HLA‐A promoter, coding, and 3′UTR sequences in a Brazilian cohort, and their evolutionary aspects

Abstract

HLA‐A is the second most polymorphic locus of the human leucocyte antigen (HLA) complex encoding a key molecule for antigen presentation and NK cell modulation. Many studies have evaluated HLA‐A variability in worldwide populations, focusing mainly on exons, but the regulatory segments have been poorly characterized. HLA‐A variability is particularly high in the segment encoding the peptide‐binding groove (exons 2 and 3), which is related to the antigen presentation function and the balancing selection in these segments. Here we evaluate the genetic diversity of the HLA‐A gene considering a continuous segment encompassing the extended promoter (1.5 kb upstream of the first translated ATG), all exons and introns, and the entire 3′ untranslated region, by using massively parallel sequencing. To achieve this goal, we used a freely available bioinformatics workflow that optimizes read mapping for HLA genes and defines complete sequences using either the phase among variable sites directly observed in sequencing data and probabilistic models. The HLA‐A variability detected in a highly admixed population sample from Brazil shows that the HLA‐A regulatory segments present few, but divergent sequences. The regulatory segments are in close association with the coding alleles. Both exons and introns are highly variable. Moreover, patterns of molecular diversity suggest that the promoter, in addition to the coding region, might be under the same selective pressure, but a different scenario arises when it comes to exon 4 and the 3′UTR segment.

HLA‐C genetic diversity and evolutionary insights in two samples from Brazil and Benin

“…This low sequence depth is not related to mapping bias because further investigations using other library preparation kits circumvented this issue, indicating the underrepresentation was due to a fragmentation bias of Nextera XT kit, which affects especially CG‐rich regions, such as intron 2. This was previously reported for the HLA‐A locus 32 .…”

“…This pattern was also observed among Mandenka 58 . Since the α3 domain interacts with the CD8 co‐receptor to facilitate TCR signaling, we expected conservation of this HLA‐C segment as was observed for HLA‐A 32 . In spite of that, the HLA‐C α3/CD8 interaction may be preserved because amino acids located within residues 222‐245 that are important to HLA/CD8 interaction are preserved 72‐74 with only one rare variant (Glu229Gln) within this region (Table S3) that is associated with allele HLA‐C*15:13 .…”

“…As observed for other HLA genes, such as HLA‐A and HLA‐G 29,32 , there was a strong LD across the entire HLA‐C locus , with a single segregation block (Figure S1) and many variants in complete LD. This LD profile supports the close relationship among the coding and regulatory sequences.…”

“…This is intriguing because exon 1 encodes the HLA‐C leader peptide and it is primarily involved in targeting HLA‐C to the cellular membrane, and it is not part of the mature HLA‐C molecule. This pattern was not observed for HLA‐A in this same Brazilian sample 32 …”

“…Thus, it is not clear (i) how HLA‐C presents both features considering its polymorphic nature and expression pattern, and (ii) how natural selection has shaped this gene's variability because the coding region of classical and nonclassical HLA genes reflect different selection profiles. Both HLA‐G and HLA‐E coding regions are under the influence of purifying selection, 29,30 while balancing selection acting on the segments encoding the peptide‐binding groove of classical HLA molecules is well documented 31‐36 …”