Differential utilization of poly (A) signals between DHFR alleles in CHL cells.
The Chinese hamster cell line, DC-3F, is heterozygous at the DHFR locus, and each allele can be distinguished on the basis of a unique DNA restriction pattern, protein isoelectric profile and in the abundancy of the DHFR mRNAs it expresses. Although each allele produces four transcripts, 1000, 1650 and 2150 nucleotides [corrected] in length, the relative distribution of these RNAs differs for each; the 2150 nt mRNA represents the major (60%) species generated from one allele, while the 1000 nt mRNA is the major species generated from the other. The allele that predominantly expresses the 2150 nt transcript is preferentially overexpressed when DC-3F cells are subjected to selection in methotrexate. We have analyzed the 3' ends of both DHFR alleles and have found that the three major mRNAs arise by readthrough of multiple polyadenylation signals. A four base deletion in one allele changes the consensus polyadenylation signal AAUAAA to AAUAAU, resulting in the utilization of a cryptic polyadenylation signal lying 21 bp upstream. Surprisingly, this mutation in the third polyadenylation signal appears to affect not only the utilization of this signal, but also the efficiency with which the first signal, located 1171 bp upstream from the third site, is utilized.
Scotto
K W
KW
Molecular Pharmacology Program, Memorial Sloan-Kettering Cancer Center, New York 10021.
Yang
H
H
Davide
J P
JP
Melera
P W
PW
D000483:Alleles; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D008168:Lung; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011061:Poly A / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D015183:Restriction Mapping; D017384:Sequence Deletion; D012689:Sequence Homology, Nucleic Acid; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*
D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D011061:Poly A; D013762:Tetrahydrofolate Dehydrogenase