The conversion of male germ cell chromatin to a nucleoprotamine structure is fundamental to the life cycle, yet the underlying molecular details remain obscure. Here we show that an essential step is the genome-wide incorporation of TH2B, a histone H2B variant of hitherto unknown function. Using mouse models in which TH2B is depleted or C-terminally modified, we show that TH2B directs the final transformation of dissociating nucleosomes into protamine-packed structures. Depletion of TH2B induces compensatory mechanisms that permit histone removal by up-regulating H2B and programming nucleosome instability through targeted histone modifications, including lysine crotonylation and arginine methylation. Furthermore, after fertilization, TH2B reassembles onto the male genome during protamine-to-histone exchange. Thus, TH2B is a unique histone variant that plays a key role in the histone-to-protamine packing of the male genome and guides genome-wide chromatin transitions that both precede and follow transmission of the male genome to the egg.

Protein lysine acetylation plays a key role in regulating chromatin dynamics, gene expression and metabolic pathways in eukaryotes, and, thus, contributes to diverse cellular processes like transcription, cell cycle regulation, and apoptosis. Although recent evidence suggests that acetylated proteins impact broadly cellular functions in prokaryotes, the substrates and localization of this modification remain widely unknown due to the limitations of analytical methods. Comprehensive identification of protein acetylation is a major bottleneck due to its dynamic property and pretty low abundance. A complete atlas of acetylome will significantly advance our understanding of this modification functions in prokaryotes. To achieve this goal, we have developed an intergraded approach to identifying lysine acetylation. Combining immunoaffinity enrichment with high sensitive mass spectrometry, we identified 349 acetylated proteins and addressed 1070 acetylation sites in Escherichia coli. To our knowledge, the acetylated proteins and acetylated sites were increased to 3 times and 8 times, respectively, compared to that in previous report. To further characterize this modification, we classified acetylated proteins into several groups according to cell components, molecular functions and biological process. Additionally, interaction networks and high confident domains architectures of acetylated proteins were investigated with the aid of bioinformatics tools. Finally, the acetylated metabolic enzymes were analyzed on the basis of acetylated proteins identified by proteomic survey in E. coli. Our study has demonstrated that the combined approach is powerful for identification and characterization of protein lysine acetylation on a large scale. These results not only greatly expand the number of acetylated proteins, but also provide a series of important information including localization, networks and characterization of acetylome.

Lysine 2-hydroxyisobutyrylation (Khib) has recently been shown to be an evolutionarily conserved histone mark. Here, we report that CobB serves as a lysine de-2-hydroxyisobutyrylation enzyme that regulates glycolysis and cell growth in prokaryotes. We identified the specific binding of CobB to Khib using a novel self-assembled multivalent photocrosslinking peptide probe and demonstrated that CobB can catalyze lysine de-2-hydroxyisobutyrylation both in vivo and in vitro. R58 of CobB is a critical site for its de-2-hydroxyisobutyrylase activity. Using a quantitative proteomics approach, we identified 99 endogenous substrates that are targeted by CobB for de-2-hydroxyisobutyrylation. We further demonstrated that CobB can regulate the catalytic activities of enolase (ENO) by removing K343hib and K326ac of ENO simultaneously, which account for changes of bacterial growth. In brief, our study dissects a Khib-mediated molecular mechanism that is catalyzed by CobB for the regulation of the activity of metabolic enzymes as well as the cell growth of bacteria.

ObjectiveThis study assessed the characteristics of pathogens identified in clinical isolates from patients with urinary tract infection (UTI) and their in vitro sensitivity to commonly used antibiotics in the clinical setting in China.Design and settingMulticenter study was conducted between January and December 2011 in 12 hospitals in China.ParticipantsUrine samples were collected from 356 symptomatic patients treated in the study hospitals for acute uncomplicated cystitis, recurrent UTI or complicated UTI.Primary and secondary outcome measuresMinimal inhibitory concentrations (MICs) were measured using broth microdilution according to the Clinical and Laboratory Standards Institute 2011 guidelines. Thirteen antimicrobial agents were tested: fosfomycin tromethamine, levofloxacin, moxifloxacin, cefdinir, cefixime, cefaclor, cefprozil, cefuroxime, amoxicillin/clavulanic acid, cefotaxime, azithromycin, nitrofurantoin and oxacillin. Escherichia coli isolates were screened and extended spectrum β-lactamases (ESBL) production was confirmed by a double-disk synergy test.Results198 urine samples were culture-positive and 175 isolates were included in the final analysis. E coli was detected in 50% of cultures, followed by Staphylococcus epidermidis (9%), Enterococcus faecalis (9%) and Klebsiella pneumoniae (5%). The detection rate of ESBL-producing E coli was 53%. Resistance to levofloxacin was the most common among all the isolates. Nitrofurantoin and fosfomycin tromethamine had the greatest activity against E coli; overall, 92% and 91% of isolates were susceptible to these antimicrobials. E faecalis had the highest susceptibility rates to fosfomycin tromethamine (100%).ConclusionsThe most frequently identified pathogens in our patients were ESBL-producing E coli and E faecalis. Fosfomycin tromethamine and nitrofurantoin showed a good antimicrobial activity against UTI pathogens. They may represent good options for the empiric treatment of patients with UTI.

Whether transcriptional regulators are functionally involved in mitosis is a fundamental question in cell biology. Here we report that the RNF20/40 complex, a major ubiquitin ligase catalysing histone H2B monoubiquitination, interacts with the motor protein Eg5 during mitosis and participates in spindle assembly. We show that the RNF20/40 complex monoubiquitinates and stabilizes Eg5. Loss of RNF20/40 results in spindle assembly defects, cell cycle arrest and apoptosis. Consistently, depletion of either RNF20/40 or Eg5 suppresses breast cancer in vivo. Significantly, RNF20/40 and Eg5 are concurrently upregulated in human breast carcinomas and high Eg5 expression is associated with poorer overall survival of patients with luminal A, or B, breast cancer. Our study uncovers an important spindle assembly role of the RNF20/40 complex, and implicates the RNF20/40-Eg5 axis in breast carcinogenesis, supporting the pursuit of these proteins as potential targets for breast cancer therapeutic interventions.