Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows

Lyubov Kalashnikova; Irina Bagal; Nina Murugina; Roman Senina; Vladimir Kalashnikov; Shamil Meshcherov

doi:10.33988/auvfd.1863540

Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows

Abstract

Toll-like receptor 2 (TLR2) is a critical component of the innate immune system. TLR2, a membrane-bound receptor, is expressed on the surface of specific immune cells, where it is involved in recognizing microbial components and initiating signaling cascades that activate immune cells, thereby playing a pivotal role in early pathogen detection and activating immune responses. This study investigated the effect of Toll-like receptor 2 gene polymorphism on the expression of genes associated with innate immune cell activation. The PCR-RFLP method was used to study single nucleotide polymorphism in exon 2 of the TLR2 gene 385 T/G (NC_037344.1 rs55617172) in 103 Holstein cows. The frequency of the TLR2T allele was 0.74, and that of the TLR2TT genotype was 53.4%. The effect of the TLR2 gene polymorphism on the expression of the genes of proinflammatory cytokines tumor necrosis factor (TNF), interleukin 6 (IL6), and interleukin 12B (IL12B) in peripheral blood mononuclear cells (PBMCs) from 29 cows was shown when PBMCs were stimulated with the TLR2 agonist lipoteichoic acid (LTA) or the Toll-like receptor 4 (TLR4) agonist lipopolysaccharide (LPS). PBMCs obtained from animals carrying the G allele of the TLR2 gene showed significantly increased expression of the proinflammatory cytokine genes TNF, IL6, and IL12B following stimulation with TLR2 or TLR4 agonists. The TLR2 genotype did not significantly affect the expression of genes encoding the Toll-like receptors TLR2 and TLR4 or the adaptor molecules MYD88 and TICAM2.

Keywords

Project Number

project No. 23-26-00235

References

Bartens MC, Gibson AJ, Etherington GJ, et al (2021): Single nucleotide polymorphisms in the bovine tlr2 extracellular domain contribute to breed and species-specific innate immune functionality. Front Immunol, 12, 764390. DOI:10.3389/fimmu.2021.764390.
Benedict CA (2003): Viruses and the TNF-related cytokines, an evolving battle. Cytokine Growth Factor Rev, 14, 349-357. DOI: 10.1016/s1359-6101(03)00030-3.
Dalkıran S, Yakan A, Özkan H (2025): Effects of melatonin on specific gene expression in the ovaries of superovulated rats. Ankara Univ Vet Fak Derg, 72, 497-507. DOI:10.33988/auvfd.1642763.
Facci MR, Auray G, Meurens F, et al (2011): Stability of expression of reference genes in porcine peripheral blood mononuclear and dendritic cells. Vet Immunol Immunopathol, 141, 11–15. DOI: 10.1016/j.vetimm.2011. 01.005.
Francisco S, Billod JM, Merino J, et al (2022): Induction of TLR4/TLR2 interaction and heterodimer formation by low endotoxic atypical LPS. Front Immunol, 12, 748303. DOI: 10.3389/fimmu.2021.748303.
Kagan JC, Su T, Horng T, et al (2008): TRAM couples’ endocytosis of Toll-like receptor 4 to the induction of interferon-beta. Nat Immunol, 9, 361–368. DOI: 10.1038/ni1569.
Kalashnikova LA, Bagal IE, Kalashnikov VE, et al (2023): Polymorphism of the immune system genes TNF and TLR2 in Kholmogory and Holstein cattle. Zh Animal Science, 12, 2-5. DOI:10.25708/ZT.023.65.55.001.
Koets A, Santema W, Mertens H, et al (2010): Susceptibility to paratuberculosis infection in cattle is associated with single nucleotide polymorphisms in Toll-like receptor 2 which modulate immune responses against Mycobacterium avium subspecies paratuberculosis. Prev Vet Med, 93, 305-315. DOI: 10.1016/j.prevetmed.2009. 11.008.

