Mts1 (S100A4) and Its Peptide Demonstrate Cytotoxic Activity in Complex with Tag7 (PGLYRP1) Peptide.

Receptors of cytokines are major regulators of the immune response. In this work, we have discovered two new ligands that can activate the TNFR1 (tumor necrosis factor receptor 1) receptor. Earlier, we found that the peptide of the Tag (PGLYRP1) protein designated 17.1 can interact with the TNFR1 receptor. Here, we have found that the Mts1 (S100A4) protein interacts with this peptide with a high affinity (Kd = 1.28 × 10-8 M), and that this complex is cytotoxic to cancer cells that have the TNFR1 receptor on their surface. This complex induces both apoptosis and necroptosis in cancer cells with the involvement of mitochondria and lysosomes in cell death signal transduction. Moreover, we have succeeded in locating the Mts1 fragment that is responsible for protein-peptide interaction, which highly specifically interacts with the Tag7 protein (Kd = 2.96 nM). The isolated Mts1 peptide M7 also forms a complex with 17.1, and this peptide-peptide complex also induces the TNFR1 receptor-dependent cell death. Molecular docking and molecular dynamics experiments show the amino acids involved in peptide binding and that may be used for peptidomimetics' development. Thus, two new cytotoxic complexes were created that were able to induce the death of tumor cells via the TNFR1 receptor. These results may be used in therapy for both cancer and autoimmune diseases.

The 12-Membered TNFR1 Peptide, as Well as the 16-Membered and 6-Membered TNF Peptides, Regulate TNFR1-Dependent Cytotoxic Activity of TNF.

Understanding the exact mechanisms of the activation of proinflammatory immune response receptors is very important for the targeted regulation of their functioning. In this work, we were able to identify the sites of the molecules in the proinflammatory cytokine TNF (tumor necrosis factor) and its TNFR1 (tumor necrosis factor receptor 1), which are necessary for the two-stage cytotoxic signal transduction required for tumor cell killing. A 12-membered TNFR1 peptide was identified and synthesized, interacting with the ligands of this receptor protein's TNF and Tag7 and blocking their binding to the receptor. Two TNF cytokine peptides interacting with different sites of TNFR1 receptors were identified and synthesized. It has been demonstrated that the long 16-membered TNF peptide interferes with the binding of TNFR1 ligands to this receptor, and the short 6-membered peptide interacts with the receptor site necessary for the transmission of a cytotoxic signal into the cell after the ligands' interaction with the binding site. This study may help in the development of therapeutic approaches to regulate the activity of the cytokine TNF.

Thymopentin plays a key role in restoring the function of macrophages to alleviate the sepsis process.

Immune dysfunction is one of the leading causes of death of sepsis. How to regulate host immune functions to improve prognoses of septic patients has always been a clinical focus. Here we elaborate on the efficacy and potential mechanism of a classical drug, thymopentin (TP5). TP5 could decrease peritoneal bacterial load, and reduce inflammatory cytokine levels both in the peritoneal lavage fluid (PLF) and serum, alleviate pathological injuries in tissue and organ, coaxed by cecal ligation and perforation (CLP) in mice, ultimately improve the prognosis of septic mice. Regarding the mechanism, using RNA-seq and flow cytometry, we found that TP5 induced peptidoglycan recognition protein 1 (PGLYRP1) expression, increased phagocytosis and restored TNF-α expression of small peritoneal macrophage (SPM) in the septic mice. This may be increased SPM's ability to clear peritoneal bacteria, thereby attenuates the inflammatory response both in the peritoneal cavity and the serum. It was shown that TP5 plays a key role in restoring the function of peritoneal macrophages to alleviate the sepsis process. We reckon that this is closely relevant to SPM phagocytosis, which might involve increased PGLYRP1 expression and restored TNF-α secretion.

Association of salivary TREM-1 and PGLYRP1 inflammatory markers with non-communicable diseases.

