ABSTRACT
Structural maintenance of chromosomes (SMC) protein complexes structure genomes by extruding DNA loops, but the molecular mechanism that underlies their activity has remained unknown. We show that the active condensin complex entraps the bases of a DNA loop transiently in two separate chambers. Single-molecule imaging and cryo-electron microscopy suggest a putative power-stroke movement at the first chamber that feeds DNA into the SMC-kleisin ring upon adenosine triphosphate binding, whereas the second chamber holds on upstream of the same DNA double helix. Unlocking the strict separation of "motor" and "anchor" chambers turns condensin from a one-sided into a bidirectional DNA loop extruder. We conclude that the orientation of two topologically bound DNA segments during the SMC reaction cycle determines the directionality of DNA loop extrusion.
Subject(s)
Adenosine Triphosphatases , DNA-Binding Proteins , DNA , Multiprotein Complexes , Adenosine Triphosphatases/chemistry , Cryoelectron Microscopy , DNA/chemistry , DNA-Binding Proteins/chemistry , Multiprotein Complexes/chemistry , Nucleic Acid Conformation , Single Molecule ImagingABSTRACT
Complexes containing a pair of structural maintenance of chromosomes (SMC) family proteins are fundamental for the three-dimensional (3D) organization of genomes in all domains of life. The eukaryotic SMC complexes cohesin and condensin are thought to fold interphase and mitotic chromosomes, respectively, into large loop domains, although the underlying molecular mechanisms have remained unknown. We used cryo-EM to investigate the nucleotide-driven reaction cycle of condensin from the budding yeast Saccharomyces cerevisiae. Our structures of the five-subunit condensin holo complex at different functional stages suggest that ATP binding induces the transition of the SMC coiled coils from a folded-rod conformation into a more open architecture. ATP binding simultaneously triggers the exchange of the two HEAT-repeat subunits bound to the SMC ATPase head domains. We propose that these steps result in the interconversion of DNA-binding sites in the catalytic core of condensin, forming the basis of the DNA translocation and loop-extrusion activities.
Subject(s)
Carrier Proteins/chemistry , Chromosomal Proteins, Non-Histone/chemistry , Nuclear Proteins/chemistry , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae/chemistry , Adenosine Triphosphatases/chemistry , Adenosine Triphosphatases/metabolism , Adenosine Triphosphatases/ultrastructure , Adenosine Triphosphate/metabolism , Carrier Proteins/metabolism , Carrier Proteins/ultrastructure , Cell Cycle Proteins , Chromosomal Proteins, Non-Histone/metabolism , Chromosomal Proteins, Non-Histone/ultrastructure , Cryoelectron Microscopy , DNA-Binding Proteins/chemistry , DNA-Binding Proteins/metabolism , DNA-Binding Proteins/ultrastructure , Models, Molecular , Multiprotein Complexes/chemistry , Multiprotein Complexes/metabolism , Multiprotein Complexes/ultrastructure , Nuclear Proteins/metabolism , Nuclear Proteins/ultrastructure , Protein Conformation , Protein Folding , Protein Multimerization , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae Proteins/ultrastructureABSTRACT
Condensin is a conserved SMC complex that uses its ATPase machinery to structure genomes, but how it does so is largely unknown. We show that condensin's ATPase has a dual role in chromosome condensation. Mutation of one ATPase site impairs condensation, while mutating the second site results in hyperactive condensin that compacts DNA faster than wild-type, both in vivo and in vitro. Whereas one site drives loop formation, the second site is involved in the formation of more stable higher-order Z loop structures. Using hyperactive condensin I, we reveal that condensin II is not intrinsically needed for the shortening of mitotic chromosomes. Condensin II rather is required for a straight chromosomal axis and enables faithful chromosome segregation by counteracting the formation of ultrafine DNA bridges. SMC complexes with distinct roles for each ATPase site likely reflect a universal principle that enables these molecular machines to intricately control chromosome architecture.
Subject(s)
Adenosine Triphosphatases/metabolism , Chromatin Assembly and Disassembly/physiology , DNA-Binding Proteins/metabolism , Multiprotein Complexes/metabolism , Adenosine Triphosphatases/genetics , Adenosine Triphosphatases/physiology , Adenosine Triphosphate/chemistry , Binding Sites/genetics , Binding Sites/physiology , Cell Cycle Proteins/metabolism , Cell Line, Tumor , Chromatin/physiology , Chromosomal Proteins, Non-Histone/metabolism , Chromosomes/metabolism , Chromosomes/physiology , DNA/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/physiology , Humans , Multiprotein Complexes/physiology , Protein Binding/physiology , Protein Subunits/metabolism , CohesinsABSTRACT
The condensin protein complex plays a key role in the structural organization of genomes. How the ATPase activity of its SMC subunits drives large-scale changes in chromosome topology has remained unknown. Here we reconstruct, at near-atomic resolution, the sequence of events that take place during the condensin ATPase cycle. We show that ATP binding induces a conformational switch in the Smc4 head domain that releases its hitherto undescribed interaction with the Ycs4 HEAT-repeat subunit and promotes its engagement with the Smc2 head into an asymmetric heterodimer. SMC head dimerization subsequently enables nucleotide binding at the second active site and disengages the Brn1 kleisin subunit from the Smc2 coiled coil to open the condensin ring. These large-scale transitions in the condensin architecture lay out a mechanistic path for its ability to extrude DNA helices into large loop structures.