Comparative molecular characterization of Forkhead box protein 3 (FoxP3) gene of swamp-type (Bubalus carabanensis) and riverine-type (Bubalus bubalis) water buffaloes

https://doi.org/10.1016/j.cimid.2019.01.022Get rights and content

Highlights

  • •

    We characterized FoxP3 gene in water buffaloes for the first time.

  • •

    The DNA sequence of the amplified FoxP3 gene in both swamp- and riverine-type water buffaloes contained 1296 nucleotides.

  • •

    There were a number of amino acid substitutions between FoxP3 of swamp- and riverine-type water buffaloes.

  • •

    Homology analysis revealed highest homology between riverine-type water buffalo and cattle.

  • •

    FoxP3 of swamp- and riverine-type water buffaloes had a high degree of homology to other animal species.

Abstract

FoxP3 is a forkhead family member that plays an important role in the development and function of a type of CD4 + T cell called T regulatory cells. Molecular characterization of FoxP3 gene in swamp- and riverine-type water buffaloes was conducted to determine its homology and compare it to the FoxP3 gene of other animal species (cattle, goat, sheep, horse, pig, cat, and dog), determine its unique characteristics in water buffaloes, and provide a reference for future studies to analyze its immunological function. FoxP3 nucleotide sequence of swamp- and riverine-type water buffaloes was 99% identical, whereas its protein translation revealed 97% homology. FoxP3 of swamp- and riverine-type water buffaloes were compared to FoxP3 of other animal species and revealed a high degree of homology which suggests that they may have the same biological properties. This study is the first report that describes the genetic characteristic of FoxP3 gene in water buffalo.

Introduction

Forkhead box protein 3 (FoxP3), an X‑linked gene (located in Xp11.23), is a forkhead family member that plays an important role in the development and function of a type of CD4+ T cell called T regulatory cells [1], thus called the “master regulator” for the Treg lineage [2], which is essential for maintaining immune tolerance to self.1 Induction of the Foxp3 gene in normal naïve T cells converts them to Treg-like cells with in vivo and in vitro suppressive function. This indicates that Foxp3 functions in controlling expression of critical suppression-mediating molecules [2]. Ziegler [3] stated that lack of Foxp3 leads to development of fatal autoimmune lymphoproliferative disease. Furthermore, retroviral-mediated introduction of Foxp3 can cause conventional CD4 + T cells to acquire a regulatory-like phenotype and be capable of suppressing immune responses both in vitro and in vivo.

The swamp-type (Bubalus carabanensis) and riverine-type (Bubalus bubalis) buffaloes are the known types of domesticated water buffalo. Swamp-type is usually gray, but there are also significant numbers which are pink, albino which are white, or piebald which are gray with white patches, whereas riverine-type is usually much darker but often have white markings on the head, legs or tail [4].

For the advancement of buffalo management and health, studies about its genetic immunity may be helpful. Genotype of Foxp3 in water buffaloes, was not yet characterized, is not well understood. Since Foxp3 is known to be responsible for the development of the regulatory T cells [1] which act to suppress immune responses, hence maintaining homeostasis and self-tolerance [5], it may also have specific significance regarding the immune responses of water buffaloes as compared to other species.

Foxp3 gene of water buffaloes has been molecularly characterized and compared to the Foxp3 gene sequence of other species. This is one of the primary steps in understanding the specific role of FoxP3 in water buffalo’s immune system. Thus, this study can be a foundation to know its involvement in disease susceptibility or resistance and probably to improve the gene selection for breeding.

Section snippets

Blood sample

A total of 23 blood samples were collected from the jugular and/or ear vein of randomly selected animals using syringes with heparin as anticoagulant. Blood samples from riverine-type buffaloes were collected from the Philippine Carabao Center (PCC) herd. Meanwhile, swamp-type buffaloes were collected from the municipalities of Nueva Ecija, Philippines. The buffy coat was then collected and transferred in a 1.5 ml centrifuge tube and washed with 1000 μL of NH4Cl through mixing in the vortex and

Sequence analysis of FoxP3

The amplicon size of FoxP3 for both types of water buffaloes was 1296 bp encoding for 432 amino acids is shown in Table 1.The Forkhead or the “winged-helix” superfamily domain, identified using the BLAST, was located from positon 337 up to position 409 (Fig. 1). The transcription factor of the Forkhead was identified as well, which was located in positions 335 to 429. All cysteine residues were all intact in the conserved region of the sequences and 3 N-linked glycosylation sites were observed

Discussion

The FoxP3 gene, as part of the Forkhead transcription factor complex or the Forkhead box genes, play an important role in the immune system including the correct performance of cellular and humoral immune response [9]. In this study, FoxP3 gene of swamp-type and riverine-type water buffaloes were amplified and characterized through DNA sequencing and phylogenetic analysis. These are the primary steps in understanding the specific role of FoxP3 in water buffalo’s immune system. FoxP3 of water

Conclusions

This study showed that both swamp- and riverine-type water buffaloes had high homology but showed differences in their FoxP3 amino acid sequences which could contribute to the claims of disease resistance in swamp buffalo. Phylogenetic and sequence analyses of different animal species (cattle, goat, sheep, horse, pig, dog and cat) were also included to compare with swamp-type and riverine-type water buffaloes. To further analyze the role of FoxP3 gene, particularly in water buffaloes, it is

Conflict of interest

The authors declare no conflicts of interest.

Acknowledgment

This research was supported by the PCC. Thanks are extended to the PCC management and the rest of the staff of the Biosafety and Environment Section in providing the needed materials and technical assistance for this research.

References (26)

  • B. Lemcke

    Water Buffalo Farming in Southern Australia, Northern Territory Government

    (2006)
  • J. Grant et al.

    Validated protocol for foxp3 reveals increased expression in Type 1 diabetes patients

    Cytometry B Clin. Cytom.

    (2009)
  • S. Tajima et al.

    Complete Bovine Leukemia Virus (BLV) provirus is conserved in BLV-infected cattle throughout the course of B-cell lymphosarcoma development

    J. Virol.

    (1998)
    • 1

    Both authors contributed equally to this work.

    View full text
    © 2019 Elsevier Ltd. All rights reserved.