MECHANISTIC ANALYSIS OF PREFERENTIAL DHRSS TARGETING

重组信号序列（RSS）靶向V（D）J重组 它们与功能性免疫受体基因紧密相连 片段虽然RSS侧翼的D/H段似乎是等同的， 并不是随机使用的。我们发现小鼠D/H基因重排 至少1000倍更频繁地通过删除比通过倒位。在 在人B细胞中，缺失D/H/J/H重排有利于10-100倍 而不是反转重排。人工重组底物 证明了在D/H-J/H中3' D/H RSS的强偏倚 重排可以在细胞系中重现，并且依赖于 几个因素，包括紧邻的编码序列， D/H RSS，D/H片段本身的重组伴侣。这些 数据表明，某些RSS组合更好地靶向重组 than others. 反向D/H片段编码约100个带正电荷的残基。 是正常方向D/H段速率的两倍。数据表单 几个不同的研究人员已经证明了以下几点：1） 抗DNA特异性与带正电荷重链之间的关联 链CDR 3/s;和2）在CDR 3/s中发现倒位D/H基因区段的倾向。 抗DNA抗体由于反向D/H重排是相对 罕见，特异性抗体中D/H倒置频率高 DNA需要进一步研究 因此，本建议的目的是划定顺式和反式- 针对特定12/23 RSS组合的作用元素， 5 ′ D/H RSS优先加入V/H RSS，3 ′ D/H RSS优先加入J/H。 并确定D/H RSS靶向是否在自身免疫性疾病中失调， 条件

V(D)J recombination is targeted by recombination signal sequences (RSS) which are found immediately adjacent to functional immune receptor gene segments. While the RSS flanking D/H segments appear to be equivalent they are not randomly utilized. We have found that murine D/H genes rearrange at least 1000 times more frequently by deletion than by inversion. In human B cells, deletion D/H/J/H rearrangement is favored by 10-100 fold over inversional rearrangement. Artificial recombination substrates demonstrate that the strong bias for the 3' D/H RSS in D/H-J/H rearrangement can be recapitulated in cell lines, and is dependent on several factors including the coding sequence immediately adjacent to the D/H RSS, the recombination partner to which the D/H segment itself. These data suggest that certain RSS combinations target recombination better than others. Inverted D/H segments encodes positively charged residues at approximately twice the rate as D/H segments in the normal orientation. Data forms several different investigators have demonstrated the following: 1) an association between anti-DNA specificity and positively charged heavy chain CDR3/s; and 2) a propensity to find inverted D/H gene segments in anti-DNA antibodies. Since inverted D/H rearrangements are relatively rare, the high frequency of D/H inversions in antibodies with specificity for DNA warrants further study. Thus, the objectives of this proposal are to delineate the cis and trans- acting elements which target specific 12/23 RSS combinations using the preferential joining of the 5'D/H RSS to V/H RSS and 3' D/H RSS to J/H. and to determine whether D/H RSS targeting is dysregulated in autoimmune conditions.