Lane K, Van Valen D, DeFelice MM, et al (2017): Measuring signaling and RNA-seq in the same cell links gene expression to dynamic patterns of NF-κB activation. Cell Syst, 4, 458–469. DOI: 10.1016/j.cels.2017.03.010.
Li S, Cao M, Song L, et al (2016): The contribution of toll-like receptor 2 on Helicobacter pylori activation of the nuclear factor-kappa B signaling pathway in gastric epithelial cells. Microb Pathog, 98, 63-68. DOI: 10.1016/j.micpath.2016.06.028.
Liu Y, Wang Y, Yamakuchi M, et al (2001): Upregulation of toll-like receptor 2 gene expression in macrophage response to peptidoglycan and high concentration of lipopolysaccharide is involved in NF-kappa b activation. Infect Immun, 69, 2788–2796. DOI: 10.1128/IAI.69.5. 2788-2796.2001.
Livak KJ, Schmittgen TD (2001): Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods, 25, 402–408. DOI: 10.1006/meth.2001.1262.
Lu YC, Yeh WC, Ohashi PS (2008): LPS/TLR4 signal transduction pathway. Cytokine, 42, 145–151. DOI: 10.1016/j.cyto.2008.01.006.
Lüttgenau J, Herzog K, Strüve K, et al (2016): LPS-mediated effects and spatio-temporal expression of TLR2 and TLR4 in the bovine corpus luteum. Reproduction, 151, 391-399. DOI: 10.1530/REP-15-0520.
Mashhadi Abolghasem Shirazi M, Sadat SM, Haghighat S, et al (2023): Alum and a TLR7 agonist combined with built-in TLR4 and 5 agonists synergistically enhance immune responses against HPV RG1 epitope. Sci Rep, 13, 16801. DOI: 10.1038/s41598-023-43965-3.
Maurić Maljković M, Vlahek I, Piplica A, et al (2023): Prospects of toll-like receptors in dairy cattle breeding. Anim. Genet, 54, 425–434. DOI: 10.1111/age.13325.
Mucha R, Bhide MR, Chakurkar EB, et al (2009): Toll-like receptors TLR1, TLR2 and TLR4 gene mutations and natural resistance to Mycobacterium avium subsp. paratuberculosis infection in cattle. Vet Immunol Immunopathol, 128, 381-388. DOI: 10.1016/j.vetimm. 2008.12.007.
Napetschnig J, Wu H (2013): Molecular basis of NF-κB signaling. Ann Rev Biophys, 42, 443–468. DOI: 10.1146/annurev-biophys-083012-130338.
Oliveira-Nascimento L, Massari P, Wetzler LM (2012): The role of TLR2 in infection and immunity. Front Immunol, 3, 79. DOI: 10.3389/fimmu.2012.00079.
Oner Y, Ata N, Karabaş M, et al (2022): Investigation of TLR2 -EcoRV, NOD2- BsaHI ve IFNγ- HphI allele frequency distribution among Turkish Native Cattle Breeds. Turk J Agric Food Sci Technol, 10, 992-996. DOI: 10.24925/turjaf.v10i6.992-996.4701.
Opsal MA, Lien S, Brenna-Hansen S, et al (2008): Association analysis of the constructed linkage maps covering TLR2 and TLR4 with clinical mastitis in Norwegian Red cattle. J Anim Breed Genet, 125, 110-118. DOI: 10.1111/j.1439-0388.2007.00704.x.
Pashenkov MV, Murugina NE, Budikhina AS, et al (2019): Synergistic interactions between NOD receptors and TLRs: Mechanisms and clinical implications. J Leukoc Biol, 105, 669–680. DOI: 10.1002/ JLB.2RU0718-290R.
Pattabiraman G, Panchal R, Medvedev AE (2017): The R753Q polymorphism in Toll-like receptor 2 (TLR2) attenuates innate immune responses to mycobacteria and impairs MyD88 adapter recruitment to TLR2. J Biol Chem, 292, 10685-10695. DOI: 10.1074/jbc.M117.784470.
Piao W, Ru LW, Toshchakov VY (2016): Differential adapter recruitment by TLR2 co-receptors. Pathog Dis, 74, ftw043. DOI: 10.1093/femspd/ftw043.
Prebavathy T, Thanislass J, Dhanammal L, et al (2015): Association between SNPs in TLR2 gene segment corresponding to LRR functional domain of TLR2 receptor and bovine mastitis. Asian J Anim Sci, 9, 45-56. DOI: 10.3923/ajas.2015.45.56.
Proença MA, de Oliveira JG, Cadamuro AC, et al (2015): TLR2 and TLR4 polymorphisms influence mRNA and protein expression in colorectal cancer. World J Gastroenterol, 21, 7730-7741. DOI: 10.3748/wjg.v21.i25. 7730.
Rainard P, Fromageau A, Cunha P, et al (2008): Staphylococcus aureus lipoteichoic acid triggers inflammation in the lactating bovine mammary gland. Vet Res, 39, 52. DOI: 10.1051/vetres:2008034.
Roy JG, McElhaney JE, Verschoor CP (2020): Reliable reference genes for the quantification of mRNA in human T-cells and PBMCs stimulated with live influenza virus. BMC Immunol, 21, 4. DOI: 10.1186/s12865-020-0334-8.
Thirunavukkarasu S, de Silva K, Whittington RJ, et al (2013): In vivo and in vitro expression pattern of Toll-like receptors in Mycobacterium avium subspecies paratuberculosis infection. Vet Immunol Immunopathol, 156, 20-31. DOI: 10.1016/j.vetimm.2013.08.008.
Yilmaz A, Onen HI, Alp E, et al (2012): Real-Time PCR for Gene Expression Analysis. 229-254. In: Polymerase Chain Reaction. InTech, Rijeka, Croatia.
Zhang LP, Gan QF, Ma TH, et al (2009). Toll-like receptor 2 gene polymorphism and its relationship with SCS in dairy cattle. Anim Biotechnol, 20, 87-95. DOI: 10.1080/10495390902873096.