AIM: Triggering receptor expressed on myeloid cells 1 (TREM-1) and peptidoglycan recognition protein 1 (PGLYRP1) are elevated in biofluids in the presence of various inflammatory conditions. This cross-sectional study aimed to evaluate the effect of age, sex, smoking and different oral and systemic non-communicable diseases on the levels of TREM-1 and PGLYRP1 in saliva. MATERIALS AND METHODS: In total, 445 individuals (mean age 48.7 ± 16.9 years, female:male 51%:49%) were included. All provided self-reported information on smoking and systemic diseases and whole stimulated saliva. Periodontal and cariological parameters were recorded. Salivary levels of TREM-1, PGLYRP1 and total protein were measured using commercially available assays. RESULTS: Salivary TREM-1 levels were significantly higher in stages III-IV periodontitis compared to other periodontal diagnoses (p < .05). Smoking, bleeding on probing (BOP), percentage of pockets ≥4 mm and the number of manifest caries were associated with TREM-1 (p < .05), while sex, BOP, number of manifest caries and muscle and joint diseases were associated with PGLYRP1 (p < .05). CONCLUSIONS: Salivary TREM-1 is associated with periodontitis and caries, while PGLYRP1 is associated with gingival inflammation and caries. Additionally, TREM-1 levels are modified by smoking, while PGLYRP1 is modified by sex and muscle and joint diseases. TREM-1 and PGLYRP1 in saliva could serve as potential biomarkers for detecting and monitoring non-communicable diseases.

N-Terminal Peptide of PGLYRP1/Tag7 Is a Novel Ligand for TREM-1 Receptor.

An investigation of innate immunity receptors sheds light on the mechanisms of inflammation and associated immune reactions. One of the key immune regulators is the TREM-1 receptor, which is involved in both inflammation and antitumor immune response. In this article, we have obtained a new ligand for the TREM-1 receptor. The peptide, named N3, is a part of the innate immune protein PGLYRP1/Tag7. It is responsible for activating the TREM-1 signaling pathway. Here, we have demonstrated that the N3 peptide acts like other TREM-1 receptor ligands: its binding results in a mild inflammation response and appearance of cytotoxic lymphocytes. We have shown that cytotoxic populations of lymphocytes in N3 peptide-treated PBMCs are similar to those treated with Tag7 or Hsp70. We also determined the part of the N3 peptide responsible for binding to TREM-1. The resulting peptide (N9) consists of nine amino acids and can be considered as a potential peptide that blocks TREM-1 signaling.

TNF upregulates peptidoglycan recognition protein 1 in esophageal cancer cells to clear the path to its signaling: Making the "enemy" a friend.

TNF, CCN1, and peptidoglycan recognition protein 1 (PGLYRP1) are often found together in the inflammatory tissue. While TNF and CCN1 promote tissue regeneration, PGLYRP1 protects it from bacterial infection. In fibroblasts, CCN1 was reported to support TNF in apoptosis induction while PGLYRP1 was found to compete with TNF for binding to TNFR1. When PGLYRP1 binds to TNFR1 by itself, it silences the receptor, but if HSP70 joins them, it leads to cell death. In cancer cells, however, CCN1 was found to antagonize TNF signaling by increasing the extracellular pool of TNFR1. In this study, we assessed their relationship in the esophageal cancer cells and found a more complex liaison among them. At first, TNF highly upregulated PGLYRP1 expression but downregulated CCN1. Secondly, PGLYRP1 bound TNFR1 and HSP70 both intracellularly and extracellularly, but TNF only promoted their extracellular interaction. Lastly, the knockdown of PGLYRP1 impaired TNF signaling. Taken together, this study shows that CCN1 interrupts TNF signaling by increasing the extracellular TNFR1 species while TNF fights back by upregulating PGLYRP1 to absorb them.

Peri-Implant Surgical Treatment Downregulates the Expression of sTREM-1 and MMP-8 in Patients with Peri-Implantitis: A Prospective Study.

sTREM-1 and its ligand PGLYRP1 play an essential role in the inflammatory process around teeth and implants. In this study, we aimed to evaluate the impact of peri-implant treatment on the salivary levels of the sTREM-1/PGLYRP-1/MMP-8 axis after 3 months. A total of 42 participants (with a mean age of 61 years old ± 7.3) were enrolled in this longitudinal study, 24 having peri-implant mucositis (MU) and 18 having peri-implantitis (PI). Clinical peri-implant parameters, such as probing pocket depth (PPD), % of plaque, and bleeding on probing (BOP), and the whole unstimulated saliva samples were evaluated at baseline and 3 months after treatment. The MU group received nonsurgical peri-implant treatment, while the PI group received open-flap procedures. The levels of sTREM-1, PGLYRP-1, MMP-8, and TIMP-1 were analyzed using enzyme-linked immunosorbent assays. BOP, plaque levels, and PPD significantly reduced after treatment in both groups. A significant decrease in the salivary levels of sTREM-1, MMP-8, and TIMP-1 in the PI group and PGLYRP1 and TIMP-1 in the MU group were observed. Salivary levels of sTREM-1 were significantly reduced in patients with PI but not with MU. Additionally, peri-implant treatment had a significantly higher impact on MMP-8 reduction in patients with PI than in those with MU.

