Polypyrimidine tract binding protein modulates efficiency of polyadenylation.
Polypyrimidine tract binding protein (PTB) is a major hnRNP protein with multiple roles in mRNA metabolism, including regulation of alternative splicing and internal ribosome entry site-driven translation. We show here that a fourfold overexpression of PTB results in a 75% reduction of mRNA levels produced from transfected gene constructs with different polyadenylation signals (pA signals). This effect is due to the reduced efficiency of mRNA 3' end cleavage, and in vitro analysis reveals that PTB competes with CstF for recognition of the pA signal's pyrimidine-rich downstream sequence element. This may be analogous to its role in alternative splicing, where PTB competes with U2AF for binding to pyrimidine-rich intronic sequences. The pA signal of the C2 complement gene unusually possesses a PTB-dependent upstream sequence, so that knockdown of PTB expression by RNA interference reduces C2 mRNA expression even though PTB overexpression still inhibits polyadenylation. Consequently, we show that PTB can act as a regulator of mRNA expression through both its negative and positive effects on mRNA 3' end processing.
Castelo-Branco
Pedro
P
Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom.
Furger
Andre
A
Wollerton
Matthew
M
Smith
Christopher
C
Moreira
Alexandra
A
Proudfoot
Nick
N
D001483:Base Sequence; D004247:DNA / Q000235:genetics; D005914:Globins / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D026723:Polyadenylation; D038941:Polypyrimidine Tract-Binding Protein / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D012333:RNA, Messenger; D011994:Recombinant Proteins; D038941:Polypyrimidine Tract-Binding Protein; D005914:Globins; D004247:DNA