Transcriptional repression at the silent yeast mating type loci is ach
ieved through the formation of a particular nucleoprotein complex at s
pecific cis-acting elements called silencers. This complex in turn app
ears to initiate the spreading of a histone binding protein complex in
to the surrounding chromatin, which restricts accessibility of the reg
ion to the transcription machinery, We have investigated long-range, c
ooperative effects between silencers by studying the repression of a r
eporter gene integrated at the HML locus flanked by various combinatio
ns of wild-type and mutated silencer sequences, Two silencers can coop
erate over >4000 bp to repress transcription efficiently. More importa
ntly, a single binding site for either the repressor activator protein
1 (Rap1), the autonomous replicating sequence (ARS) binding factor 1
(Abf1) or the origin recognition complex (ORC) can enhance the action
of a distant silencer without acting as a silencer on its own. Functio
nal cooperativity is demonstrated using a quantitative assay for repre
ssion, and varies with the affinity of the binding sites used, Since t
he repression mechanism is Sir dependent, the Rap1, ORC and/or Abf1 pr
oteins bound to distant I)NB elements may interact to create an interf
ace of sufficiently high affinity such that Sir-containing complexes b
ind, nucleating the silent chromatin state.