Association of peptidoglycan recognition protein 1 to post-myocardial infarction and periodontal inflammation: A subgroup report from the PAROKRANK (Periodontal Disease and the Relation to Myocardial Infarction) study.

BACKGROUND: Peptidoglycan recognition protein 1 (PGLYRP1) is an antimicrobial and proinflammatory innate immunity protein activated during infections. We aimed to investigate whether PGYLRP1 and associated molecules of the immune response in saliva is a cumulative outcome result of both MI and periodontal inflammation. METHODS AND RESULTS: Two hundred patients with MI and another 200 matched non-MI controls were included. A full-mouthexamination was conducted to assess periodontal inflammation and collection of stimulated saliva was performed 6 to 10 weeks after the first MI. PGLYRP1, triggering receptor expressed on myeloid cells 1 (TREM-1), interleukin-1 beta (IL-1ß) were analyzed by ELISA. Matrix metalloproteinase (MMP)-8 levels were determined by IFMA. Compared to controls, MI patients showed higher salivary PGLYRP1, but not TREM-1 levels. The difference in PGLYRP1 levels remained after adjustment for covariates. In MI patients, the PGLYRP1 levels positively correlated with BOP and PPD 4 to 5 mm. Among non-MI subjects, the levels of PGLYRP1 correlated positively and significantly with BOP and total PPD. Salivary PGLYRP1 concentrations also showed strong positive correlations with levels of TREM-1, IL-1ß and MMP-8. In multivariate linear regression analysis, in MI patients, BOP and former smokingstatus displayed an association with salivary PGLYRP1 concentration. CONCLUSION: MI patients showed higher salivary PGLYRP1 levels than healthy controls, also after adjusting for smoking, sex, age and periodontal health status. Salivary levels of PGLYRP1 may reflect the overall inflammatory burden to chronic bacterial exposure, possibly underpinning the observed associations between periodontitis and exposure with MI.

Mechanisms of Action of the PGLYRP1/Tag7 Protein in Innate and Acquired Immunity.

One of the promising fields of modern molecular biology is the search for new proteins that regulate the various stages of the immune response and the investigation of the molecular mechanisms of action of these proteins. Such proteins include the multifunctional protein PGLYRP1/Tag7, belonging to the PGRP-S protein family, whose gene was discovered in mice at the Institute of Gene Biology, Russian Academy of Sciences, in 1996. PGLYRP1/Tag7 is classified as a protein of innate immunity; however, it can also participate in the regulation of acquired immunity mechanisms. In this paper, we consider the involvement of PGLYRP1/Tag7 in the triggering of antimicrobial defense mechanisms and formation of subsets of cytotoxic lymphocytes that kill tumor cells. The paper emphasizes that the multifaceted functional activity of Tag7 in the immune response has to do with its ability to interact with various proteins to form stable protein complexes. Hsp70-associated Tag7 can induce the death of tumor cells carrying the TNFR1 receptor. Tag7, associated with the Mts1 (S100A4) protein, can stimulate the migration of innate and adaptive immune cytotoxic lymphocytes to a lesion site. Involvement of Tag7 in the regulation of immunological processes suggests that it may be considered as a promising agent in cancer therapy. These properties of Tag7 were used to develop autologous vaccines that have passed the first and second phases of clinical trials in patients with end-stage melanoma and renal cancer. The C-terminal peptide of Tag7, isolated by limited proteolysis, was shown to protect the cartilage and bone tissue of the ankle joint in mice with induced autoimmune arthritis and may be a promising drug for suppressing the development of inflammatory processes.

Survey of Candidate Single-Nucleotide Polymorphisms in SLC11A1, TLR4, NOD2, PGLYRP1, and IFNγ in Ankole Longhorn Cattle in Central Region of Uganda to Determine Their Role in Mycobacterium avium Subspecies paratuberculosis Infection Outcome.

