The Origin Recognition Complex and Sir4 Protein Recruit Sir1p to Yeast Silent Chromatin through Independent Interactions Requiring a Common Sir1p Domain
Abstract:Sir1p is one of four SIR (silent information regulator) proteins required for silencing the cryptic mating-type locus HMRa in the budding yeast Saccharomyces cerevisiae. A Sir1p interaction with Orc1p, the largest subunit of the origin recognition complex (ORC), is critical for Sir1p's ability to bind HMRa and function in the formation of silent chromatin. Here we show that a discrete domain within Sir1p, the ORC interaction region (OIR), was necessary and sufficient for a Sir1p-ORC interaction. The OIR contai… Show more
“…Direct physical interaction between Sir1p and ORC is critical for establishing silent chromatin at HMR and HML in S. cerevisiae (7,10,11). Previous studies identified the minimal OIR of Sir1p and amino acids within the OIR as important for the Sir1p͞ORC interaction (8,11) (Fig.…”
Section: Resultsmentioning
confidence: 99%
“…The crystal structure of the S. cerevisiae Orc1p-BAH domain has been determined and serves as the guide for modeling other BAH domains (10). The domain of Sir1p required for the Sir1p͞ORC interaction, the ORC interaction region (OIR) (11), is the only molecularly defined BAH-domain-binding partner and thus is key for understanding the function and mechanisms of BAH-domaincontaining proteins and their binding partners in regulating chromatin structure.…”
mentioning
confidence: 99%
“…Analyses of HMR-E, a silencer that is necessary and sufficient to target silent chromatin to HMR, has established that the primary role for ORC in silencing is to recruit Sir1p to the silencer through direct protein-protein interactions between the Orc1p subunit of ORC and Sir1p (6)(7)(8)(9)(10)(11). ORC-bound Sir1p nucleates and͞or stabilizes a silent chromatin domain through direct protein-protein interactions with other Sir proteins, in particular Sir4p (7,11). Thus, formation of the Sir1p͞ORC interaction is a critical step in establishing a repressive chromatin structure at HMR (12).…”
The Sir1 protein plays a key role in establishing a silent chromatin structure at the cryptic mating-type loci HMR and HML in Saccharomyces cerevisiae by interacting with the bromo-adjacent homology (BAH) domain of the Orc1p subunit of the origin recognition complex (ORC). Here, we present the high-resolution crystal structures of the ORC interaction region (OIR) of Sir1p and that of the complex formed between the OIR and BAH domains. Amino acids within the OIR previously shown to be required for a Sir1p͞ORC interaction are presented on a conserved, convex surface that forms a complementary interface with a concave region of the Orc1 BAH domain that is critical for transcriptional silencing. The OIR͞ BAH interaction surface comprises a network of hydrophobic and polar͞ionic interactions between discrete structural modules in each protein and involves several residues that were not implicated in previous studies. These data provide important structural insights into a protein-protein interaction critical for the formation of a specialized chromatin domain within eukaryotic chromosomes.chromatin ͉ silencers ͉ yeast ͉ chromosomes ͉ replication
“…Direct physical interaction between Sir1p and ORC is critical for establishing silent chromatin at HMR and HML in S. cerevisiae (7,10,11). Previous studies identified the minimal OIR of Sir1p and amino acids within the OIR as important for the Sir1p͞ORC interaction (8,11) (Fig.…”
Section: Resultsmentioning
confidence: 99%
“…The crystal structure of the S. cerevisiae Orc1p-BAH domain has been determined and serves as the guide for modeling other BAH domains (10). The domain of Sir1p required for the Sir1p͞ORC interaction, the ORC interaction region (OIR) (11), is the only molecularly defined BAH-domain-binding partner and thus is key for understanding the function and mechanisms of BAH-domaincontaining proteins and their binding partners in regulating chromatin structure.…”
mentioning
confidence: 99%
“…Analyses of HMR-E, a silencer that is necessary and sufficient to target silent chromatin to HMR, has established that the primary role for ORC in silencing is to recruit Sir1p to the silencer through direct protein-protein interactions between the Orc1p subunit of ORC and Sir1p (6)(7)(8)(9)(10)(11). ORC-bound Sir1p nucleates and͞or stabilizes a silent chromatin domain through direct protein-protein interactions with other Sir proteins, in particular Sir4p (7,11). Thus, formation of the Sir1p͞ORC interaction is a critical step in establishing a repressive chromatin structure at HMR (12).…”
