Identification of Siglec-1 null individuals infected with HIV-1

Abstract

Siglec-1/CD169 is a myeloid-cell surface receptor critical for HIV-1 capture and infection of bystander target cells. To dissect the role of SIGLEC1 in natura, we scan a large population genetic database and identify a loss-of-function variant (Glu88Ter) that is found in ∼1% of healthy people. Exome analysis and direct genotyping of 4,233 HIV-1-infected individuals reveals two Glu88Ter homozygous and 97 heterozygous subjects, allowing the analysis of ex vivo and in vivo consequences of SIGLEC1 loss-of-function. Cells from these individuals are functionally null or haploinsufficient for Siglec-1 activity in HIV-1 capture and trans-infection ex vivo. However, Siglec-1 protein truncation does not have a measurable impact on HIV-1 acquisition or AIDS outcomes in vivo. This result contrasts with the known in vitro functional role of Siglec-1 in HIV-1 trans-infection. Thus, it provides evidence that the classical HIV-1 infectious routes may compensate for the lack of Siglec-1 in fuelling HIV-1 dissemination within infected individuals.

Figure 1. Location and frequency of SIGLEC1 protein truncating variants.

Protein domains are represented in different colours. Data from the Exome Aggregation Consortium (exac.broadinstitute.org) identifies 70 protein truncating variants in SIGLEC1 including 33 stop gain (red), 24 frameshift (yellow) and 12 splice disrupting (blue) variants. Grey boxes indicate amino acid blocks encoded by each exon. With the exception of Glu88Ter, all protein truncating variants occur at <1% frequency. Glu88Ter is located in the V-set domain of Siglec-1, the region that recognizes sialyllactose in HIV-1 membrane gangliosides.

Figure 2. Siglec-1 expression and trans-infection across distinct SIGLEC1 genotypes.

Monocytes were isolated and cultured 24 h in the presence of 1,000 U ml⁻¹ of IFNα to induce Siglec-1 expression. (a) Quantification of Siglec-1 expression levels assessed by flow cytometry. Empty box represents a repeat analysis of one Siglec-1 null homozygote. (b) Capture of fluorescent HIV-1 VLPs by monocytes from distinct genotypes previously exposed to isotype or α-Siglec-1 mAbs. Geometric mean fluorescence intensity of monocytes not exposed to VLPs is also depicted to show the background levels of the assay (empty bars). (c) Correlation between Siglec-1 expression levels and viral capture values of isotype-treated monocytes. (d) HIV-1 transmission to a reporter CD4⁺ cell line from monocytes of opposing homozygous individuals pre-incubated with isotype or α-Siglec-1 mAbs. HIV-1 infection of reporter cells was determined by induced luciferase activity. Data show mean relative light units and SEM of cells from two homozygous individuals with the common allele and one Siglec-1 null homozygote.

Figure 3. Analysis of association between Siglec-1 Glu88Ter and HIV-1 clinical outcomes in the absence of antiretroviral treatment.

(a) Set point viral load of individuals from the SHCS with or without the Glu88Ter allele (n=2,243). (b) CD4⁺ T-cell count dynamics of individuals from the SHCS with or without the Glu88Ter allele (n=3,385). CD4⁺ T-cell counts (cells mm⁻³) were binned using 500 days windows, counting backwards from the date of antiretroviral treatment start or loss of follow-up. Median CD4⁺ T-cell values (lines) and interquartile ranges in each bin (shaded areas) are shown for individuals carrying no copies (n=3,305) or one copy (n=78) of the Siglec-1 Glu88Ter allele. Actual CD4⁺ T-cell values are shown for the two Siglec-1 Glu88Ter homozygotes. (c) Plasma viral RNA level (cp ml⁻¹) and CD4⁺ T-cell count (cells mm⁻³) dynamics of the two Siglec-1 Glu88Ter homozygotes. The dates of first HIV-1 positive report and of ART initiation are depicted. Open circles indicate negative values bellow the represented detection level. (d) Time to AIDS measured in the SHCS cohort (n=2,458 common allele Glu88; n=53 Glu88Ter including 52 heterozygous and 1 homozygous individuals) or (e) the MACS cohort (n=401 common allele Glu88; n=12 Glu88Ter which were all heterozygous).