Structures and functions of band boundaries of mammalian chromosomes

哺乳动物染色体的带区与多种基因组特征有关，如DNA在S期的复制时间和远程GC%镶嵌结构。因此，条带边界被认为与复制时间和GC%水平的过渡区域相对应。在本研究中，为了精确地确定复制开关点，我们在染色体行走和测序后确定了MHC及其周围的非MHC区域XIC （X失活中心）及其周围，以及PAB（假常染色体边界）及其周围的复制时间；然后检测GC%水平与染色体带带的相关性。在MHC II类和III类之间、MHC I类与其端粒非MHC之间、MHC II类与其着丝粒非MHC之间，以及MHC II类与其着丝粒非MHC之间，发现了复制时间和GC%水平的一致过渡。I类及其端粒non-MHC之间的连接处被细胞遗传学指定为6p21.3和p22.1之间的边界。在XIC的XIST位点的3'部分也发现了复制开关，这被认为对应于Xql3.2和q13.3之间的边界，在这些连接区域内，存在特征序列，包括具有三联体形成潜力的多嘌呤/多嘧啶束，三联体已知在多嘌呤/多嘧啶序列中容易发生，并且在体外暂停复制叉运动。使用抗三联体抗体和非变性FISH '与多嘌呤/多嘧啶探针进行免疫染色，发现三联体与人间期细胞核中的单个着丝粒在空间上具有序列特异性。

Band zones of mammalian chromosome are associated with various genome characteristics such as DNA replication timing during S phase and long-range GC% mosaic structures. Therefore, band boundaries are thought to correspond to transition regions for replication timing and GC% levels, In this study, to identify replication switch point precisely, we determined replication timing in MHC and its surrounding non-MHC regions in and around XIC (X inactivation center), and in and around PAB (pseudoautosomal boundary), after chromosome walking and sequencing ; then correlation with GC% levels and chromosome band zones was examined. Concordant transitions for replication timing and GC% level were found in the junctions between MHC classes II and III, between class I and its telomeric non-MHC, and between class II and its centromeric non-MHC.The junction between class I and its telomeric non-MHC was assigned cytogenetically as the border between 6p21.3 and p22.1. The replication switch was also found in the 3' portion of XIST locus in XIC, that is thought to correspond to the border between Xql3.2 and q13.3, Within these junction regions, there are characteristic sequences including polypurine/polypyrimidine tracts with triplex-forming potential Triplexes are known to occur readily in polypurine/polypyrimidine sequences and to pause replication-fork movement in vitro. Using immunostaining with antitriplex antibodies and non-denatured FISH 'with polypurine/polypyrimidine probes, triplexes were found to be sequence-specificaly associated spatially with individual centromeres in the human interphase nucleus.