Le M H, Duricka D, Karpen G H
Molecular Biology and Virology Laboratory, Salk Institute for Biological Studies, La Jolla, California 92037, USA.
Genetics. 1995 Sep;141(1):283-303. doi: 10.1093/genetics/141.1.283.
Heterochromatin is a ubiquitous yet poorly understood component of multicellular eukaryotic genomes. Major gaps exist in our knowledge of the nature and overall organization of DNA sequences present in heterochromatin. We have investigated the molecular structure of the 1 Mb of centric heterochromatin in the Drosophila minichromosome Dp1187. A genetic screen of irradiated minichromosomes yielded rearranged derivatives of Dp1187 whose structures were determined by pulsed-field Southern analysis and PCR. Three Dp1187 deletion derivatives and an inversion had one breakpoint in the euchromatin and one in the heterochromatin, providing direct molecular access to previously inaccessible parts of the heterochromatin. End-probed pulsed-field restriction mapping revealed the presence of at least three "islands" of complex DNA, Tahiti, Moorea, and Bora Bora, constituting approximately one half of the Dp1187 heterochromatin. Pulsed-field Southern analysis demonstrated that Drosophila heterochromatin in general is composed of alternating blocks of complex DNA and simple satellite DNA. Cloning and sequencing of a small part of one island, Tahiti, demonstrated the presence of a retroposon. The implications of these findings to heterochromatin structure and function are discussed.
异染色质是多细胞真核生物基因组中普遍存在但了解甚少的组成部分。我们对异染色质中存在的DNA序列的性质和整体组织的认识存在重大差距。我们研究了果蝇小染色体Dp1187中1 Mb着丝粒异染色质的分子结构。对辐照后的小染色体进行遗传筛选,得到了Dp1187的重排衍生物,其结构通过脉冲场Southern分析和PCR确定。三个Dp1187缺失衍生物和一个倒位在常染色质中有一个断点,在异染色质中有一个断点,为直接分子进入异染色质中以前无法进入的部分提供了途径。末端探针脉冲场限制性图谱显示存在至少三个复杂DNA的“岛”,即塔希提岛、莫雷阿岛和博拉博拉岛,约占Dp1187异染色质的一半。脉冲场Southern分析表明,果蝇异染色质一般由复杂DNA和简单卫星DNA的交替块组成。对一个岛(塔希提岛)的一小部分进行克隆和测序,证明存在一个反转录转座子。讨论了这些发现对异染色质结构和功能的影响。