Details

Primary Language

English

Subjects

Veterinary Immunology, Animal Science, Genetics and Biostatistics

Journal Section

Research Article

Authors

Lyubov Kalashnikova This is me
0000-0002-9760-5254
Russian Federation

Irina Bagal ^*
0009-0000-9365-0544
Russian Federation

Nina Murugina This is me
0000-0001-7000-5729
Russian Federation

Roman Senina This is me
0009-0007-9131-7677
Russian Federation

Vladimir Kalashnikov This is me
0009-0009-4259-1957
Russian Federation

Shamil Meshcherov This is me
0000-0001-5500-8753
Russian Federation

Early Pub Date

June 18, 2026

Publication Date

July 1, 2026

Submission Date

January 15, 2026

Acceptance Date

April 28, 2026

Published in Issue

Year 2026 Volume: 73 Number: 3

DOI

https://doi.org/10.33988/auvfd.1863540

IZ

https://izlik.org/JA84YR45KK

Cite

RIS / Bibtex

APA

Kalashnikova, L., Bagal, I., Murugina, N., Senina, R., Kalashnikov, V., & Meshcherov, S. (2026). Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows. Ankara Üniversitesi Veteriner Fakültesi Dergisi, 73(3), 307-314. https://doi.org/10.33988/auvfd.1863540

AMA

1.Kalashnikova L, Bagal I, Murugina N, Senina R, Kalashnikov V, Meshcherov S. Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows. Ankara Univ Vet Fak Derg. 2026;73(3):307-314. doi:10.33988/auvfd.1863540

Chicago

Kalashnikova, Lyubov, Irina Bagal, Nina Murugina, Roman Senina, Vladimir Kalashnikov, and Shamil Meshcherov. 2026. “Influence of TLR2 Polymorphism on Innate Immune System Gene Expression in Holstein Cows”. Ankara Üniversitesi Veteriner Fakültesi Dergisi 73 (3): 307-14. https://doi.org/10.33988/auvfd.1863540.

EndNote

Kalashnikova L, Bagal I, Murugina N, Senina R, Kalashnikov V, Meshcherov S (July 1, 2026) Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows. Ankara Üniversitesi Veteriner Fakültesi Dergisi 73 3 307–314.

IEEE

[1]L. Kalashnikova, I. Bagal, N. Murugina, R. Senina, V. Kalashnikov, and S. Meshcherov, “Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows”, Ankara Univ Vet Fak Derg, vol. 73, no. 3, pp. 307–314, July 2026, doi: 10.33988/auvfd.1863540.

ISNAD

Kalashnikova, Lyubov - Bagal, Irina - Murugina, Nina - Senina, Roman - Kalashnikov, Vladimir - Meshcherov, Shamil. “Influence of TLR2 Polymorphism on Innate Immune System Gene Expression in Holstein Cows”. Ankara Üniversitesi Veteriner Fakültesi Dergisi 73/3 (July 1, 2026): 307-314. https://doi.org/10.33988/auvfd.1863540.

JAMA

1.Kalashnikova L, Bagal I, Murugina N, Senina R, Kalashnikov V, Meshcherov S. Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows. Ankara Univ Vet Fak Derg. 2026;73:307–314.

MLA

Kalashnikova, Lyubov, et al. “Influence of TLR2 Polymorphism on Innate Immune System Gene Expression in Holstein Cows”. Ankara Üniversitesi Veteriner Fakültesi Dergisi, vol. 73, no. 3, July 2026, pp. 307-14, doi:10.33988/auvfd.1863540.

Vancouver

1.Lyubov Kalashnikova, Irina Bagal, Nina Murugina, Roman Senina, Vladimir Kalashnikov, Shamil Meshcherov. Influence of TLR2 polymorphism on innate immune system gene expression in Holstein cows. Ankara Univ Vet Fak Derg. 2026 Jul. 1;73(3):307-14. doi:10.33988/auvfd.1863540