The Structure and Sequence Analysis of TLR4 Gene in Cattle

Toll-like receptor 4 (TLR4) is essential for initiating the innate response to lipopolysaccharide (LPS) from Gram-negative bacteria by acting as a signal transducting receptor. In order to help in investigating TLR4 as a candidate disease-resistance gene in cows, we isolated the cDNA (GenBank accession no. DQ839566) by RT-PCR and rapid amplification of cDNA ends (RACE) experiments and analyzed the sequence characters by bioinformatics. The results showed that cattle TLR4 gene about 3 739 bp contains an open reading frame of 2 526 bp encoded 841 amino acids (aa), 470 bp 5′ untranslated region (UTR), and 743 bp 3′ UTR. Tissue expression profile by RT-PCR indicated that TLR4 gene expresses in mammary glands, liver, muscle, duodenum, fats, uterus, kidneys, hearts, lungs, pancreas, and ovary. TLR4 protein domain predicted by bioinformatics consists of signal peptide, transmembrane helices domain, 3 sorts of leucine-rich repeat domains (LRR, LRR-TYP, and LRRCT), and a toll-interleukin1-resistance domain (TIR). Leucine-rich repeat domains were related with recognizing a broad of pathogen-associated molecular patterns (PAMP) from pathogen, and TIR domain for downstream signaling transduction was most conservative (98% identify) than other domains after alignment of protein from ovine, porcine, human, and mouse. In addition, a 470 bp 5′-flanking region sequence was amplified by PCR, and 15 putative DNA binding sites were predicted, but this sequence lacks TATA box, CCAAT character, and GC-rich regions.