De Novo Generation of Simple Sequence during Gene Amplification

Mammalian cells that have undergone gene amplification and/or gene rearrangement have been used as resources to gain insight into the questions of chromosome structure and dynamics. The multidrug resistant murine cell line J7.V2-1 has been shown previously to contain two distinct forms of the highly amplified mdr2 gene, a member of the mouse gene family responsible for the multidrug resistant (MDR) phenotype [Kirschner, L. S. (1995) DNA Cell Biol. 14, 47-59]. Characterization of both forms of the gene revealed that one form corresponded to the wild-type structure of the gene, whereas the other represented a rearrangement. Investigation of this altered gene demonstrated a deletion of 1.6 kb of the wild-type sequence, and replacement of this region with a poly(AT) tract that appears to have been generated de novo. Analysis of the native sequence in this region demonstrated the absence of repetitive elements, but was notable for the presence of two long stretches of polypurine: polypyrimidine strand asymmetry. Analysis of mdr2 transcripts in this cell line revealed that nearly all of the mRNA is transcribed from the rearranged form of the gene. This message is unable to code for a functional mdr2 gene product, owing to a deletion of the fourth exon during this event. Mechanisms of the rearrangement, as well as the significance of this curious effect on transcription, are discussed.