Evidence for somatic rearrangement of immunoglobulin genes coding for variable and constant regions.

Abstract

A high-molecular-weight DNA from Balb/c mouse early embryo or from MOPC 321 plasmacytoma (a Kchain producer) was digested to completion with Bacillus amyloliquefaciens strain H restriction enzyme (BamH I). The resulting DNA fragments were fractionated according to size in preparative agarose gel electrophoresis. DNA fragments carrying gene sequences coding for the variable or constant region of K chains were detected by hybridization with purified, 12SI-labeled, whole MOPC 321 K mRNA and with its 3'-end half. The pattern of hybridization was completely different in the genomes of embryo cells and of the plasmacytoma. The pattern of embryo DNA showed two components, one of which (molecular weight = 6.0 million) hybridized with C-gene sequences and the other (molecular weight = 3.9 million) with V-gene sequences. The pattern of the tumor DNA showed a single component that hybridized with both V-gene and C-gene sequences and that is smaller (molecular weight = 2.4 million) than either of the components in embryo DNA. The results were interpreted to mean that the VK and CK genes, which are some distance away from each other in the embryo cells, are joined to form a contiguous polynucleotide stretch during differentiation of lymph~c~es. Such joining occurs in both of the homologous chromosomes. Relevance of these findings with respect to models for V-C gene joining, activation of a specific Vk.gene, and allelic exclusion in immunoglobulin gene loci is discussed. Both light and heavy chains of immunoglobulin molecules consist of two regions: the variable region (V region) and the constant region (C region) (1, 2). Uniqueness (i.e., one copy per haploid genome) of the genetic material coding for C region ("C gene") has been conjectured from normal Mendelian segregation of allotypic markers (3). Nucleic acid hybridization studies have confirmed this notion (4-10). Hybridization studies have also demonstrated that a group of closely related V regions are somatically generated from a few, conceivably even a single, germline gene(s) (V gene) (4-6). (12), except that 10% glycerol was present in all buffers during the phosphocellulose step. Five milligrams of high-molecular-weight embryo or MOPC 321 tumor DNA in a buffer consisting of 6 mM TrisHCl, pH 7.4,6 mM MgCl2, and 6 mM 2-mercaptoethanol was incubated with 104 units (1 unit is defined as the amount of the enzyme sufficient for digesting 1 ,ug of phage X DNA in 60 min at 370 under the above conditions) of the purified enzyme at 370 for 4 hr. In order to investigate completeness of digestion we followed the following scheme. An aliquot of the reaction mixture containing mouse DNA was removed before incubation and mixed with a small amount (ratio of X DNA to mouse DNA, 1:10) of phage X DNA. This pilot mixture was incubated in parallel with the main reaction mixture under the conditions described above. After incubation the pilot mixture was electrophoresed in 0.9% agarose. DNA was visualized under ultraviolet light after staining with 1 ,g/ml of ethidium bromide ...

Immunoglobulin Gene Transcripts Have Distinct VHDJH Recombination Characteristics in Human Epithelial Cancer Cells

“…Ig gene recombination, as theorized previously by Dreyer and Bennett, was confirmed to exist in mouse myeloma cells using a probe against the Ig mRNA kappa chain (4,5).…”

Developmental hierarchy of immunoglobulin gene rearrangements in human leukemic pre-B-cells.

“…multiple constant (CL) regions (1)(2)(3)(4)(5)(6)(7). Heavy chain genes are similarly organized (VH, JH, CH) but appear to have an additional diversity (DH) segment incorporated between their VH and JH regions (8)(9)(10)(11)(12)(13).…”

Immunoglobulin gene conversion: Insights from bursal B cells and the DT40 cell line

“…In humans and mice, functional V, D, and J gene segments exist in large clusters and V(D)J recombination generates combinatorial diversity by the random assortment of individual V, D, and J segments (Hozumi and Tonegawa, 1976). After antigen stimulation, the functionally rearranged V segments are further modified by somatic hypermutation (Jacob et al, 1991) and the Ig isotope can be changed by class switch recombination (Honjo and Kataoka, 1978).…”