Recent studies have shown that the protein CTCF, which plays an important role 
in insulation and in large-scale organization of chromatin within the eukaryotic 
nucleus, depends for both activities on recruitment of the cohesin complex. We 
show here that the interaction of CTCF with the cohesin complex involves direct 
contacts between the cohesin subunit SA2 and specific regions of the C-terminal 
tail of CTCF. All other cohesin components are recruited through their 
interaction with SA2. Expression in vivo of CTCF mutants lacking the C-terminal 
domain, or with mutations at sites within it required for SA2 binding, disrupts 
the normal expression profile of the imprinted genes IGF2-H19 and also results 
in a loss of insulation activity. Taken together, our results demonstrate that 
specific sites on the C terminus of CTCF are essential for cohesin binding and 
insulator function. The only direct interaction between CTCF and cohesin 
involves contact with SA2, which is external to the cohesin ring. This suggests 
that in recruiting cohesin to CTCF, SA2 could bind first and the ring could 
assemble subsequently.