Evolution of the Bovine TLR Gene Family and Member Associations with Mycobacterium avium Subspecies paratuberculosis Infection

Fisher, Colleen A.; Bhattarai, Eric K.; Osterstock, Jason B.; Dowd, Scot E.; Seabury, Paul M.; Vikram, Meenu; Whitlock, Robert H.; Schukken, Ynte H.; Schnabel, Robert D.; Taylor, Jeremy F.; Womack, James E.; Seabury, Christopher M.

Surprisingly, the region of TLR3 demonstrating the strongest deviation from neutrality does not include the two nonsynonymous SNPs predicted to impact protein function ( Table 3, Table 4 ), but includes a 59 putative promoter region (PROSCAN 1.7: http:// www-bimas.cit.nih.gov/molbio/proscan/index.html) [23] harboring several transcription factor binding sites (NF-kB, PEA1, AP-1, TFIID; Positions 2852041-2852291 of NW_001494406.2) as well as the first two exons and introns of TLR3. No variation was detected within the predicted promoter itself. However, 40 validated SNPs were found to flank the putative promoter (see Table S2 for coordinates), with nearly half of this variation occurring immediately upstream (n = 19 SNPs). Further evaluation of LD between adjacent variable sites for taurine cattle revealed two regions of TLR3 with persistent, unbroken r 2 .0.50 between all adjacent sites as follows: 1) Variable sites 1-5 upstream of the predicted promoter (Table S2) ; and 2) Variable sites 10-19, which span the predicted promoter. This unbroken pattern of persistent r 2 was also detected in our pooled analysis of all cattle, but did not extend across as many adjacent variable sites (Table S2 , sites 13-17; region also spans the predicted promoter), and was only found in one upstream region. Therefore, it is possible that selection is primarily operating on noncoding variation within the genomic regions flanking the predicted promoter. Future functional studies will be needed to determine whether the SNPs flanking the predicted TLR3 promoter actually modulate differences in gene expression.