The isolation and characterization of HeLa cell messenger RNA-like molecules containing uridylic acid-rich oligonucleotide sequences.
HeLa cell mRNA, isolated from mRNPs released from polysomes by EDTA to minimize contamination from nuclear RNA, has been separated into four classes of molecules: 1) those containing poly(A) sequences [poly(A+)] as well as oligo(U) sequences [oligo(U+)] (4 to 13%); 2) poly(A+) oligo(U-) (approximately 52%); 3) poly(A-) oligo(U+) (approximately 2%); 4) poly(A-) oligo(U-) (approximately 33%). The oligo(u) segments are 89% UMP and range from 20 to 50 nucleotides in length. The poly(A+) oligo(U+) mRNAs appear to contain the oligo(U) in a region with secondary structure (possibly an intramolecular duplex with the 3'-poly(A) since they did not bind to poly(A) Sepharose without prior HCHO modification. HCHO modifies the exocyclic amino groups of CMP, GMP, and AMP and prevents hydrogen bonding but reacts only slightly with UMP. After removal of the excess HCHO, the oligo(U) of the mRNA was free to bind to the poly(A) Sepharose. Control experiments indicated that the binding of oligo(U+) mRNA to poly(A) Sepharose was specific and the HCHO did not cause cross-linking of the RNAs. The poly(A+) oligo(U+) RNAs appeared to contain one oligo(U)/molecule and could re-bind to poly(A) Sepharose with 31 to 75% efficiency. The poly(A+) oligo(U+) mRNA class was more heterogeneous in size and slightly larger (approximately 3 kb average length) than the other mRNA classes.
D001483:Base Sequence; D002846:Chromatography, Affinity; D006367:HeLa Cells / Q000032:analysis; D006801:Humans; D008970:Molecular Weight; D009843:Oligoribonucleotides / Q000032:analysis; D011061:Poly A; D012333:RNA, Messenger / Q000302:isolation & purification*; D012685:Sepharose; D014500:Uracil Nucleotides / Q000032:analysis*; D014542:Uridine Monophosphate / Q000032:analysis*
D009843:Oligoribonucleotides; D012333:RNA, Messenger; D014500:Uracil Nucleotides; D011061:Poly A; D012685:Sepharose; D014542:Uridine Monophosphate