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Sister chromatid cohesion establishment occurs in concert with lagging strand synthesis.

Rudra S, Skibbens RV - Cell Cycle (2012)

Bottom Line: While the mechanism of cohesion establishment is largely unknown, a popular model is that Ctf7/Eco1 acetylates cohesins encountered by and located in front of the fork.Our genetic and biochemical studies link Ctf7/Eco1 to the Okazaki fragment flap endonuclease, Fen1.We further report genetic and biochemical interactions between Fen1 and the cohesion-associated DNA helicase, Chl1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Lehigh University, Bethlehem, PA, USA.

ABSTRACT
Cohesion establishment is central to sister chromatid tethering reactions and requires Ctf7/Eco1-dependent acetylation of the cohesin subunit Smc3. Ctf7/Eco1 is essential during S phase, and a number of replication proteins (RFC complexes, PCNA and the DNA helicase Chl1) all play individual roles in sister chromatid cohesion. While the mechanism of cohesion establishment is largely unknown, a popular model is that Ctf7/Eco1 acetylates cohesins encountered by and located in front of the fork. In turn, acetylation is posited both to allow fork passage past cohesin barriers and convert cohesins to a state competent to capture subsequent production of sister chromatids. Here, we report evidence that challenges this pre-replicative cohesion establishment model. Our genetic and biochemical studies link Ctf7/Eco1 to the Okazaki fragment flap endonuclease, Fen1. We further report genetic and biochemical interactions between Fen1 and the cohesion-associated DNA helicase, Chl1. These results raise a new model wherein cohesin deposition and establishment occur in concert with lagging strand-processing events and in the presence of both sister chromatids.

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Related in: MedlinePlus

Figure 4. Cohesion establishment coupled to lagging strand processing. Replication fork (Pol = DNA polymerase coupled to PCNA) moves to the left: leading strand replication on the top and lagging strand replication on the bottom (RNA primers shadowed). Immediately behind the fork, PCNA associates with Fen1 (green) and Chl1 (orange). Ctf7/Eco1 (yellow) is not stably recruited to chromatin by any factor, but transiently interacts with chromatin to establish cohesion. Therefore, both cohesin deposition and subsequent cohesion establishment occur behind the replication fork. Cohesins (purple) depicted as unstructured to highlight the many models currently posited in the literature.1,44,45 MCM helicase, primase and RPA not shown (based on Burgers46).
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Figure 4: Figure 4. Cohesion establishment coupled to lagging strand processing. Replication fork (Pol = DNA polymerase coupled to PCNA) moves to the left: leading strand replication on the top and lagging strand replication on the bottom (RNA primers shadowed). Immediately behind the fork, PCNA associates with Fen1 (green) and Chl1 (orange). Ctf7/Eco1 (yellow) is not stably recruited to chromatin by any factor, but transiently interacts with chromatin to establish cohesion. Therefore, both cohesin deposition and subsequent cohesion establishment occur behind the replication fork. Cohesins (purple) depicted as unstructured to highlight the many models currently posited in the literature.1,44,45 MCM helicase, primase and RPA not shown (based on Burgers46).

