ParB spreading on DNA requires cytidine triphosphate in vitro

• Published in: eLife 2020-02, Vol.9
• Main Authors: Jalal, Adam Sb, Tran, Ngat T, Le, Tung Bk
• Format: Article
• Language: English
• Subjects: Bacteria; Bacterial genetics; Bacterial Proteins - metabolism; Binding proteins; Binding sites; Caulobacter crescentus; Caulobacter crescentus - metabolism; chromosome organization; chromosome segregation; Chromosomes; Chromosomes and Gene Expression; CTP; Cytidine triphosphate; Cytidine Triphosphate - metabolism; Deoxyribonucleic acid; DNA; DNA Primase - metabolism; DNA, Bacterial - metabolism; DNA-binding protein; DNA-Binding Proteins - metabolism; Hydrolysis; Microbiology and Infectious Disease; ParB-parS; Protein binding; Proteins; Spreading; Transcription; Transcription (Genetics)
• ISSN: 2050-084X; 2050-084X
• DOI: 10.7554/elife.53515

In all living organisms, it is essential to transmit genetic information faithfully to the next generation. The SMC-ParAB- system is widely employed for chromosome segregation in bacteria. A DNA-binding protein ParB nucleates on sites and must associate with neighboring DNA, a process known as spreading, to enable efficient chromosome segregation. Despite its importance, how the initial few ParB molecules nucleating at sites recruit hundreds of further ParB to spread is not fully understood. Here, we reconstitute a -dependent ParB spreading event using purified proteins from and show that CTP is required for spreading. We further show that ParB spreading requires a closed DNA substrate, and a DNA-binding transcriptional regulator can act as a roadblock to attenuate spreading unidirectionally in vitro. Our biochemical reconstitutions recapitulate many observed in vivo properties of ParB and opens up avenues to investigate the interactions between ParB- with ParA and SMC.