Lymphoid V(D)J recombination: nucleotide insertion at signal joints as well as coding joints

Area of Science:

• Immunology
• Molecular Biology
• Genetics

Background:

• Variable-diversity-joining region (V(D)J) recombination assembles antigen receptor genes.
• Coding joints in V(D)J recombination often exhibit nucleotide deletions and insertions.
• The role of terminal deoxynucleotidyltransferase (TdT) in signal joint modification is less understood.

Purpose of the Study:

• To investigate nucleotide additions and deletions at signal joints during V(D)J recombination.
• To determine the influence of terminal deoxynucleotidyltransferase (TdT) on signal joint modifications.
• To compare modifications at signal joints with those at coding joints.

Main Methods:

• Transfection of lymphoid cell lines with extrachromosomal recombination substrates.
• Analysis of nucleotide sequences at both signal and coding joints.
• Quantification of terminal deoxynucleotidyltransferase (TdT) activity in cell lines.

Main Results:

• Signal joints, unlike coding joints, rarely undergo nucleotide loss but frequently incorporate insertions.
• Nucleotide addition frequency at signal joints correlates with terminal deoxynucleotidyltransferase (TdT) levels in pre-B-cell lines.
• Insertions at both signal and coding joints are G+C rich, consistent with TdT's base preference.

Conclusions:

• Terminal deoxynucleotidyltransferase (TdT) plays a significant role in nucleotide additions at signal joints during V(D)J recombination.
• Signal joint modification by TdT differs from coding joint modification, with less nucleotide loss.
• The G+C richness of insertions suggests a conserved mechanism of TdT activity in V(D)J recombination.