Codon Optimization of the Tat Antigen of Human Immunodeficiency Virus Type 1 Generates Strong Immune Responses in Mice following Genetic Immunization

Abstract

DNA vaccines have been successful in eliciting potent immune responses in mice. Their efficiency, however, is restricted in larger animals. One reason for the limited performance of the DNA vaccines is the lack of molecular strategies to enhance immune responses. Additionally, genes directly cloned from pathogenic organisms may not be efficiently translated in a heterologous host expression system as a consequence of codon bias. To evaluate the influence of codon optimization on the immune response, we elected to use the Tat antigens of human immunodeficiency virus type 1 (HIV-1) (subtype C) and HIV-2, as these viral antigens are poorly immunogenic in natural infection and in experimental immunization and they are functionally important in viral infectivity and pathogenesis. Substituting codons that are optimally used in the mammalian system, we synthetically assembled Tat genes and compared them with the wild-type counterparts in two different mouse strains. Codon-optimized Tat genes induced qualitatively and quantitatively superior immune responses as measured in a T-cell proliferation assay, enzyme-linked immunospot assay, and chromium release assay. Importantly, while the wild-type genes promoted a mixed Th1-Th2-type cytokine profile, the codonoptimized genes induced a predominantly Th1 profile. Using a pepscan strategy, we mapped an immunodominant T-helper epitope to the core and basic domains of HIV-1 Tat. We also identified cross-clade immune responses between HIV-1 subtype B and C Tat proteins mapped to this T-helper epitope. Developing molecular strategies to optimize the immunogenicity of DNA vaccines is critical for inducing strong immune responses, especially to antigens like Tat. Our identification of a highly conserved T-helper epitope in the first exon of HIV-1 Tat of subtype C and the demonstration of a cross-clade immune response between subtypes B and C are important for a more rational design of an HIV vaccine.

HIV-1 Tat-Based Vaccines: An Overview and Perspectives in the Field of HIV/AIDS Vaccine Development

“…This evidence thereby suggests that the presence of anti-Tat antibodies may predict the clinical outcome and that the induction of humoral responses against Tat, either during the course of the natural infection or by vaccination, may contrast the progression to the disease. The predominant antibody responses to Tat have been mapped at amino acids 1-20 and 36-50, which are highly conserved among different HIV subtypes [120] and are key for Tat function and contain B [133,144] and T cell [115,[145][146][147][148][149][150][151] epitopes. However, antibody responses against other functional domains of Tat are also detected during infection, including the cysteine, basic, and carboxy-terminal regions [120-122, 126, 129, 152, 153].…”

“…Another important feature of Tat rendering it a relevant vaccine antigen is the conservation of its immunogenic regions among the HIV-1 M group at the sequence, functional, and conformational level [120,142,151,152,154,163]. Indeed, the Tat protein of 86 aa from a clade B lab-adapted HIV-1 virus (HTLV-IIIB strain, clone BH-10) isolated more than 20 years ago [164] is very well recognized by sera from African individuals infected with different virus clades, including clade C, which is responsible for more than half of new HIV-1 infections worldwide [165], with similar prevalence and epitope mapping titers of anti-Tat antibodies [120].…”

Development of Enzyme-Linked Immunosorbent Assays Using 2 Truncated ORF2 Proteins for Detection of IgG Antibodies Against Hepatitis E Virus

“…To overcome poor expression, we optimized our gene sequence for expression in E. coli; this provided several advantages, including elimination of polyadenylation sites, splicing sites, killer motifs, repeated sequences, RNA secondary structure, and high GC content [25,26]. The expression level can be improved through using host-favorite codons and lowering the GC content, since rare codons and high GC content can decrease or even prevent expression [27]. There are several approaches for expressing the HEV structural gene [21,28].…”

Tipping the Proteome with Gene-Based Vaccines: Weighing in on the Role of Nanomaterials

“…As previously mentioned, the route and method of delivery can have a profound effect on the outcome of vaccination. Codon optimization, promoter sequences, introns, enhancers, polyadenylation signals, and unmethylated cytosine-guanine dinucleotide (CpG) repeats can be modified within the plasmid to yield maximum expression of the protein product [32,[36][37][38][39][40][41]. Another popular option is using formulation adjuvants.…”