Mentions: Here, we exploited the well-characterized lagging strand processing factor Fen1 to uncover novel Fen1-Ctf7/Eco1 binding and thus position Ctf7/Eco1 relative to the DNA replication fork. Findings from multiple genetic and biochemical studies indeed support a model in which cohesion establishment occurs immediately behind the fork and possibly coupled to lagging-strand processing.18,27,34 The genetic and physical interactions identified between Fen1, Ctf7/Eco1 and Chl1 (current study and Skibbens24) does not necessarily link Ctf7/Eco1 recruitment to Okazaki fragment maturation. For instance, neither Fen1 nor Chl1 are essential for cell viability20,21,35,36 and thus are unlikely as platforms critical for Ctf7/Eco1 recruitment. Currently, we cannot exclude the possibility that both cooperatively contribute to recruitment—a model consistent with the lethality of cells harboring deletions of both fen1 and chl1 (this study). Instead, we favor a model of cohesion establishment in which Ctf7/Eco1 transiently cycles in associating with cohesins loaded behind the DNA replication fork. We currently view these transient associations as occurring independent of replication fork factors, per se, but that establishment occurs in a post-fork chromatin context that exists in the vicinity of Okazaki fragment maturation as well as on leading-strand synthesis. This post-fork establishment model benefits from several accompanying features (Fig. 4). First, it posits that establishment occurs when sister chromatids are actually present, in contrast to current models that posit establishment requires acetylation of cohesins that reside in front of the fork and before sister chromatids are synthesized.13,16 Second, our model places Ctf7/Eco1 proximal to PCNA but does not require PCNA-dependent chromatin recruitment of Ctf7/Eco1. Nor do we envision Ctf7/Eco1 recruitment requiring Fen1 or Chl1. Instead, we favor a model that Ctf7/Eco1 transiently functions in the local environment that exists immediately behind the replication fork to convert cohesins to a pairing-competent state. Third, this model posits that the cohesins that participate in sister chromatid tethering reactions are most likely loaded behind the DNA replication fork. This is consistent with early reports in which the essential activity of Scc2 (and Ctf7/Eco1) was mapped to S phase.6,8 Thus, both cohesin deposition and Ctf7/Eco1-dependent cohesion establishment likely occur in this post-fork context—cohesin “association” prior to the replication fork being irrelevant to cohesion. This latter point is supported by the highly dynamic nature of cohesins during G1.7,33,37 Our current view of establishment parallels that of chromatinization, in which newly synthesized histone complexes are deposited onto nascent sister chromatid strands and subsequently posttranslationally modified to engender epigenetic states.38 Cohesin deposition/modifications that engender sister chromatin pairing and transcription regulation may be similarly temporally coupled. Toward this end, we note several chromatin-modifying complexes (INO80, RSC and SWI/SNF components) that promote efficient cohesion establishment.2 The physical link reported here between Chl1 and Fen1, coupled with recent findings that Chl1 exhibits 5'-3' unwindase capabilities, removes chromatin-bound proteins (possibly resolving chromatin structures such as G-quads) and is critical for cohesin deposition or stabilization, provides additional motivation in considering a chromatin-based post-fork model of cohesion establishment.39-41 Future research focused on testing cohesion establishment in the context of chromatinization is likely to provide significant insights applicable to multiple fields of inquiry and, given the role of cohesion pathway mutations in tumorigenesis and developmental maladies, may be of clinical interest.5,42


Sister chromatid cohesion establishment occurs in concert with lagging strand synthesis.

Rudra S, Skibbens RV - Cell Cycle (2012)

