"In the processing of an mRNA in frog eggs, the mRNA is first transcribed from the DNA, spliced to removed introns, clipped at the 3' end, and a polyadenylate tail is added. The mRNA is then transported to the cytoplasm. Now, many of the mRNAs in Xenopus oocytes are stockpiled and stored for use later in development- so, they sit in the cytoplasm, not being translated into proteins. In fact, as soon as these "stored" mRNAs are transported into the cytoplasm, their polyadenylate (poly A) tails are shortened to about 15-40 nucleotides. mRNAs are then activated for translation into proteins at particular times and in particular regions of the oocyte and developing embryo. The CPE (cytoplasmic polyadenylation element) sequence comes into play at one of the times- when the oocyte is reinitiating its development in preparation for fertilization. Basically, the CPE recruits the cytoplasmic polyadenylation element binding protein (CPEB). CPEB then recruits a protein complex, called CPSF to the formerly quiescent mRNA. CPSF physically interacts with both CPEB and another sequence in the mRNA, to the 3' side of the CPE, called the nuclear polyadenylation hexanucleotide. This particular sequence (AAUAAA) is required for the polyadenylation of mRNAs that occurs en masse in the nucleus. It also turns out, that AAUAAA is ALSO necessary for cytoplasmic polyadenylation (as is apparent by the requirement for the CPSF complex). CPSF, once bound to the mRNA, recruits the enzyme poly A polymerase to the mRNA. THis enzyme then goes about the business of elongating the poly (A) tail of the mRNA. The elongated poly (A) tail then binds up another set of proteins, which interact with proteins bound to the 5' end of the mRNA. These 5' end bound proteins are necessary for translation initiation. Thus, this interaction between the 3' and 5' ends of the mRNA enhances the translation of the mRNA."
Examples of CPE:
Origin CPE CPE separation hexanucleotide B4 RNA uuuuuaau-13nt-AAUAAA G10 uuuuuuauaaag-7nt-AAUAAA c-mos uuuuauAAUAAA (CPE/ hex overlap) cdk2 uuuuau-15nt-uuuuuaauuuuau-57nt-AAUAAA cyclin A1 uuuuuAAUAAA (CPE/ hex overlap) cyclin B1 uuuuuaau-10nt-uuuuAAUAAA cyclin B2 uuuuuauu-45t-aauaaa-8nt-uuuuuuauuu wee1 uuuuuau-12nt-uuuuaAAUAAA-2nt-uuuuaau
Note: In the examples of c-mos and cyclin A1, the CPE overlaps the hexanucleotide! It appears that the CPE and hex can be up to 100nts apart.
This problem motivates the following questions, in building up to a program for the recognition of the motif consisting of CPE sequence, followed by separating nucleotides, followed by AAUAAA.
In part this problem cannot be answered, because currently there is no fully sequenced frog genome; moreover, the frog is tetraploid, unlike the diploid fly, human, etc. Nevertheless, the programs could be developed to run on partial genomic sequences.
3'utr sequence of the Xenopus cyclin B1 cDNA
agg actacgtggc attccaattg tgtattgttg gcaccatgtg cttctgtaaa tagtgtattg tgtttttATAAA gctcatttta acatgWe write the previous 3'UTR EST, which is cDNA (not RNA) without intervening spaces.
aggactacgtggcattccaattgtgtattgttggcaccatgtgcttctgtaaatagtgtattgtgtttttaatgttttactggttttAATAAAgctcattttaacatgWe write and apply a Python to compute the reverse complement of the previous sequence, which yields the reverse complement RNA, and then apply RNAfold from Vienna RNA Package.
catgttaaaatgagcTTTATTaaaaccagtaaaacattaaaaacacaatacactatttacagaagcacatggtgccaacaatacacaattggaatgccacgtagtcct ..((((..((((...((((((......)))))).))))...))))................((...((.((((.((((..........))))...)))).))..)).. minimum free energy = -9.90 kcal/mol