Protein compactness and interaction valency define the architecture of a biomolecular condensate across scales.

Non-membrane-bound biomolecular condensates have been proposed to represent an important mode of subcellular organization in diverse biological settings. However, the fundamental principles governing the spatial organization and dynamics of condensates at the atomistic level remain unclear. The Lge1 protein is required for histone H2B ubiquitination and its N-terminal intrinsically disordered fragment (Lge1) undergoes robust phase separation.

Phase separation directs ubiquitination of gene-body nucleosomes.

The conserved yeast E3 ubiquitin ligase Bre1 and its partner, the E2 ubiquitin-conjugating enzyme Rad6, monoubiquitinate histone H2B across gene bodies during the transcription cycle. Although processive ubiquitination might-in principle-arise from Bre1 and Rad6 travelling with RNA polymerase II, the mechanism of H2B ubiquitination across genic nucleosomes remains unclear. Here we implicate liquid-liquid phase separation as the underlying mechanism.

The COMA complex interacts with Cse4 and positions Sli15/Ipl1 at the budding yeast inner kinetochore.

Kinetochores are macromolecular protein complexes at centromeres that ensure accurate chromosome segregation by attaching chromosomes to spindle microtubules and integrating safeguard mechanisms. The inner kinetochore is assembled on CENP-A nucleosomes and has been implicated in establishing a kinetochore-associated pool of Aurora B kinase, a chromosomal passenger complex (CPC) subunit, which is essential for chromosome biorientation. By performing crosslink-guided in vitro reconstitution of budding yeast kinetochore complexes we showed that the Ame1/Okp1 heterodimer, which forms the COMA complex with Ctf19/Mcm21, selectively bound Cse4 nucleosomes through the Cse4 N-terminus.

Structural mechanism for the recognition and ubiquitination of a single nucleosome residue by Rad6-Bre1.

Cotranscriptional ubiquitination of histone H2B is key to gene regulation. The yeast E3 ubiquitin ligase Bre1 (human RNF20/40) pairs with the E2 ubiquitin conjugating enzyme Rad6 to monoubiquitinate H2B at Lys123. How this single lysine residue on the nucleosome core particle (NCP) is targeted by the Rad6-Bre1 machinery is unknown.

Monoubiquitination of histone H2B is intrinsic to the Bre1 RING domain-Rad6 interaction and augmented by a second Rad6-binding site on Bre1.

Ubiquitin signaling on chromatin is linked to diverse aspects of genome regulation, including gene expression and DNA repair. The yeast RING E3 ligase Bre1 combines with the E2 Rad6 to monoubiquitinate histone H2B during transcription. Little is known about how Bre1 directs Rad6 toward transferring only a single ubiquitin to a specific lysine residue.

New methods for the isolation and characterization of biofilm-persistent mutants in Pseudomonas putida.

Here we describe two new methods for the genetic characterization of bacterial biofilm development. First, we have designed a microtitre dish-based approach for high-throughput screening of Pseudomonas putida mutants showing increased biofilm under dispersal conditions. Using this method, nine such biofilm-persistent mutants, bearing transposon insertions in four loci: lapG, bifA, mvaB and dksA, were isolated.