Biology subjects

Bjorkegren, C.

Publications and source records attributed to Bjorkegren, C..

4 recordsLinked to original sources

bioRxiv2023

Loop-extruding Smc5/6 organizes transcription-induced positive DNA supercoils

The Structural Maintenance of Chromosome (SMC) protein complexes cohesin, condensin and the Smc5/6 complex (Smc5/6) are essential for chromosome function. At the molecular level, these complexes fold DNA by loop extrusion. Accordingly, cohesin creates chromosome loops in interphase, and condensin compacts mitotic chromosomes. However, the role of Smc5/6s recently discovered DNA loop extrusion activity is unknown. Here, we uncover that Smc5/6 associates with transcription-induced positively supercoiled chromosomal DNA at cohesin-dependent chromosome loop boundaries. Mechanistically, single-molecule imaging reveals that dimers of Smc5/6 specifically recognize the tip of positively supercoiled DNA plectonemes, and efficiently initiates loop extrusion to gather the supercoiled DNA into a large plectonemic loop. Finally, Hi-C analysis shows that Smc5/6 links chromosomal regions containing transcription-induced positive supercoiling in cis. Altogether, our findings indicate that Smc5/6 controls the three-dimensional organization of chromosomes by recognizing and initiating loop extrusion on positively supercoiled DNA.

molecular biology↗

bioRxiv2022

A system for inducible mitochondria-specific protein degradation in vivo

Targeted protein degradation systems developed for eukaryotes employ cytoplasmic machineries to perform proteolysis. This has prevented mitochondria-specific analysis of genome maintaining proteins that localize to both mitochondria and nucleus. Here, we present an inducible mitochondria-specific protein degradation system in Saccharomyces cerevisiae based on the Mesoplasma florum Lon (mf-Lon) protease and its corresponding ssrA tag (called PDT). We show that mitochondrially targeted mf-Lon protease efficiently and selectively degrades a PDT-tagged reporter protein localized to the mitochondrial matrix. The degradation can be induced by depleting adenine from the medium and tuned by altering the promoter strength of the MF-LON gene. Finally, we demonstrate that mf-Lon degrades endogenous, dually localized proteins inside mitochondria. In summary, our system is an efficient tool for analysis of intricate mitochondria-nuclear crosstalk essential for proper mitochondrial function. One-Sentence SummaryMitochondria-specific protein degradation of dually localized proteins

cell biology↗

bioRxiv2022

The Smc5/6 complex is a DNA loop extruding motor

Structural Maintenance of Chromosomes (SMC) protein complexes are essential for the spatial organization of chromosomes. While cohesin and condensin organize chromosomes by extruding DNA loops, the molecular functions of the third eukaryotic SMC complex, Smc5/6, remain largely unknown. Using single-molecule imaging, we reveal that Smc5/6 forms DNA loops by extrusion. Upon ATP-hydrolysis, Smc5/6 symmetrically reels DNA into loops at a force-dependent rate of 1 kilobase pairs per second. Smc5/6 extrudes loops in the form of a dimer, while monomeric Smc5/6 unidirectionally translocate along DNA. We also find that Nse5 and Nse6 (Nse5/6) subunits act as negative regulators of Smc5/6-mediated loop initiation and stability. Our findings reveal Smc5/6s molecular functions, and establish loop extrusion as a conserved mechanism among eukaryotic SMC complexes. One-Sentence SummarySmc5/6 is a DNA-loop-extruding motor, establishing loop extrusion as a conserved mechanism among eukaryotic SMC complexes.

biophysics↗

bioRxiv2018

14-3-3 proteins activate Pseudomonas exotoxins-S and -T by chaperoning a hydrophobic surface

Pseudomonas are a common cause of hospital acquired infections that may be lethal. ADP-ribosyltransferase activities of Pseudomonas exotoxin-S and -T depend on 14-3-3 proteins inside the host cell. By binding in the 14-3-3 phosphopeptide binding groove, a hydrophobic C-terminal helix of ExoS and ExoT has been thought to be crucial for their activation. However, crystal structures of the 14-3-3{beta}:ExoS and -ExoT complexes presented here reveal an extensive novel binding interface that is sufficient for complex formation and toxin activation. We show that C-terminally truncated ExoS ADP-ribosyltransferase domain lacking the hydrophobic binding motif is active when co-expressed with 14-3-3. Moreover, swapping the hydrophobic C-terminus with a fragment from Vibrio Vis toxin creates a 14-3-3 independent toxin that ADP-ribosylates known ExoS targets. Finally, we show that 14-3-3 stabilizes ExoS against thermal aggregation. Together, this indicates that 14-3-3 proteins activate exotoxin ADP-ribosyltransferase domains by chaperoning their hydrophobic surfaces independently of the hydrophobic C-terminal segment.\n\nShort summaryCrystal structures of Pseudomonas exotoxins-S and -T identify a novel hydrophobic interface with 14-3-3 proteins, and we show that 14-3-3 activates these toxins independent of their phosphopeptide groove binding C-termini, by preventing their aggregation.

biochemistry↗