogeneity and antibody glycan specificity are therefore important parameters in HIV-1 vaccine design.

scholarly journals P03.15 Site-specific immune evasion and substantial heterogeneity within entities provide evidence for personalized immunotherapy

2020 ◽  
Vol 8 (Suppl 2) ◽  
pp. A28-A29
Author(s):  
M Thelen ◽  
K Wennhold ◽  
J Lehmann ◽  
E Staib ◽  
MA Garcia Marquez ◽  
...  
Keyword(s):  
T Cells ◽  
Antigen Presentation ◽  
Immune Evasion ◽  
Immune Cell ◽  
Immune Escape ◽  
Immune Cell Infiltration ◽  
Site Specific ◽  
Immune Escape Mechanisms ◽  
Substantial Heterogeneity ◽  
Research Grant

BackgroundImmune-checkpoint inhibition (CKI) demonstrated remarkable therapeutic efficacy in several kinds of cancer. However, immune escape mechanisms lead to primary or secondary resistance in the majority of patients. Most predictive biomarkers failed, as the primary target of CKI is not the tumor cell itself, but the crosstalk between immune- and cancer cells. We aimed to characterize the immune evasion landscape in primary tumors across different entities.Materials and MethodsExpression of 32 immune-regulatory molecules on lymphocytes was analyzed in peripheral blood and tumor infiltrating lymphocytes (TILs) of 146 primary tumor patients across 10 different entities using flow cytometry. NanoString was applied to determine RNA expression of the respective ligands and 20 genes associated with antigen presentation. Expression of coinhibitory ligands on tumor cells was assessed by immunohistochemistry. To quantify the immune cell infiltration, digital pathology was used and the Immunoscore was generated for each patient.ResultsWhile an increase of regulatory T cells was a common feature across all entities, we found site-specific differences regarding other lymphocyte subsets and expression of immune-regulatory molecules by TILs and tumor cells. Expression of co-inhibitory molecules on tumor infiltrating T cells accumulated especially in advanced stage cancers whereas immune cell infiltration was mainly associated with enhanced antigen presentation. Co-expression of multiple immune-inhibitory ligands was most frequent in colorectal, lung and ovarian carcinoma. Genes related to antigen presentation were frequently dysregulated in seminoma, liver and lung cancer.ConclusionsImmune evasion is a common feature of cancer and frequently detected co-occurrence of multiple mechanisms probably contributes to resistance against immunotherapy. We describe substantial heterogeneity regarding immune escape mechanisms between patients with the same primary tumor. Individualized immunotherapeutic strategies based on pretherapeutic evaluation of the immune evasion landscape might help to improve response to CKI.Disclosure InformationM. Thelen: None. K. Wennhold: None. J. Lehmann: None. E. Staib: None. M.A. Garcia Marquez: None. P. Lohneis: None. A. Lechner: None. S. Wagener-Ryczek: None. P.S. Plum: None. D. Pfister: None. F. Dörr: None. D. Beutner: None. F. Thangarajah: None. D. Ratiu: None. W. Malter: None. S. Merkelbach-Bruse: None. C.J. Bruns: None. A. Quaas: None. M.S. von Bergwelt-Baildon: B. Research Grant (principal investigator, collaborator or consultant and pending grants as well as grants already received); Significant; Astellas. F. Consultant/Advisory Board; Modest; Bristol-Myers Squibb. H.A. Schlößer: B. Research Grant (principal investigator, collaborator or consultant and pending grants as well as grants already received); Significant; Astra Zeneca.

QUASI analysis of the HIV-1 envelope sequences in the Los Alamos National Laboratory HIV sequence database: pattern and distribution of positive selection sites and their frequencies over yearsThis paper is one of a selection of papers in this Special Issue, entitled International Symposium on Recent Advances in Molecular, Clinical, and Social Medicine, and has undergone the Journal's usual peer-review process.

2007 ◽  
Vol 85 (2) ◽  
pp. 259-264 ◽  
Author(s):  
Binhua Liang ◽  
Ma Luo ◽  
T. Blake Ball ◽  
Francis A. Plummer
Keyword(s):  
Positive Selection ◽  
National Laboratory ◽  
Sequence Database ◽  
Alamos National Laboratory ◽  
Los Alamos ◽  
Los Alamos National Laboratory ◽  
Central Target ◽  
Conserved Regions ◽  
Geographical Regions ◽  
Hiv 1

The envelope (env) protein of human immunodeficiency virus type 1 (HIV-1) plays a crucial role in virus entry and is a central target for HIV vaccine design. Using the QUASI program, we analyzed the conserved regions of all currently available env sequences in the Los Alamos National Laboratory HIV Sequence Database and identified positive selection (PS) sites that are likely to be restricted by host immune responses. We found that PS sites are dispersed across conserved regions of env sequence, and that the C3, C4, and C5 regions were the most targeted. Several regions were identified as being PS free and were mainly distributed in the C1 and C2 regions. When comparing individual QUASI PS site frequencies across clades and geographical regions with the overall frequency of the entire env database, the env sequences from North America showed significantly lower PS site frequency, while those from Asia were significantly higher using Student's t test. The QUASI PS site frequency of env proteins from viruses isolated from different years showed that the PS site frequencies of the env population increased over time. Our study provides an overview of PS sites across the conserved regions of HIV-1 env sequences.

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