Cryptic Signals and the Fidelity of V(D)J Joining

Abstract

V(D)J recombination is responsible for the de novo creation of antigen receptor genes in T-and B-cell precursors. To the extent that lymphopoiesis takes place throughout an animal's lifetime, recombination errors present an ongoing problem. One type of aberrant rearrangement ensues when DNA sequences resembling a V(D)J joining signal are targeted by mistake. This study investigates the type of sequence likely to be subject to mistargeting, the level of joining-signal function associated with these sequences, and the number of such cryptic joining signals in the genome.In a vertebrate, many millions of B and T cells are produced daily. A pivotal event in B-and T-cell differentiation is the assembly, through DNA recombination, of antigen receptor genes (reviewed in reference 43). Recombination is mediated by two key proteins, called RAG1 and RAG2, as well as an incompletely defined collection of other enzymes and factors (reviewed in references 28 and 56). During pre-B-cell and pre-T-cell development, DNA segments termed V, D, and J are assembled to form the variable exon of the expressed immunoglobulin (Ig) and T-cell receptor (TCR) genes. The joined gene segments encode the highly variable antigen-binding domain of the antigen receptor protein.V(D)J assembly is accomplished through DNA rearrangements that can encompass up to several megabases. The sequence motifs that serve as recognition elements for this process (joining signals) are surprisingly compact. A joining signal, found adjacent to every V, D, or J segment, is comprised of a heptamer (CACAGTG), a spacer (of 12 or 23 bp), and a nonamer (ACAAAAACC) (Fig. 1A). No sequence apart from the 28 (or 39)-bp joining signal motif is required for sitespecific recognition and recombination (reference 33 and references cited therein).Any V(D)J recombination event is normally an interaction between two gene segments, one of which possesses a 12-bp spacer (12-spacer) signal, and the other of which possesses a 23-bp spacer (23-spacer) signal. The molecular signature of V(D)J recombination is production of two types of junctions. One of these, the coding joint, is formed from the fusion of the two coding sequences (e.g., V and J), and the other, a signal joint, is formed from the corresponding joining signals. It is usual for a coding joint to have a small and variable degree of base loss and insertion at the point where the two coding elements connect, whereas the signal joint is typified by a precise fusion, heptamer to heptamer, of the two signals.Not surprisingly, DNA sequences that resemble an authentic joining signal (cryptic signals) are sometimes rearranged by mistake (first described in reference 36). The result is the production of recombinant junctions that are structurally analogous to coding joints and signal joints (Fig. 1) (17, 36). In this report, we take production of the characteristic junctions as the defining, diagnostic feature of a cryptic signal.Cryptic signals are seen at several of the Ig and TCR loci in both mice and humans and are active in re...

DNA structures at chromosomal translocation sites

“…(19,24) Recent studies have shown that the LMO-2, Ttg1, Tal1(SCL1), Tal2 (SIL), MTS1 fragile sites represent unintended or cryptic sites (pseudo-signals is another term for these), which the RAGs treat as normal RSS sequences because their sequence is quite close to that of an RSS. (25)(26)(27)(28) This misrecognition of the target sequence leads to these translocations or deletions.…”

Clonal diversification of primary BALB/c plasmacytomas harboring T(12;15) chromosomal translocations

“…The reason for the putative allele-specific instability is not known, but it is possible that it is related to the known instability of the Igh locus in plasmacytoma cells. [15][16][17][18] Another attempt to explain the instability considers a role for the 3Ј-C␣ enhancers, which are Since the 3Ј-C␣ enhancers have been invoked before as possible control elements for isotype switching, 19 illegitimate recombinations in the C H gene cluster [20][21][22] and, indeed, for the general activation of the chromatin domain containing the Igh locus, 23 it seems plausible that the enhancer may also facilitate the deletional mutagenesis in Chr T(12;15)-typical Igh/c-myc junctions. This mutagenesis, which appeared to take three distinct forms, will be discussed in the following three paragraphs.…”

Cytosines, but Not Purines, Determine Recombination Activating Gene (RAG)-induced Breaks on Heteroduplex DNA Structures

“…In the recent past, studies have shown that cryptic RSS sites present elsewhere in the genome can also act as off-target sites for RAG misrecognition, leading to chromosomal translocations in lymphoid cancers such as leukemia (21)(22)(23)(24). In addition to its sequence-specific endonuclease activities, recent studies have shown that the RAG complex can act as a structure-specific nuclease (22).…”

“…After cleavage, the RAG complex remains tightly bound to the two signal ends and less tightly bound to the coding end, in a postcleavage complex (17,18). Finally, the complete exon coding for antibody or TCR (T-cell receptor) is generated by joining of the broken subexons by nonhomologous DNA endjoining (NHEJ) (19,20).In the recent past, studies have shown that cryptic RSS sites present elsewhere in the genome can also act as off-target sites for RAG misrecognition, leading to chromosomal translocations in lymphoid cancers such as leukemia (21)(22)(23)(24). In addition to its sequence-specific endonuclease activities, recent studies have shown that the RAG complex can act as a structure-specific nuclease (22).…”