Nicking is asynchronous and stimulated by synapsis in 12/23 rule-regulated V(D)J cleavage.

Abstract:

:The first step in DNA cleavage at V(D)J recombination signals by RAG1 and RAG2 is creation of a nick at the heptamer/coding flank border. Under proper conditions in vitro the second step, hairpin formation, requires two signals with spacers of 12 and 23 bp, a restriction referred to as the 12/23 rule. Under these conditions hairpin formation occurs at the two signals at or near the same time. In contrast, we find that under the same conditions nicking occurs at isolated signals and hence is not subject to the 12/23 rule. With two signals the nicking events are not concerted and the signal with a 12 bp spacer is usually nicked first. However, the extent and rate of nicking at a given signal are diminished by mutations of the other signal. The appearance of DNA nicked at both signals is stimulated by more than an order of magnitude by the ability of the signals to synapse, indicating that synapsis accelerates nicking and often precedes it. These observations allow formulation of a more complete model of catalysis of DNA cleavage and how the 12/23 rule is enforced.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Eastman QM,Schatz DG

doi

10.1093/nar/25.21.4370

subject

Has Abstract

pub_date

1997-11-01 00:00:00

pages

4370-8

issue

21

eissn

0305-1048

issn

1362-4962

pii

gka677

journal_volume

25

pub_type

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