Expression of a Dromedary Heavy Chain-Only Antibody and B Cell Development in the Mouse

Zou, Xiangang; Smith, Jennifer A.; Nguyen, Viet Khong; Ren, Liming; Luyten, Kattie; Brüggemann, Marianne

doi:10.4049/jimmunol.175.6.3769

Cited by 28 publications

(21 citation statements)

References 61 publications

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“…Moreover, we see no LC rearrangement and conclude that LC are not required for further B cell differentiation. The difference in our results and those of Zou et al (13) may be explained by the level of expression of the locus (and, thus, signaling) because of the LCR on our constructs. Our results confirm that truncated HCAb lacking CH1 (24) or VH and CH1 (36) cannot associate with SLCs and fail to activate gene rearrangement.…”

Section: Discussioncontrasting

confidence: 56%

“…IgG1 expression from the pro-B cell stage onwards, was shown to substitute for IgM in Rag2 Ϫ/Ϫ B cell development (35). Recently, it also was shown that a prerearranged camelid IgG2a partially rescues B cell development in one transgenic line in a MT (and a C⌬ Ϫ/Ϫ ) background (13). In our case, IgM or IgG lacking CH1 rescue B cell development in 10 of 10 independent lines.…”

Section: Discussionmentioning

confidence: 58%

“…Murine NSO myeloma cells can express a rearranged camelid VHH-␥2a gene (12) and, recently, the same gene was expressed in transgenic mice (13). Here, we describe transgenic mice containing various nonrearranged chimeric HCAb loci and show they rearrange properly, result in allelic exclusion, efficiently rescue B cell development, and undergo class switch recombination and affinity maturation.…”

Section: Onventional Antibodies Contain Two Heavy and Light Chains mentioning

confidence: 99%

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Generation of heavy-chain-only antibodies in mice

Janssens¹,

Dekker²,

Hendriks

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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We have generated transgenic mice containing hybrid llama͞ human antibody loci that contain two llama variable regions and the human D, J, and C and͞or C␥ constant regions. Such loci rearrange productively and rescue B cell development efficiently without LC rearrangement. Heavy-chain-only antibodies (HCAb) are expressed at high levels, provided that the CH1 domain is deleted from the constant regions. HCAb production does not require an IgM stage for effective pre-B cell signaling. Antigenspecific heavy-chain-only IgM or IgGs are produced upon immunization. The IgG is dimeric, whereas IgM is multimeric. The chimeric HCAb loci are subject to allelic exclusion, but several copies of the transgenic locus can be rearranged and expressed successfully on the same allele in the same cell. Such cells are not subject to negative selection. The mice produce a full antibody repertoire and provide a previously undescribed avenue to produce specific human HCAb in the future.immunoglobulin rearrangement ͉ transgenic C onventional antibodies contain two heavy and light chains (LC) coded for by heavy and LC loci. B cell development and antibody production starts in the bone marrow (BM) by heavy chain (HC) VDJ recombination and expression of IgM associated with a surrogate LC on the cell surface. In a second round of recombination, one of the LC rearranges in pre-B cells. If successful, the B cells undergo selection, affinity maturation, and switching to different HC constant regions to result in B cells, which express tetrameric antibodies of different isotypes (IgA, IgG, and IgE). Normally absence of HC or LC expression leads to arrest of B cell development. However, some species produce HC-only antibodies (HCAb) as part of their normal B cell development and repertoire. The best-known HCAb (i.e., no LC) are IgG2 and IgG3 in camelids (1). They undergo antigen-mediated selection and affinity maturation, and their variable domains are subject to somatic hypermutation (2, 3). HCAb are thought to recognize unusual epitopes, such as clefts on the antigen surface (4). The first domain of the constant region, CH1, is spliced out because of the loss of a consensus splice signal (5, 6). CH1 exon loss also has been described in other mammals, albeit associated with disease, e.g., in mouse myelomas (7) and human HC disease (HCD) (8-10).Camelid HCAbs contain a complete VDJ region. Its size, stability, specificity, and solubility have generated considerable biotechnological interest. The antigen-binding site, a single-variable domain (VHH), resembles VH of conventional Abs. However, differences in FR2 and CDR3 prevent VHH to pair with a variable LC, whereas hydrophilic amino acids provide solubility (11). HCAb of the IgM class have not been found in camelids, suggesting that the IgM ϩ stage of HCAb formation is very transient and͞or circumvented.Murine NSO myeloma cells can express a rearranged camelid VHH-␥2a gene (12) and, recently, the same gene was expressed in transgenic mice (13). Here, we describe transgenic mice containing various...

