Cohesin mediates chromatin interactions that regulate mammalian β-globin expression

The β-globin locus undergoes dynamic chromatin interaction changes in differentiating erythroid cells that are thought to be important for proper globin gene expression. However, the underlying mechanisms are unclear. The CCCTC-binding factor, CTCF, binds to the insulator elements at the 5' and 3' boundaries of the locus, but these sites were shown to be dispensable for globin gene activation. We found that, upon induction of differentiation, cohesin and the cohesin loading factor Nipped-B-like (Nipbl) bind to the locus control region (LCR) at the CTCF insulator and distal enhancer regions as well as at the specific target globin gene that undergoes activation upon differentiation. Nipbl-dependent cohesin binding is critical for long-range chromatin interactions, both between the CTCF insulator elements and between the LCR distal enhancer and the target gene. We show that the latter interaction is important for globin gene expression in vivo and in vitro. Furthermore, the results indicate that such cohesin-mediated chromatin interactions associated with gene regulation are sensitive to the partial reduction of Nipbl caused by heterozygous mutation. This provides the first direct evidence that Nipbl haploinsufficiency affects cohesin-mediated chromatin interactions and gene expression. Our results reveal that dynamic Nipbl/cohesin binding is critical for developmental chromatin organization and the gene activation function of the LCR in mammalian cells.     

Coevolutionary analysis enabled rational deregulation of allosteric enzyme inhibition in Corynebacterium glutamicum for lysine production

Product feedback inhibition of allosteric enzymes is an essential issue for the development of highly efficient microbial strains for bioproduction. Here we used aspartokinase from Corynebacterium glutamicum (CgAK), a key enzyme controlling the biosynthesis of industrially important aspartate family amino acids, as a model to demonstrate a fast and efficient approach to the deregulation of allostery. In the last 50 years many researchers and companies have made considerable efforts to deregulate this enzyme from allosteric inhibition by lysine and threonine. However, only a limited number of positive mutants have been identified so far, almost exclusively by random mutation and selection. In this study, we used statistical coupling analysis of protein sequences, a method based on coevolutionary analysis, to systematically clarify the interaction network within the regulatory domain of CgAK that is essential for allosteric inhibition. A cluster of interconnected residues linking different inhibitors' binding sites as well as other regions of the protein have been identified, including most of the previously reported positions of successful mutations. Beyond these mutation positions, we have created another 14 mutants that can partially or completely desensitize CgAK from allosteric inhibition, as shown by enzyme activity assays. The introduction of only one of the inhibition-insensitive CgAK mutations (here Q298G) into a wild-type C. glutamicum strain by homologous recombination resulted in an accumulation of 58 g/liter L-lysine within 30 h of fed-batch fermentation in a bioreactor.     

Excisionase in Pf filamentous prophage controls lysis‐lysogeny decision‐making in Pseudomonas aeruginosa

Pf filamentous prophages are prevalent among clinical and environmental Pseudomonas aeruginosa isolates. Pf4 and Pf5 prophages are integrated into the host genomes of PAO1 and PA14, respectively, and play an important role in biofilm development. However, the genetic factors that directly control the lysis‐lysogeny switch in Pf prophages remain unclear. Here, we identified and characterized the excisionase genes in Pf4 and Pf5 (named xisF4 and xisF5, respectively). XisF4 and XisF5 represent two major subfamilies of functional excisionases and are commonly found in Pf prophages. While both of them can significantly promote prophage excision, only XisF5 is essential for Pf5 excision. XisF4 activates Pf4 phage replication by upregulating the phage initiator gene (PA0727). In addition, xisF4 and the neighboring phage repressor c gene pf4r are transcribed divergently and their 5′‐untranslated regions overlap. XisF4 and Pf4r not only auto‐activate their own expression but also repress each other. Furthermore, two H‐NS family proteins, MvaT and MvaU, coordinately repress Pf4 production by directly repressing xisF4. Collectively, we reveal that Pf prophage excisionases cooperate in controlling lysogeny and phage production.     

The Great Chinese Famine Leads to Shorter and Overweight Females in Chongqing Chinese Population After 50 Years

This study investigated a possible association between early nutritional status during the famine, and the risk of overweight and obesity in adulthood in Chongqing Chinese population. The body weight, height, and BMI data were obtained from records of population (17,023) that had annual physical evaluations in the Public Health Center (in our hospital). Subjects born during 1956–1964 were divided into three groups: toddler group, all subjects who were born 1–3 years before the famine (1956–1958); gestational group, who were born during the famine period (1959–1961), and control group, who were born after the famine (1962–1964). The body weight and BMI were significantly higher, but the body height was significantly lower in the toddler and gestational groups (P < 0.05) in the female population as compared to the control group. The odds ratio of being overweight in females is more pronounced in the toddler group (1.48 times, 95% confidence interval (CI): 1.288–1.689) than in the gestational group (1.26 times, 95% CI: 1.089–1.457). The odds ratio of being obese in females is significantly higher in the toddler group (1.46 times, 95% CI: 1.288–1.689) than the control group. For males, the famine had no impact at all on the adulthood body weight in males. The Great Chinese Famine that affected the Chongqing population during 1959–1961 leads to shorter and overweight females, and the former is a risk factor for increased BMI in Chongqing. Second, the famine seems to be producing shorter but slimmer males in Chongqing. Furthermore, toddler's and maternal's malnutrition during the famine had important late consequences on the health status.     

