J. Biol. Chem., Vol. 268, Issue 5, 3180-3183, 02, 1993
Lymphoid V(D)J recombination. Functional analysis of the spacer sequence within the recombination signal
Z Wei and MR Lieber
Department of Pathology, Stanford University School of Medicine, California 94305-5324.
The V(D)J recombination reaction is directed by a pair of signal sequences,
each consisting of a palindromic heptamer, an A/T-rich nonamer, and an
intervening spacer sequence of 12 or 23 base pairs. The spacer sequence
previously has not been analyzed for a functional role. In this study,
numerous simultaneous sequence changes have been made in the spacer of each
signal to test their functional importance. All of the AT base pairs in
each signal were changed to GC base pairs. This particular change is of
interest because it markedly increases the energy that would be required to
melt out the two strands of each signal to permit the intersignal base
pairing proposed in a commonly invoked model for signal-signal interaction
in V(D)J recombination. We find that changing 6 to 12 AT base pairs in the
12-signal to GC does not affect V(D)J recombination, nor does changing 11
of 23 AT base pairs in the 23-signal. Substrates with all-GC spacer
sequences in both the 12- and the 23-signal also recombine at efficiencies
that are not significantly reduced. These studies demonstrate that the
sequences at these particular positions are not recognized by the
recombinase. In addition, the data do not support models invoking
signal-signal base pairing.
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