Mycobacterium avium ssp. paratuberculosis (MAP) is the cause of Johne's disease (JD) in a wide range of domestic and wild ruminants. Single-nucleotide polymorphisms (SNPs) in several genes including solute-like carrier 11A1 (SLC11A1), interferon gamma (IFNγ), Toll-like receptor 4 (TLR4), nucleotide-binding oligomerization domain 2 gene (NOD2), and bovine peptidoglycan recognition protein 1 (PGLYRP1) have been implicated in influencing the infection outcome of MAP in cattle. We have carried out a survey in a population of Ankole cattle from three districts in the central region of Uganda including Isingiro, Lyantonde, and Rakai to determine the role played by several SNPs on the above genes in the infection outcome of local cattle in Uganda. Nine hundred fifty-five heads of cattle obtained from 93 herds were tested using ELISA. Thirty-five ELISA-positive cattle and 35 negative herd mates from a total of 955 cattle tested for MAP were genotyped using iPLEX MassARRAY genotyping systems to detect the presence of a total of 13 SNPS in five different genes (SLC11A1, IFNγ, TLR4, NOD2, and PGLYRP1). The cow-level prevalence of MAP infection in Ankole Longhorn cattle in the three districts was 3.98% (35/955), while the herd-level prevalence was 27.9% and within-herd prevalence was 12 ± 1.5% (95% CI = 9.1-14.8%). The genotypes and allele frequencies of the MAP-positive cattle were compared with those of their ELISA-negative herd mates to determine the significance of the polymorphisms. The results showed that SNPs rs109915208, rs110514940, and rs110905610 on SLC11A1, c.480G>A and c.625C>A on PGLYRP1, and c.2021C>T on TLR4 were monomorphic in both seropositive and seronegative cattle and therefore had no influence on the infection outcome. The remaining SNPs studied in the five genes [SLC11A1: rs109614179; TLR4: rs29017188 (c.226G>C), c.2021C>T; NOD2: rs110536091, rs111009394; PGLYRP1: c.102G>C, c.480G>A, c.625C>A; IFNγ: rs110853455] were polymorphic, but their allele and genotype frequencies did not show any significant difference between the seropositive and seronegative cattle. No significant difference was observed for any haplotype at the gene level.

Novel ligands and modulators of triggering receptor expressed on myeloid cells receptor family: 2015-2020 updates.

Introduction: Triggering receptors expressed on myeloid cells (TREMs) are inflammatory amplifiers with defined pathophysiological role in various infectious diseases, acute and chronic aseptic inflammations, and a variety of cancers, depicting TREMs as prominent therapeutic targets.Areas covered: Herein, updates from 2015 to 2020 are discussed to divulge the TREM ligands, as well as their peptide blockers, claimed to modulate their expression. The article also presents different strategies employed during the last five years to block interactions between TREMs and their ligands to treat various disease conditions by modulating their expression and activity.Expert opinion: There has been significant progress in the discovery of novel ligands and modulators of TREMs in the last five years that mainly revolved around the function of TREM molecules. A few peptides showed encouraging results to modulate the expression and activity of TREMs in preclinical studies, and these peptides are currently under clinical investigation. Based on the findings so far in several careful studies, we expect novel therapeutics in the near future which could have the ability to treat various disease conditions associated with TREM expression.

Cytokines TNFα, IFNγ and IL-2 Are Responsible for Signal Transmission from the Innate Immunity Protein Tag7 (PGLYRP1) to Cytotoxic Effector Lymphocytes.

Studies on the mechanisms of activation of cytotoxic lymphocyte subpopulations are an important research direction in modern immunology. This study provides a detailed analysis of the effect of Tag7 (PGRP-S, PGLYRP1) on the development of lymphocyte subpopulations cytotoxic against MHC-negative tumor cells in a pool of peripheral blood mononuclear cells (PBMCs). The results show that Tag7 can bind to the TREM-1 receptor on the surfaces of monocytes, thereby triggering the expression of mRNA TNFα and IFNγ. The appearance of these cytokines in conditioned medium leads to IL-2 cytokine secretion by CD3+CD4+ lymphocytes. In turn, IL-2 facilitates unspecific activation of three cytotoxic cell subpopulations in the PBMC pool: NK (CD16+CD56+), CD3+CD4+ and CD3+CD8+. These subpopulations appear after a certain period of incubation with Tag7 and show toxicity against tumor cells.

