Analysis of Genes for DNA Replication Enzymes and Homeobox Proteins in Congenital Multiple Anomalies.

1. DNA聚合酶δ辅助蛋白（PCNA）和同源蛋白：我们分离了编码水稻PCNA的基因组克隆，并对其中一个克隆进行了部分测序。水稻的氨基酸序列与大鼠PCNA（P.N. a. S. 86：3189 - 3193，1989）。我们还测定了果蝇PCNA基因的完整核苷酸序列，并在5 ′-侧翼区发现了一簇含有同源结构域的基因产物的假定结合位点。足迹分析显示果蝇同源结构域蛋白（偶跳跃和zerknullt）可以特异性地将这些位点结合（M。C. B。10：872 - 879，1990）。为了证明人类发育程序化分化和细胞增殖调控之间的密切关系，我们分离了人类PCNA和同源框4A.2的基因组克隆。先天性畸形DNA复制酶基因和同源盒基因的研究：用PCNA、DNA聚合酶β和DNA聚合酶α探针对15例先天性畸形中的1例进行DNA印迹杂交，发现异常限制性片段。以DNA聚合酶β和γ-晶体为探针，在95例正常人中检出1例异常限制性片段。使用盒4A和4B探针的DNA印迹分析未显示先天性异常的DNA样本中存在任何不寻常的限制性片段。用染色体2同源框4位点附近的γ-结晶DNA片段检测了30例严重先天畸形的DNA样本的多态性。先天性畸形和γ-晶体加内斯的RFLP之间存在统计学显著相关性。该结果表明，与先天性畸形密切相关的基因座存在于γ-晶体和同源框4基因座附近（Nucleic Acids Symp. 21：19 - 20，1989，IRL出版社）。

1. DNA polymerase delta auxiliary protein (PCNA) and homeopox protein : We isolated genomic clones encoding rice PCNA and partially sequenced one of the clones. The rice amino acid sequence shared 60% homology with the sequence of the rat PCNA (P. N. A. S. 86 : 3189-3193, 1989). We also determined the c omplete nucleotide sequence of the Drosophila PCNA gene and found a cluster of putative binding sites for the homeodomain-containing gene products in the 5^,-flanking region. Footprint analysis revealed that Drosophila homeodomain proteins (even-skipped and zerknullt) can specifically bi these sites (M. C. B. 10 : 872-879, 1990). In order to demonstrate the close relationship between the regulation of developmentally programmed differentiation and that of cell proliferation in man, we isolated genomic clones for human PCNA and homeobox 4A.2. Studies on genes for DNA replication enzyme and on homeobox genes in congenital anomalies : Abnormal restriction fragments were detected by DNA blot hybridization with PCNA, DNA polymerase beta and DNA polymerase alpha probes in one of 15 cases of congenital malformations. Abnormal restriction frgments were detected in one case of 95 normal individuals when DNA polymerase beta and gamma-crystalline were used as probes. DNA blot analysis using box 4A and 4B probes did not show any unusual restriction fragments in the DNA samples of congenital anomalies. Thirty DNA samples of severe degree of congenital malformations were examined for polymorphisms with gamma-crystalline DNA fragment which has been mapped near the homeobox 4 locus in chomosome2. Statistically significant association was demonstrated between congenital malformations and RFLPs of gamma-crystalline ganes. This result indicates that the locus closely linked to congenital malformations is present in the proximity of gamma-crystalline and homeobox 4 loci (Nucleic Acids Symp. Ser. 21 : 19-20, 1989, IRL press).