Position independent and copy-number-related expression of the bovine neonatal Fc receptor α-chain in transgenic mice carrying a 102 kb BAC genomic fragment

Abstract

We generated and characterized transgenic mice carrying a 102 kb bovine genomic fragment, encoding the neonatal Fc receptor alpha-chain (bFcRn). FcRn plays a crucial role in the maternal IgG transport and it also regulates the IgG and albumin homeostasis. Some of its functions and transcriptional regulation show species specific differences. The FcRn heterodimer is composed of the alpha-chain and beta-2-microglobulin (beta2 m). A bacterial artificial chromosome containing the bovine FcRn alpha-chain gene (bFCGRT) with its 44 kb 5' and 50 kb long 3' flanking sequences was microinjected into fertilized mouse oocytes. Two of the transgenic lines generated, showed copy number related and integration site independent bFcRn expression. The bFcRn alpha-chain forms a functional receptor with the mouse beta2-microglobulin and extends the half-life of the mouse IgG in transgenic mice. Our results underline the feasibility of creating BAC transgenic mouse models of economically important bovine genes.

Neonatal FcR Overexpression Boosts Humoral Immune Response in Transgenic Mice

“…The generation and basic phenotype of the Tg mice have been recently published (14). Briefly, a BAC containing the bovine FcRn a-chain gene (bFCGRT) with its 44-kb-long 59 and 50-kb-long 39 flanking sequences was microinjected into fertilized mouse oocytes.…”

“…Therefore, one of our goals was to analyze the effect of FcRn overexpression on the Ag-specific IgG response. We performed experiments employing the transgenic (Tg) mice we have recently created (14), using a bacterial artificial chromosome (BAC) that contains the bovine FcRn (bFcRn) a-chain gene (bFCGRT). Two of the Tg lines showed copy numberrelated bFcRn expression and demonstrated a significantly extended half-life of mouse IgG (mIgG), indicating that bFcRn forms a functional complex with the mouse b 2 m and thus binds and protects mIgG (14).…”

NFκB induces overexpression of bovine FcRn

“…To study whether NFκB transcription factors bind to and induce the bFC-GRT promoter (bFP) regions, a luciferase reporter gene construct pGL3-bFP2 (-1787 + 92) was first created that harbors a 1787 bp long segment of the promoter and a 92 bp long segment of the non-coding exon1 of the bFCGRT ( Fig. S1; numbers are relative to the first nucleotide of exon1 of the bFCGRT 31 ). Human p65 overexpression resulted in 4-, 12-, and 10-fold increased luciferase activity of the pGL3-bFP2 construct in HEK-293, HC11 and JAR cell lines, respectively, compared with the activity of the empty vector, indicating that this DNA segment can be induced by NFκB (Fig.…”

“…33 Vascular endothelium is proposed to be a major site at which FcRn protects serum IgG and albumin from lysosomal degradation and it is known that FcRn overexpression enhances IgG recycling and elongates its biological half-life. 31,43 However, a local inflammation that may lead to the upregulation of bFcRn affects only a relatively small portion of blood vessels, and thus one may not expect that it would meaningfully influence the biological half-life of IgG. On the other hand, FcRn upregulation in capillary endothelial cells may increase local IgG tissue distribution via IgG transcytosis in inflamed tissues, and thus may contribute in local, IgG mediated protection.…”

“…Total RNA was isolated from spleens and 3 μg of total RNA was size fractionated on 1% agarose/2.2 M formaldehyde gel, transferred to Hybond N+ membrane (Amersham) and hybridized with the 32 P-labeled cDNA probe synthesized by PCR primers as described. 31 The signals obtained were evaluated using Fuji medical X-Ray film granted a European patent for this technology (EP2097444), and seek patent in other major markets. The technology has been licensed exclusively to ImmunoGenes Ltd.…”