Regulation of Salivary Peptidoglycan Recognition Protein 1 in Adolescents.

INTRODUCTION: Peptidoglycan recognition protein 1 (PGLYRP1), a member of peptidoglycan recognition proteins, is known to be involved in the proinflammatory response toward bacterial infections. Recently, PGLYRP1 was identified as a ligand for triggering receptor expressed on myeloid cells 1 (TREM-1). Although PGLYRP1 is involved in immune and inflammatory responses, its levels in initial stages of periodontal disease in adolescents are currently unknown. OBJECTIVES: We aimed to investigate salivary levels of PGLYRP1 and its correlation with TREM-1, polymorphonuclear leukocyte elastase (PMN elastase), and an active matrix metalloproteinase 8 (aMMP-8) in adolescents. METHODS: Whole saliva samples (n = 537) were collected from 15- to 16-y-old adolescents at Kotka Health Center, Finland, prior to periodontal examination, including measurement of periodontal pocket depth (PPD), visible plaque index (VPI), and bleeding on probing (BOP). Adolescents, clustered as periodontally healthy, gingivitis, or subclinical periodontitis, were tested for salivary levels of TREM-1, PGLYRP1, and PMN elastase by enzyme-linked immunosorbent assay and aMMP-8 by a time-resolved immunofluorometric assay (IFMA). RESULTS: Salivary levels of PGLYRP1 and aMMP-8 were significantly higher in adolescents with subclinical periodontitis and gingivitis compared to individuals with healthy periodontium. TREM-1 and PMN elastase levels were higher in adolescents with subclinical periodontitis compared to healthy individuals but did not reach significance. PGLYRP1 correlated positively with BOP, PPD, VPI, aMMP-8, and TREM-1. CONCLUSIONS: Elevated PGLYRP1 levels in adolescents with gingivitis and subclinical periodontitis and its positive correlation with TREM-1 and aMMP-8 may indicate an association of PGLYRP1 with initial stages of periodontal disease. Sex and poor oral hygiene but not smoking are also associated with higher levels of PGLYRP1. However, PGLYRP1 has a lower discriminating capacity and is therefore a less reliable marker alone in the diagnosis of initial stages of periodontal disease in adolescents. KNOWLEDGE TRANSFER STATEMENT: PGLYRP1, a member of peptidoglycan recognition proteins, is a ligand for TREM-1. Elevated PGLYRP1 levels in adolescents with gingivitis and subclinical periodontitis and its positive correlation with TREM-1 and aMMP-8 may indicate an association of PGLYRP1 with initial stages of periodontal disease. However, it has a lower discriminating capacity and is therefore a less reliable marker alone in the diagnosis of periodontal disease in adolescents.

The modulation of the TREM-1/PGLYRP1/MMP-8 axis in peri-implant diseases.

OBJECTIVES: The aim of this study is to investigate the expression of sTREM-1 and its ligand PGLYRP1, as well as the expression of MMP-8 and its inhibitor TIMP-1, in peri-implant diseases. As a secondary aim, we analyzed the influence of the concomitant existence of periodontitis in the expression of these biomarkers. MATERIALS AND METHODS: This study included 77 patients (29 males and 48 females; mean age 55.0 ± 11.5), 18 having gingivitis, 16 having periodontitis, 20 having mucositis, and 23 having peri-implantitis. Patients were clinically examined, and unstimulated whole saliva was collected. sTREM-1, PGLYRP1, MMP-8, TIMP-1, and MMP-8/TIMP1 ratio were determined by ELISA. RESULTS: The periodontitis group presented higher probing depth (PD) mean, and higher clinical attachment loss, compared with the other groups. The peri-implantitis group presented higher PD mean in implants compared to the mucositis group. Patients with PD ≥ 6 mm showed significantly higher levels of PGLYRP1, MMP-8, and MMP-8/TIMP-1 ratio than patients with PD < 6 mm. When all four markers were assessed, there were no significant differences between mucositis and peri-implantitis groups. Concomitant periodontitis resulted in higher significant levels of MMP-8 in patients with peri-implant disease. CONCLUSION: We did not observe significant differences in the levels of the sTREM-1/PGLYRP1/MMP-8 axis between patients with periodontal and peri-implant diseases, suggesting that these markers are also involved in the inflammatory process around implants. Besides, the presence of periodontitis may affect the levels of MMP-8 in patients with peri-implant disease. CLINICAL RELEVANCE: The sTREM-1/PGLYRP1/MMP-8 axis could be useful as potent markers in periodontal and peri-implant diseases.

