To investigate the relationship between transcription and polyadenylation, an in vitro system has been developed in which endogenously transcribed pre-mRNAs containing functional polyadenylation sites are rapidly and accurately cleaved in a HeLa nuclear extract. Cleavage of endogenously transcribed substrates differed from that of exogenous substrates in that a proximal 3' terminus was not requ...

Expression vectors normally contain sequences that stimulate polyadenylation of the expressed mRNA. These sequences usually include an AATAAA consensus sequence 20-30 nucleotides 5' of a diffuse GTor T-rich stretch immediately following the sequences that form the 3' end of the mRNA. The efficiency of the two minimal elements is also influenced by additional upor downstream elements (1). Very o...

Polyadenylation of RNA is a posttranscriptional modification that can play two somewhat opposite roles: stable polyadenylation of RNA encoded in the nuclear genomes of eukaryote cells contributes to nuclear export, translation initiation, and possibly transcript longevity as well. Conversely, transient polyadenylation targets RNA molecules to rapid exonucleolytic degradation. The latter role ha...

The maternal mRNAs that drive meiotic progression in oocytes contain short poly(A) tails and it is only when these tails are elongated that translation takes place. Cytoplasmic polyadenylation requires two elements in the 3'-UTR (3'-untranslated region), the hexanucleotide AAUAAA and the CPE (cytoplasmic polyadenylation element), which also participates in the transport and localization, in a q...

Polypyrimidine tract-binding protein (PTB) is a splicing regulator that also plays a positive role in pre-mRNA 3' end processing when bound upstream of the polyadenylation signal (pA signal). Here, we address the mechanism of PTB stimulatory function in mRNA 3' end formation. We identify PTB as the protein factor whose binding to the human beta-globin (HBB) 3' UTR is abrogated by a 3' end proce...

The L1 and L2 capsid genes of human papillomavirus type 31 (HPV-31) are expressed late in the differentiation-dependent life cycle from a promoter located in the E7 open reading frame (ORF) of the early region. These late HPV genes are transcribed by RNA polymerase II which reads through the region containing early polyadenylation signals and proceeds to a poly(A) site downstream of L1. In this...

Most genes have multiple polyadenylation sites (PAS), which are often selected in a tissue-specific manner, altering protein products and affecting mRNA stability, subcellular localization and/or translability. Here we studied the polyadenylation mechanisms associated to the beta-adducin gene (Add2). We have previously shown that the Add2 gene has a very tight regulation of alternative polyaden...

Toll encodes a receptor that is critical for dorsal-ventral patterning in the early Drosophila embryo. Previous data have suggested that the accumulation of Toll protein in the embryo temporally correlates with elongation of the poly (A) tail of the message. Here, we demonstrate that Toll mRNA is translationally activated by regulated cytoplasmic polyadenylation. We also identify a 192 nucleoti...

Transcription termination by RNA polymerase II is coupled to transcript 3' end formation. A large cleavage and polyadenylation complex containing the major poly(A) polymerase Pap1 produces mRNA 3' ends, whereas those of nonpolyadenylated snoRNAs in yeast are formed either by endonucleolytic cleavage or by termination, followed by trimming by the nuclear exosome. We show that synthesis of indepe...

3' polyadenylation is a key step in eukaryotic mRNA biogenesis. In mammalian cells, this process is dependent on the recognition of the hexanucleotide AAUAAA motif in the pre-mRNA polyadenylation signal by the cleavage and polyadenylation specificity factor (CPSF) complex. A core CPSF complex comprising CPSF160, WDR33, CPSF30 and Fip1 is sufficient for AAUAAA motif recognition, yet the molecula...