SummaryHuntington’s disease is caused by an abnormally long polyglutamine tract in the huntingtin protein. This leads to the generation and deposition of N-terminal exon1 fragments of the protein in intracellular aggregates. We combined electron tomography and quantitative fluorescence microscopy to analyze the structural and material properties of huntingtin exon1 assemblies in mammalian cells, in yeast, and in vitro. We found that huntingtin exon1 proteins can form reversible liquid-like assemblies, a process driven by huntingtin’s polyQ tract and proline-rich region. In cells and in vitro, the liquid-like assemblies converted to solid-like assemblies with a fibrillar structure. Intracellular phase transitions of polyglutamine proteins could play a role in initiating irreversible pathological aggregation.

SummaryIn silico analysis of the Bifidobacterium breve UCC2003 genome predicted two distinct loci, which encode three different restriction/modification systems, each comprising a modification methylase and a restriction endonuclease. Based on sequence homology and observed protection against restriction we conclude that the first restriction endonuclease, designated BbrI, is an isoschizomer of BbeI, the second, BbrII, is a neoschizomer of SalI, while the third, BbrIII, is an isoschizomer of PstI. Expression of each of the B. breve UCC2003 methylase‐encoding genes in B. breve JCM 7017 established that BbrII and BbrIII are active and restrict incoming DNA. By exploiting knowledge on restriction/modification in B. breve UCC2003 we successfully increased the transformation efficiency to a level that allows the reliable generation of mutants by homologous recombination using a non‐replicative plasmid.

A novel restriction-modification system, designated LlaJI, was identified on pNP40, a naturally occurring 65-kb plasmid from Lactococcus lactis. The system comprises four adjacent similarly oriented genes that are predicted to encode two m 5 C methylases and two restriction endonucleases. The LlaJI system, when cloned into a low-copy-number vector, was shown to confer resistance against representatives of the three most common lactococcal phage species. This phage resistance phenotype was found to be strongly temperature dependent, being most effective at 19°C. A functional analysis confirmed that the predicted methylase-encoding genes, llaJIM1 and llaJIM2, were both required to mediate complete methylation, while the assumed restriction enzymes, specified by llaJIR1 and llaJIR2, were both necessary for the complete restriction phenotype. A Northern blot analysis revealed that the four LlaJI genes are part of a 6-kb operon and that the relative abundance of the LlaJI-specific mRNA in the cells does not appear to contribute to the observed temperaturesensitive profile. This was substantiated by use of a LlaJI promoter-lacZ fusion, which further revealed that the LlaJI operon appears to be subject to transcriptional regulation by an as yet unidentified element(s) encoded by pNP40.Restriction-modification (R/M) systems are the most abundant bacteriophage resistance mechanism found in bacteria thus far. These systems are classified into four groups (designated type I, II, III, or IV) on the basis of their cofactor requirements, subunit composition, recognition sequence structure, and cleavage site relative to the recognition sequence (51). Members of type I are composed of the products of three distinct structural genes, which encode the subunits of a hetero-oligomeric enzyme complex that is required for restriction (R), modification (M), and specificity (S). Type I enzymes are present in two different oligomeric forms in vivo. The M 2 S oligomeric enzyme is capable of methylation in the presence of S-adenosylmethionine (AdoMet) and Mg 2ϩ , while the R 2 M 2 S oligomer is responsible for the restriction of unmethylated DNA in the presence of ATP, AdoMet, and Mg 2ϩ . Type I system representatives cleave target DNAs at sites that are distant from an asymmetrical recognition sequence. A model for cleavage by type I endonucleases has been proposed by Bourniquel and Bickle (4), who suggested that the endonuclease binds to its unmethylated recognition site, forming a dimer of two R 2 M 2 S oligomers. In the presence of ATP, all four HsdR subunits of this dimer complex promote the independent translocation of DNA directed toward the bound complex, and cleavage occurs when a physical barrier forces the translocation process to stop.A typical R/M system belonging to type II, which is the simplest group with respect to genetic structure and cofactor requirements, is composed of two distinct gene products, one of which acts as a Mg 2ϩ -dependent sequence-specific endonuclease (REase) while the other functions as a cognate AdoM...