Polymorphisms in the promoter region of TANK-binding kinase1 are associated with Gram-positive bloodstream infections

Abstract number: O353

Wolffs P., Beuken E., Ouburg S., Morré S., Stassen F.

Objectives: It is long known that not all individuals with a specific disease present with the same clinical manifestations, nor do they have identical prognoses or responses to treatments. It has become clear that variations in the human genome are likely to have an impact on these aspects. TANK-binding kinase 1 (TBK1) is a central molecule in the induction of a.o. the type I interferon response to pathogens. Our goals for this study were 1) to investigate the frequency of single nucleotide polymorphisms (SNPs) in the promoter and coding region of TBK1 in a Dutch Caucasian population and 2) to search for potential associations between these SNPs and bloodstream infections.

Methods: Whole blood samples or samples of positive blood cultures were collected and after genomic DNA was isolated, PCR and sequencing were performed for SNP identification. Functional studies included promoter activity measurements using a luciferase assay as well as electrophoretic mobility shift assays (EMSA) to study binding of the transcription factor USF1 to the wt and mutant promoter. SNP incidences were studied in a case control study.

Results: In samples from Dutch Caucasian healthy volunteers, 4 SNPs were found with allele frequencies higher than 5% whereas 6 other known SNPs had frequencies lower than 5% in our cohort. Two SNPs (rs89208169 and rs89208163) located in the promoter region were studied in a larger cohort of 350 anonymised patients from the Maastricht University Medical Center with either Gram-positive or Gram-negative blood cultures. We found that the prevalence of rs89208169 was significantly increased in patients with positive blood cultures in comparison with those with negative blood cultures or healthy volunteers. Further investigation of this SNP showed that it is located just outside a USF1-binding site. Measuring the promoter activity using luciferase assays, the mutant promoter exhibited a decreased activity of <35%. This observation was confirmed by EMSA which showed that recombinant USF1 protein had a reduced binding affinity to the mutant promoter.

Conclusions: SNP rs89208169 in the promoter region of TBK1 has a significant association with Gram-positive infections. Our results demonstrate that this is likely due to a decreased expression of TBK1 due to reduced binding of the transcription factor USF1 to the mutant promoter. Our results support recent findings that TBK1 plays also an important role in the host response to Gram-positive infections.