Project description:Many human monoclonal antibodies that neutralize multiple clades of HIV-1 are polyreactive and bind avidly to mammalian autoantigens. Indeed, the generation of neutralizing antibodies to the 2F5 and 4E10 epitopes of HIV-1 gp41 in man may be proscribed by immune tolerance since mice expressing the VH and VL regions of 2F5 have a block in B-cell development characteristic of central tolerance. This developmental blockade implies the presence of tolerizing autoantigens that are mimicked by the membrane-proximal external region of HIV-1 gp41. Here we identify human kynureninase (KYNU) and splicing factor 3b subunit 3 (SF3B3) as the primary conserved, vertebrate self-antigens recognized by the 2F5 and 4E10 antibodies, respectively. 2F5 binds the H4 domain of KYNU which contains the complete 2F5 linear epitope (ELDKWA). 4E10 recognizes a conformational epitope of SF3B3 that is strongly dependent on hydrophobic interactions. Opossums carry a rare KYNU H4 domain that abolishes 2F5 binding, but retain all SF3B3 4E10 epitopes. Immunization of opossums with HIV-1 gp140 induced extraordinary titers of serum antibody to the 2F5 ELDKWA epitope but little or nothing to the 4E10 determinant. Identification of structural motif shared by vertebrates and HIV-1 provides direct evidence that immunological tolerance can impair humoral responses to HIV-1. The invitrogen protoarray that contains >9,400 recombinant human proteins was used to identify self-ligands that are recognized by broadly neutralizing HIV-1 antibodies 2F5 and 4E10. An isotype-matched human myeloma protein (151K, Southern Biotech) was used as control.
Project description:A protective vaccine against HIV will likely need to induce broadly neutralizing antibodies (bnAbs) that engage relatively conserved epitopes on the HIV envelope glycoprotein (Env) trimer. We report the design of mRNA-delivered membrane-bound form of a stabilized native-like Env trimer (BG505 MD39.3) and comprehensive evaluation of B cell, T cell, and antibody responses in non-human primates.
Project description:Tissue-like memory, activated memory and resting memory B cells were sorted by FACS from the individual living with HIV (EC17) who was aviremic and transcriptomes generated using the SmartSeq2 protocol. This was to provide a reference set for each memory B cell subset in the context of HIV. Next, HIV-specific memory B cells from the individual with broadly neutralizing plasma were then also sorted by FACS and single cell transcriptomes generated using the SmartSeq2 protocol. The phenotypes of memory B cells from the individual with broadly neutralizing plasma (T125) were then inferred from the reference set using Glmnet and Celltypist packages.
