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Identification and characterization of two CD4 alleles in Microminipigs

Overview of attention for article published in BMC Veterinary Research, October 2016
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Title
Identification and characterization of two CD4 alleles in Microminipigs
Published in
BMC Veterinary Research, October 2016
DOI 10.1186/s12917-016-0856-8
Pubmed ID
Authors

Tatsuya Matsubara, Naohito Nishii, Satoshi Takashima, Masaki Takasu, Noriaki Imaeda, Kayo Aiki-Oshimo, Kazuaki Yamazoe, Michinori Kakisaka, Shin-nosuke Takeshima, Yoko Aida, Yoshie Kametani, Jerzy K. Kulski, Asako Ando, Hitoshi Kitagawa

Abstract

We previously identified two phenotypes of CD4+ cells with and without reactions to anti-pig CD4 monoclonal antibodies by flow cytometry in a herd of Microminipigs. In this study, we analyzed the coding sequences of CD4 and certified the expression of CD4 molecules in order to identify the genetic sequence variants responsible for the positive and negative PBMCs reactivity to anti-pig CD4 monoclonal antibodies. We identified two CD4 alleles, CD4.A and CD4.B, corresponding to antibody positive and negative, respectively, by nucleotide sequencing of PCR products using CD4 specific primer pairs. In comparison with the swine CD4 amino-acid sequence [GenBank: NP_001001908], CD4.A had seven amino-acid substitutions and CD4.B had 15 amino-acid substitutions. The amino-acid sequences within domain 1 of CD4.B were identical to the swine CD4.2 [GenBank: CAA46584] sequence that had been reported previously to be a modified CD4 molecule that had lost reactivity with an anti-pig CD4 antibody in NIH miniature pigs. Homozygous and heterozygous CD4.A and CD4.B alleles in the Microminipigs herd were characterised by using the RFLP technique with the restriction endonuclease, BseRI. The anti-pig CD4 antibody recognized pig PBMCs with CD4.AA and CD4.AB, but did not recognized those with CD4.BB. We transfected HeLa cells with the FLAG-tagged CD4.A or CD4.B vectors, and certified that transfected HeLa cells expressed FLAG in both vectors. The failure of cells to react with anti-CD4 antibodies in CD4.B pigs was associated to ten amino-acid substitutions in domain 1 and/or one amino-acid substitution in joining region 3 of CD4.B. We also found exon 8 was defective in some CD4.A and CD4.B resulting in the loss of the transmembrane domain, which implies that these CD4 proteins are secreted from helper T cells into the circulation. We identified that amino-acids substitutions of domain 1 in CD4.B gave rise to the failure of some CD4 expressing cells to react with particular anti-pig CD4 monoclonal antibodies. In addition, we developed a PCR-RFLP method that enabled us to simply identify the CD4 sequence variant and the positive and negative PBMCs reactivity to our anti-pig CD4 monoclonal antibodies without the need to use flow cytometric analysis.

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Mendeley readers

The data shown below were compiled from readership statistics for 3 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Unknown 3 100%

Demographic breakdown

Readers by professional status Count As %
Student > Bachelor 1 33%
Researcher 1 33%
Unknown 1 33%
Readers by discipline Count As %
Agricultural and Biological Sciences 1 33%
Medicine and Dentistry 1 33%
Unknown 1 33%

Attention Score in Context

This research output has an Altmetric Attention Score of 1. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 10 October 2016.
All research outputs
#6,447,705
of 8,497,409 outputs
Outputs from BMC Veterinary Research
#822
of 1,298 outputs
Outputs of similar age
#176,314
of 253,286 outputs
Outputs of similar age from BMC Veterinary Research
#35
of 63 outputs
Altmetric has tracked 8,497,409 research outputs across all sources so far. This one is in the 13th percentile – i.e., 13% of other outputs scored the same or lower than it.
So far Altmetric has tracked 1,298 research outputs from this source. They receive a mean Attention Score of 2.8. This one is in the 19th percentile – i.e., 19% of its peers scored the same or lower than it.
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We're also able to compare this research output to 63 others from the same source and published within six weeks on either side of this one. This one is in the 20th percentile – i.e., 20% of its contemporaries scored the same or lower than it.