Abstract 321: The E12/id3 Axis Regulates Ccr6 Expression In B Cells Which Are Associated With Human Atherosclerosis

Abstract

Background Aortic homing of B cells and B cell mediated atheroprotection depend on the helix-loop-helix (HLH) transcription factor Id3. We identified that CCR6, a downstream target of Id3, is also essential for B cell homing to the aorta. Id3 functions through dominant negative inhibition of basic-HLH transcription factors such as E proteins. We have shown that a non-synonomous single nucleotide polymorphism in Id3 correlates with human carotid intimal medial thickness (cIMT). The protein encoded by this risk allele shows attenuated interaction with E12 yet whether this property is shared with other E proteins and the potential impact of Id3 and E12 on CCR6 expression in B cell subsets is unknown. Results Co-IPs of ancestral and polymorphic Id3 with the four known E proteins showed that E12, and not other E proteins, had significantly decreased interaction with polymorphic Id3. To address the regulatory role of Id3 on CCR6 in B cells, B cell subsetting by flow cytometry was done showing that B cell specific Id3 knockout mice (Id3fl/flApoe-/-CD19cre/+) indeed have significantly fewer CCR6 positive B cells which is in agreement with our previous findings in global Id3 knockout mice. To address the regulatory role of E12 on CCR6 expression similar subsetting of global E12 knockout mice (E12-/-Apoe-/-) was done showing they have significantly more CCR6 positive B cells of all subsets (B1a, B1b, B2) compared to wildtype. In particular the atheroprotective B1a subset had the greatest percentage increase due to loss of E12 compared to other subsets. Finally, we addressed whether CCR6 on human B cells correlates with human atherosclerosis. We found that coronary plaque burden and stenosis as measured by intravascular ultrasound (IVUS) in patients undergoing coronary catheterization correlated inversely with the average amount of CCR6 on B cells. Conclusions Our data suggests that Id3 is a key activator of CCR6 expression in B cells through dominant negative inhibition of E12. Furthermore, it raises the interesting possibility that CCR6 could play an important atheroprotective role through its regulation of B cell homing in humans.