Figure 4. Cohesion establishment coupled to lagging strand processing. Replication fork (Pol = DNA polymerase coupled to PCNA) moves to the left: leading strand replication on the top and lagging strand replication on the bottom (RNA primers shadowed). Immediately behind the fork, PCNA associates with Fen1 (green) and Chl1 (orange). Ctf7/Eco1 (yellow) is not stably recruited to chromatin by any factor, but transiently interacts with chromatin to establish cohesion. Therefore, both cohesin deposition and subsequent cohesion establishment occur behind the replication fork. Cohesins (purple) depicted as unstructured to highlight the many models currently posited in the literature.1,44,45 MCM helicase, primase and RPA not shown (based on Burgers46).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
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Figure 4: Figure 4. Cohesion establishment coupled to lagging strand processing. Replication fork (Pol = DNA polymerase coupled to PCNA) moves to the left: leading strand replication on the top and lagging strand replication on the bottom (RNA primers shadowed). Immediately behind the fork, PCNA associates with Fen1 (green) and Chl1 (orange). Ctf7/Eco1 (yellow) is not stably recruited to chromatin by any factor, but transiently interacts with chromatin to establish cohesion. Therefore, both cohesin deposition and subsequent cohesion establishment occur behind the replication fork. Cohesins (purple) depicted as unstructured to highlight the many models currently posited in the literature.1,44,45 MCM helicase, primase and RPA not shown (based on Burgers46).
Mentions: Here, we exploited the well-characterized lagging strand processing factor Fen1 to uncover novel Fen1-Ctf7/Eco1 binding and thus position Ctf7/Eco1 relative to the DNA replication fork. Findings from multiple genetic and biochemical studies indeed support a model in which cohesion establishment occurs immediately behind the fork and possibly coupled to lagging-strand processing.18,27,34 The genetic and physical interactions identified between Fen1, Ctf7/Eco1 and Chl1 (current study and Skibbens24) does not necessarily link Ctf7/Eco1 recruitment to Okazaki fragment maturation. For instance, neither Fen1 nor Chl1 are essential for cell viability20,21,35,36 and thus are unlikely as platforms critical for Ctf7/Eco1 recruitment. Currently, we cannot exclude the possibility that both cooperatively contribute to recruitment—a model consistent with the lethality of cells harboring deletions of both fen1 and chl1 (this study). Instead, we favor a model of cohesion establishment in which Ctf7/Eco1 transiently cycles in associating with cohesins loaded behind the DNA replication fork. We currently view these transient associations as occurring independent of replication fork factors, per se, but that establishment occurs in a post-fork chromatin context that exists in the vicinity of Okazaki fragment maturation as well as on leading-strand synthesis. This post-fork establishment model benefits from several accompanying features (Fig. 4). First, it posits that establishment occurs when sister chromatids are actually present, in contrast to current models that posit establishment requires acetylation of cohesins that reside in front of the fork and before sister chromatids are synthesized.13,16 Second, our model places Ctf7/Eco1 proximal to PCNA but does not require PCNA-dependent chromatin recruitment of Ctf7/Eco1. Nor do we envision Ctf7/Eco1 recruitment requiring Fen1 or Chl1. Instead, we favor a model that Ctf7/Eco1 transiently functions in the local environment that exists immediately behind the replication fork to convert cohesins to a pairing-competent state. Third, this model posits that the cohesins that participate in sister chromatid tethering reactions are most likely loaded behind the DNA replication fork. This is consistent with early reports in which the essential activity of Scc2 (and Ctf7/Eco1) was mapped to S phase.6,8 Thus, both cohesin deposition and Ctf7/Eco1-dependent cohesion establishment likely occur in this post-fork context—cohesin “association” prior to the replication fork being irrelevant to cohesion. This latter point is supported by the highly dynamic nature of cohesins during G1.7,33,37 Our current view of establishment parallels that of chromatinization, in which newly synthesized histone complexes are deposited onto nascent sister chromatid strands and subsequently posttranslationally modified to engender epigenetic states.38 Cohesin deposition/modifications that engender sister chromatin pairing and transcription regulation may be similarly temporally coupled. Toward this end, we note several chromatin-modifying complexes (INO80, RSC and SWI/SNF components) that promote efficient cohesion establishment.2 The physical link reported here between Chl1 and Fen1, coupled with recent findings that Chl1 exhibits 5'-3' unwindase capabilities, removes chromatin-bound proteins (possibly resolving chromatin structures such as G-quads) and is critical for cohesin deposition or stabilization, provides additional motivation in considering a chromatin-based post-fork model of cohesion establishment.39-41 Future research focused on testing cohesion establishment in the context of chromatinization is likely to provide significant insights applicable to multiple fields of inquiry and, given the role of cohesion pathway mutations in tumorigenesis and developmental maladies, may be of clinical interest.5,42

Bottom Line: While the mechanism of cohesion establishment is largely unknown, a popular model is that Ctf7/Eco1 acetylates cohesins encountered by and located in front of the fork.Our genetic and biochemical studies link Ctf7/Eco1 to the Okazaki fragment flap endonuclease, Fen1.We further report genetic and biochemical interactions between Fen1 and the cohesion-associated DNA helicase, Chl1.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Lehigh University, Bethlehem, PA, USA.

ABSTRACT
Cohesion establishment is central to sister chromatid tethering reactions and requires Ctf7/Eco1-dependent acetylation of the cohesin subunit Smc3. Ctf7/Eco1 is essential during S phase, and a number of replication proteins (RFC complexes, PCNA and the DNA helicase Chl1) all play individual roles in sister chromatid cohesion. While the mechanism of cohesion establishment is largely unknown, a popular model is that Ctf7/Eco1 acetylates cohesins encountered by and located in front of the fork. In turn, acetylation is posited both to allow fork passage past cohesin barriers and convert cohesins to a state competent to capture subsequent production of sister chromatids. Here, we report evidence that challenges this pre-replicative cohesion establishment model. Our genetic and biochemical studies link Ctf7/Eco1 to the Okazaki fragment flap endonuclease, Fen1. We further report genetic and biochemical interactions between Fen1 and the cohesion-associated DNA helicase, Chl1. These results raise a new model wherein cohesin deposition and establishment occur in concert with lagging strand-processing events and in the presence of both sister chromatids.

Show MeSH
Related in: MedlinePlus