Inhibition kinetics of green crab (Scylla serrata) alkaline phosphatase by zinc ions: a new type of complexing inhibition

The Tsou method was used to study the kinetic course of inactivation of green crab alkaline phosphatase by zinc ions. The results show that the enzyme was inactivated by a complexing scheme which has not been previously identified. The enzyme first reversibly and quickly binds Zn(2+) and then undergoes a slow reversible course to inactivation and slow conformational change. The inactivation reaction is a single molecule reaction and the apparent inactivation rate constant is for a saturated reaction being independent of Zn(2+) concentration if the concentration is sufficiently high. The microscopic rate constants of inactivation and the association constant were determined from the measurements.     

Highly immunogenic and totally synthetic lipopeptides as self-adjuvanting immunocontraceptive vaccines

In this study, we describe the synthesis of various lipopeptides based on the sequence of luteinizing hormone-releasing hormone (LHRH) and report on their abilities to induce Abs against this "self" hormone when inoculated into mice in the absence of additional adjuvant. The peptides consisted of a colinear CD4(+) T helper cell epitope from the L chain of influenza virus hemagglutinin and LHRH, which has B cell epitopes but no T cell epitopes present in its sequence. Lipids were attached either at the N terminus or between the T cell epitope and LHRH, in the approximate center of the peptide. The lipopeptide constructs displayed different solubilities and immunological properties that depended not only on the lipid content but also on the position of attachment of the lipids. Some of these constructs were highly immunogenic, inducing high titers of Ab, which were capable of efficiently sterilizing female mice when administered in saline by s.c. or intranasal routes. The most effective vaccines were highly soluble, contained the dipalmitoyl-S-glyceryl cysteine moiety, and had this lipid attached at the center of the molecule. The relative ability of the lipopeptides to induce an Ab response in the absence of external adjuvant was reflected by their ability to up-regulate the surface expression of MHC class II molecules on immature dendritic cells. These results demonstrate that the composition and position within peptide vaccines of self-adjuvanting lipid groups can influence the ability to induce the maturation of dendritic cells and, in turn, the magnitude of the resulting Ab response.     

Sinorhizobium fredii HH103 has a truncated nolO gene due to a -1 frameshift mutation that is conserved among other geographically distant S. fredii strains

Strain SVQ121 is a mutant derivative of Sinorhizobium fredii HH103 carrying a transposon Tn5-lacZ insertion into the nolO-coding region. Sequence analysis of the wild-type gene revealed that it is homologous to that of Rhizobium sp. NGR234, which is involved in the 3 (or 4)-O-carbamoylation of the nonreducing terminus of Nod factors. Downstream of nolO, as in Rhizobium sp. NGR234, the noeI gene responsible for methylation of the fucose moiety of Nod factors was found. SVQ121 Nod factors showed lower levels of methylation into the fucosyl residue than those of HH103-suggesting a polar effect of the transposon insertion into nolO over the noel gene. A noeI HH103 mutant was constructed. This mutant, SVQ503, produced Nod factors devoid of methyl groups, confirming that the S. fredii noeI gene is functional. Neither the nolO nor the noeI mutation affected the ability of HH103 to nodulate several host plants, but both mutations reduced competitiveness to nodulate soybean. The Nod factors produced by strain HH103, like those of other S. fredii isolates, lack carbamoyl residues. By using specific polymerase chain reaction primers, we sequenced the nolO gene of S. fredii strains USDA192, USDA193, USDA257, and 042B(s). All the analyzed strains showed the same -1 frameshift mutation that is present in the HH103 nolO-coding region. From these results, it is concluded that, regardless of their geographical origin, S. fredii strains carry the nolO-coding region but that it is truncated by the same base-pair deletion.     

PBMC activation via the ERK and STAT signaling pathways enhances the anti-tumor activity of <Emphasis Type="Italic">Staphylococcal</Emphasis> enterotoxin A

In a recent study, we highlighted the importance of cationic charge and arginine residues for the neuroprotective properties of poly-arginine and arginine-rich peptides. In this study, using cortical neuronal cultures and an in vitro glutamic acid excitotoxicity model, we examined the neuroprotective efficacy of different modifications to the poly-arginine-9 peptide (R9). We compared an unmodified R9 peptide with R9 peptides containing the following modifications: (i) C-terminal amidation (R9-NH2); (ii) N-terminal acetylation (Ac-R9); (iii) C-terminal amidation with N-terminal acetylation (Ac-R9-NH2); and (iv) C-terminal amidation with d-amino acids (R9D-NH2). The three C-terminal amidated peptides (R9-NH2, Ac-R9-NH2, and R9D-NH2) displayed neuroprotective effects greater than the unmodified R9 peptide, while the N-terminal acetylated peptide (Ac-R9) had reduced efficacy. Using the R9-NH2 peptide, neuroprotection could be induced with a 10 min peptide pre-treatment, 1–6 h before glutamic acid insult, or when added to neuronal cultures up to 45 min post-insult. In addition, all peptides were capable of reducing glutamic acid-mediated neuronal intracellular calcium influx, in a manner that reflected their neuroprotective efficacy. This study further highlights the neuroprotective properties of poly-arginine peptides and provides insight into peptide modifications that affect efficacy.