Identification of c-myc coding region determinant RNA sequences and structures cleaved by an RNase1-like endoribonuclease.
The coding region of c-myc mRNA encompassing the coding region determinant (CRD) nucleotides (nts) 1705-1792 is critical in regulating c-myc mRNA stability. This is in part due to the susceptibility of c-myc CRD RNA to attack by an endoribonuclease. We have previously purified and characterized a mammalian endoribonuclease that cleaves c-myc CRD RNA in vitro. This enzyme is tentatively identified as a 35 kDa RNase1-like endonuclease. In an effort to understand the sequence and secondary structure requirements for RNA cleavage by this enzyme, we have determined the secondary structure of the c-myc CRD RNA nts 1705-1792 using RNase probing technique. The secondary structure of c-myc CRD RNA possesses five stems; two of which contain 4 base pairs (stems I and V) and three consisting of 3 base pairs (stems II, III, and IV). Endonucleolytic assays using the c-myc CRD and several c-myc CRD mutants as substrates led to the following conclusions: (i) the enzyme prefers to cleave in between the dinucleotides UA, CA, and UG in single-stranded regions; (ii) the enzyme is more specific towards UA dinucleotides. These properties further distinguish the enzyme from previously described mammalian endonuclease that cleaves c-myc mRNA in vitro.
Tafech
Alaeddin
A
Chemistry Program, University of Northern British Columbia, 3333 University Way, Prince George, Canada BC V2N 4Z9.
Bennett
William R
WR
Mills
Fergil
F
Lee
Chow H
CH
D001483:Base Sequence; D004722:Endoribonucleases / Q000378:metabolism*; D016259:Genes, myc; D006801:Humans; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016366:Open Reading Frames; D012333:RNA, Messenger / Q000737:chemistry*
D012333:RNA, Messenger; D004722:Endoribonucleases