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Section: Discussioncontrasting

confidence: 56%

Section: Discussionmentioning

confidence: 58%

Section: Onventional Antibodies Contain Two Heavy and Light Chains mentioning

confidence: 99%

See 1 more Smart Citation

Generation of heavy-chain-only antibodies in mice

Janssens¹,

Dekker²,

Hendriks

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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“…Previous studies using mice transgenic for chimeric dromedary or llama/human IgH loci reported that homodimeric IgG2 can be expressed at the cell surface of developing and mature B cells and can replace IgM in its function during B cell development and as an Ag sensor (33,34). These data could be interpreted as evidence that camelid B cells expressing homodimeric IgGs develop without passing through an IgM ϩ stage.…”

Section: Membrane Igg2 Igg1 and Igm Expressed By Peripheral Blood Bmentioning

confidence: 65%

Tetrameric and Homodimeric Camelid IgGs Originate from the Same IgH Locus

Achour

Cavelier

Tichit

et al. 2008

The Journal of Immunology

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In addition to producing conventional tetrameric IgGs, camelids have the particularity of producing a functional homodimeric IgG type lacking L (light) chains and only made up of two H (heavy) chains. This nonconventional IgG type is characterized by variable and constant regions referred to as VHH and CHH, respectively, and which differ from conventional VH and CH counterparts. Although the structural properties of homodimeric IgGs have been well investigated, the genetic bases involved in their generation are still largely unknown. In this study, we characterized the organization of genes coding for the H chains of tetrameric and homodimeric IgGs by constructing an alpaca (Lama pacos) genomic cosmid library. We showed that a single IgH locus in alpaca chromosome 4 contains all of the genetic elements required for the generation of the two types of Igs. The alpaca IgH locus is composed of a V region that contains both VHH and VH genes followed by a unique DH-JH cluster and C region genes, which include both CHH and CH genes. Although this general gene organization greatly resembles that of other typical mammalian Vn-Dn-Jn-Cn translocon IgH loci, the intermixed gene organization within the alpaca V and C regions reveals a new type of translocon IgH locus. Furthermore, analyses of cDNA coding for the membrane forms of IgG and IgM present in alpaca peripheral blood B cells are most consistent with the notion that the development of a B cell bearing homodimeric IgG passes through an IgM+ stage, similar to the case for conventional IgG.

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“…Bacterial species enable highyield expression of sdAb fragments in the cytosol or in periplasm, whereas yeast cells utilize secretory pathways resulting in efficient disulfide bond formation, N-glycosylation and secretion of the recombinant product into the growth medium. Recent papers also describe several alternative strategies for sdAb fragment engineering and production, such as yeast or ribosome display systems (Yau et al 2003;Ryckaert et al 2010), expression of recombinant fragments in filamentous fungi (Joosten et al 2005), in mammalian cells (Bazl et al 2007) and in transgenic mice (Zou et al 2005). Mutagenesis and recombination of CDR regions to improve the affinity and specificity of sdAb fragments have also been discussed Fanning and Horn 2011).…”

Section: Production Of Sdab Fragments Using Recombinant Technologymentioning

confidence: 99%

Single-domain antibody fragments derived from heavy-chain antibodies: a review

Eyer¹,

Hruška²

2012

Vet. Med.

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Single-domain antibody (sdAb) fragments derived from heavy-chain antibodies of camelids and cartilaginous fish represent a new generation of therapeutic agents and immunoreagents. Due to their unique characteristics, such as low molecular weight, high physical-chemical stability, good water solubility, and the ability to bind antigens inaccessible to conventional antibodies, they could potentially act as a substitute for conventional therapeutic drugs in the treatment of serious human diseases, and, moreover, could be broadly used in analyses and diagnostics. In this review article, an analysis of 826 publications oriented to heavy-chain antibodies and their sdAb fragments indexed in the Web of Science ® database since 1993 has been carried out. Attention has predominantly been paid to papers published from 2010 to June 2012. Key publications are presented in tables and are characterised by descriptive words, abstracts and references. The presented publications have been sorted according to seven basic criteria: review articles and monographs, heavy-chain antibodies of camelids and sharks, production of sdAb fragments using recombinant technology, characteristic properties of sdAb fragments, application of sdAb fragments in therapy, application of sdAb fragments in diagnostic and immunoanalytical methods and other prospective uses of sdAb fragments. This review article should highlight the typical properties of heavy-chain antibodies and sdAb fragments which differentiate them from conventional antibodies and other available recombinant fragments, and also emphasize their extremely broad application potential, mainly in human disease therapy. At the same time it allows an easy and rapid orientation in numerous publications written on this subject, and facilitates the search for the required data.

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Expression of a Dromedary Heavy Chain-Only Antibody and B Cell Development in the Mouse

Abstract: Material

Cited by 28 publications

References 61 publications

Generation of heavy-chain-only antibodies in mice

Generation of heavy-chain-only antibodies in mice

Tetrameric and Homodimeric Camelid IgGs Originate from the Same IgH Locus

Single-domain antibody fragments derived from heavy-chain antibodies: a review

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