The products of pre-B cell-specific genes (lambda 5 and VpreB) and the immunoglobulin mu chain form a complex that is transported onto the cell surface.

Abstract

F ully assembled Ig H and L chains are transported onto the surface of B cells, while free H chains are retained in the endoplasmic reticulum (ER) in association with BiP (1) . However, in pre-B lines that produce only lc H chains, small amounts of P. chains are expressed on the cell surface, suggesting that some other proteins substitute for the missing L chain in these cells. In fact, proteins of 14-22 kD have been shown to bind to the cytoplasmic and surface p, chains in murine and human pre-B lines (2-4). The X5 and VpreB genes, which are specifically expressed in pre-13 cells, have strong homology to the C and the V regions of the X L chain genes, respectively (5, 6), and thus, are likely to encode the p-binding proteins. Antibodies specific for the human X5 and VpreB peptides react against A-binding molecules of 22 and 18 kD, respectively, in human pre-13 lines (7,8) . However, whether these proteins are expressed in association with the A chain on the cell surface has not yet been shown.We introduced expression vectors coding for the murine A5 and VpreB genes, together with a it vector (pSVAtm), into Ig -myeloma cells and we demonstrate that the products of the two genes are required and sufficient for p, expression on the cell surface. This result suggests that X5 and VpreB are essential components of sIg in pre-B cells . Materials and MethodsBrief Definitive Report SummaryWe constructed expression vectors coding for the two pre-B-specific genes, VpreB and X5, and transfected them together with a A vector (Atm) into Ig-, myeloma cells. In a transfectant expressing all three introduced genes, the P,tm chain is transported on the cell surface. A biochemical analysis demonstrated that, in these cells, the P.tm chain is associated noncovalently with an 18-kD protein and covalently with a 22-kD protein, 'which are most likely the products of VpreB and X5, respectively. Our results, thus, strongly suggest that the products of X5 and VpreB bind to A chains and have the same capacity as conventional Ig L chains to allow surface expression of A. chains.The expression vectors coding for X5 and Vp,,Bl, pEPX5gpt and pEPVpBneo, are shown in Fig . 1, B and C. Genomic fragments ofA5 and VprcBl genes were provided by Dr. A . Kudo, Basel Institute for Immunology, Basel, Switzerland. The Bg1II-PstI fragment of X5 was cloned into a pSV2gpt vector containing the VH promoter and the IgH enhancer. The Bg1II fragment of Vpr,Bl gene was similarly cloned into a pSV2neo vector. pSVAtm (9) is an expression vector for a chimeric A chain where the membrane exon of Am was replaced by the transmembrane and cytoplasmic part of the H-2K1 gene (Fig. 1 A). The vector for the X, L chain, pSVX,neo, was described previously (10). The vectors were transfected into myeloma cells by electroporation. Cell surface expression of 1A chains in the transfectants was analyzed by flow cytometry as described previously (9) .For biochemical analysis, biosynthetic labeling, and surface iodination were done as described previously (9, 10) . Cells were the...

Transgenic Human λ5 Rescues the Murine λ5 Nullizygous Phenotype

“…Our results suggest that the unique 50-aa region of hu 5 (which shows the highest percent amino acid difference from the mouse at 50%) is not hindering mVpreB or m association. The presence of mVpreB is likely a crucial player in assembly of the mouse/human chimeric pre-BCR in hu 5 transgenic mice with deletions of both m 5 alleles, by extension from the mouse studies (8,20,22,63). Consistent with the Minegishi et al report (62), our results suggest that the unique amino-terminal region of the hu 5 protein can function in the m 5 Ϫ/Ϫ mouse for assembly into the chimeric pre-BCR.…”

“…Association of m with the m 5 and mVpreB proteins is necessary to get cell-surface expression of m (8,20,58). Recently Minegishi et al (62) demonstrated that the unique 50-aa region of the hu 5 protein, located carboxylterminal to the signal peptide, serves as an intramolecular chaperone to prevent folding of hu 5 protein in the absence of its partner, VpreB.…”

“…The mouse SLC genes, m 5 and mVpreB, were initially identified by their exclusive expression in pre-B cells (3)(4)(5)(6). The SLC together with a rearranged Ig ( ) HC and the Ig-associated transducing chains, Ig␣ and Ig␤, comprise the pre-B cell receptor (pre-BCR) (7)(8)(9). The exact function of the SLC, including 5, is unknown, but the pre-BCR apparently plays a critical role in B lineage differentiation as targeted disruption of the m 5 gene blocks B lymphocyte development at the transition between the pro-B cell and large pre-B cell stages and drastically reduces the number of mature B lymphocytes in the periphery (10 -12).…”

Tolerance checkpoints in B‐cell development: Johnny B good

“…At this stage of differentiation, the IgH chain loci are V to D-J rearranged and those that have done this successfully (synthesized a m-heavy (mH) chain) are positively selected within this compartment. This positive selection is mediated by the pre-BCR, which is composed of the mH chain and the surrogate light chain proteins Vpre-B and l5 [10][11][12][13][14][15]. Moreover, the pre-BCR induces a strong proliferation of these positively selected pre-B cells [12,13].…”

Pre-B Cell Receptor Assesses the Quality of IgH Chains and Tunes the Pre-B Cell Repertoire by Delivering Differential Signals

“…The rearrangement at the IgH locus usually takes place before that at the IgL locus during B cell development (1,2). Once a productive rearrangement occurs at the IgH locus at the pro-B cell stage, H chains are produced and assembled with invariant VpreB/ 5 surrogate light (SL) 3 chains to form the pre-BCR (4,5). A deficiency in pre-BCR formation or signaling results in severe impairment of B cell differentiation in both humans and mice (6).…”