PUF proteins are a conserved family of eukaryotic RNA-binding proteins that 
regulate specific mRNAs: they control many processes including stem cell 
proliferation, fertility, and memory formation. PUFs repress protein expression 
from their target mRNAs but the mechanism by which they do so remains unclear, 
especially for humans. Humans possess two PUF proteins, PUM1 and PUM2, which 
exhibit similar RNA binding specificities. Here we report new insights into 
their regulatory activities and mechanisms of action. We developed functional 
assays to measure sequence-specific repression by PUM1 and PUM2. Both robustly 
inhibit translation and promote mRNA degradation. Purified PUM complexes were 
found to contain subunits of the CCR4-NOT (CNOT) complex, which contains 
multiple enzymes that catalyze mRNA deadenylation. PUMs interact with the CNOT 
deadenylase subunits in vitro. We used three approaches to determine the 
importance of deadenylases for PUM repression. First, dominant-negative mutants 
of CNOT7 and CNOT8 reduced PUM repression. Second, RNA interference depletion of 
the deadenylases alleviated PUM repression. Third, the poly(A) tail was 
necessary for maximal PUM repression. These findings demonstrate a conserved 
mechanism of PUF-mediated repression via direct recruitment of the CCR4-POP2-NOT 
deadenylase leading to translational inhibition and mRNA degradation. A second, 
deadenylation independent mechanism was revealed by the finding that PUMs 
repress an mRNA that lacks a poly(A) tail. Thus, human PUMs are repressors 
capable of deadenylation-dependent and -independent modes of repression.