RESUMEN
Mot1 is an essential, conserved, TATA-binding protein (TBP)-associated factor in Saccharomyces cerevisiae with well-established roles in the global control of RNA polymerase II (Pol II) transcription. Previous results have suggested that Mot1 functions exclusively in Pol II transcription, but here we report a novel role for Mot1 in regulating transcription by RNA polymerase I (Pol I). In vivo, Mot1 is associated with the ribosomal DNA, and loss of Mot1 results in decreased rRNA synthesis. Consistent with a direct role for Mot1 in Pol I transcription, Mot1 also associates with the Pol I promoter in vitro in a reaction that depends on components of the Pol I general transcription machinery. Remarkably, in addition to Mot1's role in initiation, rRNA processing is delayed in mot1 cells. Taken together, these results support a model in which Mot1 affects the rate and efficiency of rRNA synthesis by both direct and indirect mechanisms, with resulting effects on transcription activation and the coupling of rRNA synthesis to processing.
Asunto(s)
ADN Helicasas/metabolismo , ARN Ribosómico/biosíntesis , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Factores Asociados con la Proteína de Unión a TATA/metabolismo , Adenosina Trifosfatasas/metabolismo , Cromatina/metabolismo , ADN Ribosómico/ultraestructura , Regulación Fúngica de la Expresión Génica , Genes Fúngicos , Mutación/genética , Regiones Promotoras Genéticas/genética , Transporte de Proteínas , ARN Polimerasa I/metabolismo , Procesamiento Postranscripcional del ARN/genética , ARN Ribosómico/genética , ARN Ribosómico/ultraestructura , Secuencias Repetitivas de Ácidos Nucleicos/genética , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/ultraestructura , Factores de Transcripción/metabolismo , Transcripción GenéticaRESUMEN
Mot1 is an essential Snf2/Swi2-related ATPase and TATA-binding protein (TBP)-associated factor (TAF). In vitro, Mot1 utilizes ATP hydrolysis to disrupt TBP-DNA complexes, but the relationship of this activity to Mot1's in vivo function is unclear. Chromatin immunoprecipitation was used to determine how Mot1 affects the assembly of preinitiation complexes (PICs) at Mot1-controlled promoters in vivo. We find that the Mot1-repressed HSP26 and INO1 promoters are both regulated by TBP recruitment; inactivation of Mot1 leads to increased PIC formation coincident with derepression of transcription. For the Mot1-activated genes BNA1 and URA1, inactivation of Mot1 also leads, remarkably, to increased TBP binding to the promoters, despite the fact that transcription of these genes is obliterated in mot1 cells. In contrast, levels of Taf1, TFIIB, and RNA polymerase II are reduced at Mot1-activated promoters in mot1 cells. These results suggest that Mot1-mediated displacement of TBP underlies its mechanism of repression and activation at these genes. We suggest that at activated promoters, Mot1 disassembles transcriptionally inactive TBP, thereby facilitating the formation of a TBP complex that supports functional PIC assembly.