The effect of smoking on gingival crevicular fluid peptidoglycan recognition protein-1 level following initial periodontal therapy in chronic periodontitis.

OBJECTIVE: To determine the effect of smoking on peptidoglycan recognition protein (PGRP)-1 levels in gingival crevicular fluid (GCF) in smokers and non-smokers with chronic periodontitis (CP) following initial periodontal therapy. SUBJECTS AND METHODS: A total of 40 patients with CP (20 smokers (S + CP) and 20 non-smokers (S-CP)) and 40 subjects with healthy periodontium (20 smokers (S + PH) and 20 non-smokers (S-PH)), comprising 80 subjects, were included in this study. Baseline GCF samples were obtained from all subjects, and clinical periodontal measurements were recorded. In patients who had received initial periodontal therapy, GCF samples were obtained and all clinical periodontal measurements were recorded again during the 6th-8th weeks. GCF PGRP-1 levels were analyzed by enzyme-linked immunosorbent assay. RESULTS: At baseline, GCF PGRP-1 levels were significantly higher in both groups with CP than in both groups with healthy periodontium, whereas these levels were significantly lower in S + CP than in S-CP. GCF PGRP-1 levels decreased significantly in both CP groups after periodontal therapy, and this reduction was significantly greater in non-smokers than in smokers. CONCLUSION: Smoking might have a suppressive effect on GCF PGRP-1 levels in CP. Initial periodontal therapy is effective in decreasing GCF PGRP-1 levels in both smokers and non-smokers with CP.

Human Short Peptidoglycan Recognition Protein PGLYRP1/Tag-7/PGRP-S Inhibits Listeria monocytogenes Intracellular Survival in Macrophages.

PGLYRP1/Tag-7/PGRP-S is one of mammalian peptidoglycan recognition proteins (PGRPs). Here, we demonstrate that human recombinant PGLYRP1/Tag-7/PGRP-S potentiates the response of murine macrophage-like ANA-1 cells and human macrophages to facultative intracellular pathogen Listeria monocytogenes. PGLYRP1/Tag-7/PGRP-S binds to the surface of L. monocytogenes and other bacterial cells but has no effect on their growth in culture. While PGLYRP1/Tag-7/PGRP-S treatment modestly enhanced phagocytosis of bacteria by ANA-1 cells, the intracellular survival of PGLYRP1/Tag-7/PGRP-S treated L. monocytogenes was strongly inhibited 2 h after internalization. PGLYRP1/Tag-7/PGRP-S treatment of bacteria boosted oxidative burst induction and increased the level of proinflammatory cytokine IL-6 produced by ANA-1, however, these effects happened too late to be responsible for decreased intracellular survival of bacteria. Our results thus suggest that PGLYRP1/Tag-7/PGRP-S acts as a molecular sensor for detection of L. monocytogenes infection of mammalian cells that leads to increased killing through a mechanism(s) that remains to be defined.

Polymorphism in PGLYRP-1 gene by PCR-RFLP and its association with somatic cell score in Chinese Holstein.

Bovine peptidoglycan recognition protein 1 (PGLYRP-1), an important pattern recognition molecule (PRM) of the innate immune system, is an effector molecule in killing different microorganisms directly. To investigate whether the PGLYRP-1 gene was associated with mastitis and milk production traits in dairy cattle, the polymorphism of this gene was analyzed by PCR-RFLP in a population of 524 Chinese Holstein. A total of ten single nucleotide polymorphism (SNP) loci were identified. The association analysis of single SNP locus showed that T-35A, T-12G and G+102C were significantly associated (P<0.05) with somatic cell score (SCS), while G+102C and G+649C were significantly associated (P<0.05) with 305-day milk yield. Association analysis between combined haplotypes and SCS, milk production traits indicated that H3H3 was associated with the lower SCS (P<0.01), and H2H2 was associated with the lower 305-day milk yield (P<0.01). These findings demonstrated that polymorphisms in PGLYRP-1 gene associated with mastitis resistance and 305-day milk yield, and the H3H3 would provide a useful genetic marker of combined haplotypes for mastitis resistance selection and breeding in Chinese Holstein.