A comprehensive analysis of immunoglobulin heavy chain genes in the Bactrian camel (<i>Camelus bactrianus</i>)

doi:10.15302/J-FASE-2015056

Front. Agr. Sci. Eng.

2015, Vol. 2

Issue (3) : 249-259 https://doi.org/10.15302/J-FASE-2015056

RESEARCH ARTICLE

A comprehensive analysis of immunoglobulin heavy chain genes in the Bactrian camel (Camelus bactrianus)

Zuoxiang LIANG^1,²,Tao WANG²,Yi SUN³,Wenlong YANG¹,Zhihong LIU⁴,Jing FEI²,Ying GUO²,Qingwei MA⁵,Qingjie PAN^1,^*(

),Liming REN^2,^*(

)

1. Key Laboratory of Animal Reproduction and Germplasm Enhancement in Universities of Shandong, College of Animal Science and Technology, Qingdao Agricultural University, Qingdao 266109, China
2. State Key Laboratory of Agrobiotechnology, China Agricultural University, Beijing 100193, China
3. Laboratory of Animal Molecular Genetics, College of Animal Science and Veterinary Medicine, Shandong Agricultural University, Taian 271018, China
4. Key Laboratory of Animal Genetics, Breeding and Reproduction, Inner Mongolia Agricultural University, Hohhot 010018, China
5. Bioyong Technology Inc., Beijing 100080, China

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Abstract

Heavy chain only antibodies (HCAbs) represent a rare type of antibody that is devoid of light chains and the CH1 domain that have been reported in cartilaginous fish and camelids. By analyzing transcript data and genome sequences, we conducted a comprehensive analysis of Bactrian camel immunoglobulin heavy chain genes. Based on the transcript data, one μ gene, five γ genes, one α gene and one ε gene were found. Additionally, the variable region of HCAbs (VHH) and the conventional antibodies (VH) sequences associated with the γ3, γ1a/b and μ genes were amplified. Based on these genome sequences, seven DH, six JH, μ, γ2a, γ2c, α, and ε genes and a portion of a γ3 gene were observed. Different Kozak sequences within different VH families were found in our analysis, and the variability index differed between the VHH3 and VH3 families. Phylogenetic analysis of the constant regions of the camelid immunoglobulin genes indicates that these genes appeared before the evolutionary divergence of Bactrian camels and dromedaries.

Keywords Bactrian camel heavy-chain antibodies VHH γ3

Corresponding Author(s): Qingjie PAN,Liming REN

Just Accepted Date: 28 May 2015 Online First Date: 18 June 2015 Issue Date: 10 November 2015

Cite this article:

Zuoxiang LIANG,Tao WANG,Yi SUN, et al. A comprehensive analysis of immunoglobulin heavy chain genes in the Bactrian camel (Camelus bactrianus)[J]. Front. Agr. Sci. Eng. , 2015, 2(3): 249-259.

URL:

https://academic.hep.com.cn/fase/EN/10.15302/J-FASE-2015056
https://academic.hep.com.cn/fase/EN/Y2015/V2/I3/249

Fig.1 Five VH amino acid sequences were found among 192 HCAbs using a cDNA database. The numbering used refers to the position in the locus in the 5′ to 3′ direction. V region sequences were analyzed using the multiple sequence alignment MEGA 5.0 software. CDR (reseda shading) and FR (black and white box) regions of the V genes are determined according to IMGT nomenclature and are indicated at the top. Purple and wathet shaded amino acid sequences represent hallmarks for the VHH and VH genes, respectively.

Fig.2 Five conserved bases located at the 5' UTR end of the heavy chain variable domains of IgM, IgG1a/b and IgG3. (a) Variable region sequences of IgHM; (b) variable region sequences of IgHG1a/b; (c) variable region sequences of IgHG3.

Fig.3 Phylogenetic analysis of Bactrian camel VH families. Nurse shark VH1 was used as the outgroup. The scale bar shows the genetic distance, and the credibility value is noted beside each node. Shaded boxes represent the clustering of sequences into clans. With the exception of the Bactrian camel sequences, all VH sequences were obtained from IMGT.

Fig.4 Comparison of the variability of the VH3 sequences derived from IgM or IgG1a/b with IgG3 VHH3 sequences. (a) Amino acid sequence variability plots for the cDNA sequences of VHH3s derived from IgG3 (upper left panel) and VH3s derived from IgG1a/b (lower panels). The variability index was calculated from FR1 to FR3 as described in the results section. Grey and open bars represent amino acids in the CDR and FR regions, respectively, and dark gray bars represent residues 27−30 in the VHH, which we postulate is the extra hypervariable region that has been reported in the literature using Kabat nomenclature. CDR1 and CDR2, as defined by IMGT nomenclature, are shaded and placed between the dotted lines; (b) amino acid sequence variability plots comparing the sequences predicted from cDNA sequences of VH3s derived from IgM (upper right panel) and VH3s derived from IgG1a/b (lower panels).

Fig.5 Frequency distribution histograms of heavy chain CDR3 lengths of the VH3 and VHH3 families. CDRs are defined according to the IMGT nomenclature.

Fig.6 WebLogo plots presenting sequence differences between the VHH3 domains derived from IgG3 and the VH3 domains derived from IgM and IgG1a/b. (a) VH3 sequences derived from IgM; (b) VH3 sequences derived from IgG1a/b; (c) VHH3 sequences derived from IgG3.

Fig.7 Bactrian camel immunoglobulins: structure and gene organization. (a) Details of the Bactrian camel IgH gene organization and the domain structure of each immunoglobulin heavy chain gene constant region. (b) Nucleotide/amino acid sequences of Bactrian camel DH and JH genes.

Fig.8 Phylogenetic analysis of IgHC genes in mammals. Nurse shark IgM was used as an outgroup. The scale bar shows the genetic distance. The tree was constructed using amino acid sequences. Bactrian camel Ig classes are boxed.

Fig.9 The 3′ CH1 boundary and hinge region analysis in γ genes. The point mutation destroying the conventional splicing signal is shaded in gray. The amino acids encoded by the hinge region are listed at the right, and the size of the hinge (number of amino acids) is given within brackets. Bactrian camel sequences are in bold font. The γ1a and γ1b genes encode conventional Abs, whereas the γ2a, γ2b, γ2c and γ3 genes encode HCAbs.

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