We have previously identified a G-rich sequence (GRS) as an auxiliary downstream 
element (AUX DSE) which influences the processing efficiency of the SV40 late 
polyadenylation signal. We have now determined that sequences downstream of the 
core U-rich element (URE) form a fundamental part of mammalian polyadenylation 
signals. These novel AUX DSEs all influenced the efficiency of 3'-end processing 
in vitro by stabilizing the assembly of CstF on the core downstream URE. Three 
possible mechanisms by which AUX DSEs mediate efficient in vitro 3'-end 
processing have been explored. First, AUX DSEs can promote processing efficiency 
by maintaining the core elements in an unstructured domain which allows the 
general polyadenylation factors to efficiently assemble on the RNA substrate. 
Second, AUX DSEs can enhance processing by forming a stable structure which 
helps focus binding of CstF to the core downstream URE. Finally, the GRS 
element, but not the binding site for the bacteriophage R17 coat protein, can 
substitute for the auxiliary downstream region of the adenovirus L3 
polyadenylation signal. This suggests that AUX DSE binding proteins may play an 
active role in stimulating 3'-end processing by stabilizing the association of 
CstF with the RNA substrate. AUX DSEs, therefore, serve as a integral part of 
the polyadenylation signal and can affect signal strength and possibly 
regulation.