Biology subjects

Branzei, D.

Publications and source records attributed to Branzei, D..

6 recordsLinked to original sources

bioRxiv2025

PDS5 proteins control genome architecture by limiting the lifetime of cohesin-NIPBL complexes

Cohesin-NIPBL complexes extrude genomic DNA into loops that are constrained by CTCF boundaries. This process has important regulatory functions and weakens the separation between euchromatic and heterochromatic compartments. Cohesin can also bind PDS5A or PDS5B, which do not support loop extrusion but are required for the formation of CTCF boundaries. How PDS5 proteins perform this function is unknown. Here we show by in vitro single-molecule imaging that PDS5 proteins stop loop extrusion by facilitating the dissociation of NIPBL from cohesin. Hi-C experiments suggest that this function is required for the establishment of CTCF boundaries in cells. In silico modelling indicates that PDS5 proteins enable the separation between compartments by limiting cohesins velocity and chromatin-residence time. The degree of this compartmentalization depends on the frequency with which chromatin is extruded relative to the time it takes for compartments to form. These results identify PDS5 proteins as key regulators of genome organization. HighlightsO_LIPDS5 proteins stop loop extrusion by facilitating dissociation of NIPBL from cohesin. C_LIO_LIPDS5 proteins strengthen CTCF boundaries by limiting the lifetime of cohesin-NIPBL. C_LIO_LIPDS5 proteins regulate compartmentalization. C_LIO_LICompartmentalization is governed by polymer relaxation and loop extrusion dynamics. C_LI

molecular biology↗

bioRxiv2022

FANCJ DNA helicase is recruited to the replisome by AND-1 to ensure genome stability

FANCJ is a DNA helicase linked to Fanconi anemia and frequently mutated in breast and ovarian cancers. If and how FANCJ is recruited to the replisome is unknown. Here, we report that FANCJ directly binds to AND-1 (the vertebrate ortholog of budding yeast Ctf4), a homo-trimeric protein adaptor that connects the CDC45/MCM2-7/GINS replicative DNA helicase with DNA polymerase and several factors at DNA replication forks. We find that the interaction between FANCJ and AND-1 requires the integrity of an evolutionarily conserved Ctf4-interacting protein (CIP) box located between the FANCJ helicase motifs IV and V. Disruption of the FANCJ CIP box significantly reduces FANCJ association with the replisome, causing enhanced DNA damage, decreased replication fork recovery and fork asymmetry in stressful conditions. Cancer-relevant FANCJ CIP box variants display reduced AND-1-binding, a finding that suggests a potential role of the mutated FANCJ alleles in cancer predisposition.

molecular biology↗

bioRxiv2022

PCNA recruits cohesin loader Scc2/NIPBL to ensure sister chromatid cohesion

Sister chromatid cohesion is essential for faithful chromosome segregation and genome duplication during cell division. Failure to establish cohesion during S phase by the ring-shaped multiprotein complex cohesin leads to genomic instability1-4. Replisome-associated proteins are required to generate cohesion by two independent pathways5. One mediates conversion of cohesins bound to unreplicated DNA ahead of replication forks into cohesive entities behind them, while the second promotes cohesin de novo loading onto newly-replicated DNAs6. The latter process depends on the cohesin loader Scc2/NIPBL and the alternative PCNA loader CTF18-RFC. However, the precise mechanism of de novo cohesin loading during replication is unknown. Here we show that PCNA physically recruits yeast cohesin loader Scc2 via its C-terminal PCNA-interacting protein motif. Binding to PCNA is crucial, as scc2-pip mutant deficient in Scc2-PCNA interaction is defective in cohesion when combined with replisome mutants of the cohesin conversion pathway. Moreover, scc2-pip mutant becomes inviable without its partner Scc4/MAU2 that localizes cohesin loader to centromeres. Importantly, the role of NIPBL recruitment to PCNA for cohesion generation is conserved in vertebrate cells. Our results demonstrate that PCNA, the maestro of replication-linked functions, is also crucially involved in the cohesion establishment through de novo cohesin loading onto replicated DNA.

cell biology↗

bioRxiv2021

Structures of core eukaryotic protein complexes

Protein-protein interactions play critical roles in biology, but despite decades of effort, the structures of many eukaryotic protein complexes are unknown, and there are likely many interactions that have not yet been identified. Here, we take advantage of recent advances in proteome-wide amino acid coevolution analysis and deep-learning-based structure modeling to systematically identify and build accurate models of core eukaryotic protein complexes, as represented within the Saccharomyces cerevisiae proteome. We use a combination of RoseTTAFold and AlphaFold to screen through paired multiple sequence alignments for 8.3 million pairs of S. cerevisiae proteins and build models for strongly predicted protein assemblies with two to five components. Comparison to existing interaction and structural data suggests that these predictions are likely to be quite accurate. We provide structure models spanning almost all key processes in Eukaryotic cells for 104 protein assemblies which have not been previously identified, and 608 which have not been structurally characterized. One-sentence summaryWe take advantage of recent advances in proteome-wide amino acid coevolution analysis and deep-learning-based structure modeling to systematically identify and build accurate models of core eukaryotic protein complexes.

biochemistry↗

bioRxiv2020

SMC Complexes Are Guarded by the SUMO Protease Ulp2 Against SUMO-Chain-Mediated Turnover

Structural maintenance of chromosomes (SMC) complexes, cohesin, condensin and Smc5/6, are essential for viability and participate in multiple processes, including sister chromatid cohesion, chromosome condensation, and DNA repair. Here we show that SUMO chains target all three SMC complexes and are antagonized by the SUMO protease Ulp2 to prevent their turnover. We uncover that the essential role of the cohesin-associated subunit Pds5 is to counteract SUMO chains jointly with Ulp2. Importantly, fusion of Ulp2 to kleisin Scc1 supports viability of PDS5 null cells and protects cohesin from proteasomal degradation mediated by the SUMO-targeted ubiquitin ligase Slx5/Slx8. The lethality of PDS5 deleted cells can also be bypassed by simultaneous loss of the PCNA unloader, Elg1, and the cohesin releaser, Wpl1, but only when Ulp2 is functional. Condensin and Smc5/6 complex are similarly guarded by Ulp2 against unscheduled SUMO-chain assembly, which we propose to time the availability of SMC complexes on chromatin.

cell biology↗

bioRxiv2019

Timeless couples G quadruplex detection with processing by DDX11 during DNA replication

Regions of the genome with the potential to form secondary structure pose a frequent and significant impediment to DNA replication and must be actively managed in order to preserve genetic and epigenetic integrity. The fork protection complex (FPC), a conserved group of replisome-associated proteins including Timeless, Tipin, and Claspin, plays an important role in maintaining efficient replisome activation, ensuring optimum fork rates, sister chromatid cohesion and checkpoint function. It also helps maintain the stability of sequences prone to secondary structure formation through an incompletely understood mechanism. Here, we report a previously unappreciated DNA binding domain in the C-terminus of Timeless, which exhibits specific binding to G quadruplex (G4) structures. We show that, in vivo, both the C-terminus of Timeless and the DDX11 helicase act collaboratively to ensure processive replication of G4 structures to prevent genetic and epigenetic instability.

molecular biology↗