The Sir1 protein plays a key role in establishing a silent chromatin structure at the cryptic mating-type loci HMR and HML in Saccharomyces cerevisiae by interacting with the bromo-adjacent homology (BAH) domain of the Orc1p subunit of the origin recognition complex (ORC). Here, we present the high-resolution crystal structures of the ORC interaction region (OIR) of Sir1p and that of the complex formed between the OIR and BAH domains. Amino acids within the OIR previously shown to be required for a Sir1p͞ORC interaction are presented on a conserved, convex surface that forms a complementary interface with a concave region of the Orc1 BAH domain that is critical for transcriptional silencing. The OIR͞ BAH interaction surface comprises a network of hydrophobic and polar͞ionic interactions between discrete structural modules in each protein and involves several residues that were not implicated in previous studies. These data provide important structural insights into a protein-protein interaction critical for the formation of a specialized chromatin domain within eukaryotic chromosomes.chromatin ͉ silencers ͉ yeast ͉ chromosomes ͉ replication
“…Interestingly, an eso mutant of Sir2, which has no obvious connection to interaction with Sir4, is also located in the Sir4-interacting region of Sir2 (Garcia and Pillus 2002). It is known that establishment of a silenced chromatin domain in the mating type loci involves a cascade of protein-protein interactions, including the binding of Sir1 to the silencerbound ORC1 and the recruitment of the Sir2-Sir4 complex by the interaction between Sir1 and Sir4 (Triolo and Sternglanz 1996;Bose et al 2004;Hou et al 2005;Hsu et al 2005). Thus, a weakened or disrupted Sir2-Sir4 interaction can undermine the Sir1 function of recruiting a histone deacetylase to the targeted chromatin region, and our discovery that a Sir2 mutant affecting Sir1 function is located in the Sir4 interaction region is testimony to the network of protein-protein interaction required for transcriptional silencing in yeast.…”
The budding yeast Sir2 (silent information regulator 2) protein is the founding member of the sirtuin family of NAD-dependent histone/protein deacetylases. Its function in transcriptional silencing requires both the highly conserved catalytic domain and a poorly understood N-terminal regulatory domain (Sir2N). We determined the structure of Sir2 in complex with a fragment of Sir4, a component of the transcriptional silencing complex in Saccharomyces cerevisiae. The structure shows that Sir4 is anchored to Sir2N and contacts the interface between the Sir2N and the catalytic domains through a long loop. We discovered that the interaction between the Sir4 loop and the interdomain interface in Sir2 is critical for allosteric stimulation of the deacetylase activity of Sir2. These results bring to light the structure and function of the regulatory domain of Sir2, and the knowledge should be useful for understanding allosteric regulation of sirtuins in general.
“…ORC1 plays a key role in this process (Figure 1a). ORC1 interacts with the SIR1 protein and recruits the remaining components of the SIR complex including histone deacetylase SIR2 to the chromatin [7,27–30]. 10.1080/19491034.2018.1516484-F0001Figure 1.Chromatin related functions of ORC in different species.…”
Section: Heterochromatin Formation and Transcription Regulationmentioning
Origin recognition complex (ORC), a heteromeric six-subunit complex, is the central component of the eukaryotic pre-replication complex. Recent data from yeast, frogs, flies and mammals present compelling evidence that ORC and its individual subunits have nonreplicative functions as well. The majority of these functions, such as heterochromatin formation, chromosome condensation, and segregation are dependent on ORC-DNA interactions. Furthermore, ORC is involved in the control of cell division via its participation in centrosome duplication and cytokinesis. Recent findings have also demonstrated a direct interaction between ORC and mRNPs and highlighted an essential role of ORC in mRNA nuclear export. Along with the growth of evolutionary complexity of organisms, ORC complex functions become more elaborate and new functions of the ORC sub-complexes and individual subunits have emerged.
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