Periplasmic Acid Stress Increases Cell Division Asymmetry (Polar Aging) of Escherichia coli

Under certain kinds of cytoplasmic stress, Escherichia coli selectively reproduce by distributing the newer cytoplasmic components to new-pole cells while sequestering older, damaged components in cells inheriting the old pole. This phenomenon is termed polar aging or cell division asymmetry. It is unknown whether cell division asymmetry can arise from a periplasmic stress, such as the stress of extracellular acid, which is mediated by the periplasm. We tested the effect of periplasmic acid stress on growth and division of adherent single cells. We tracked individual cell lineages over five or more generations, using fluorescence microscopy with ratiometric pHluorin to measure cytoplasmic pH. Adherent colonies were perfused continually with LBK medium buffered at pH 6.00 or at pH 7.50; the external pH determines periplasmic pH. In each experiment, cell lineages were mapped to correlate division time, pole age and cell generation number. In colonies perfused at pH 6.0, the cells inheriting the oldest pole divided significantly more slowly than the cells inheriting the newest pole. In colonies perfused at pH 7.50 (near or above cytoplasmic pH), no significant cell division asymmetry was observed. Under both conditions (periplasmic pH 6.0 or pH 7.5) the cells maintained cytoplasmic pH values at 7.2–7.3. No evidence of cytoplasmic protein aggregation was seen. Thus, periplasmic acid stress leads to cell division asymmetry with minimal cytoplasmic stress.     

Epstein-Barr Virus Polypeptides: Identification of Early Proteins and Their Synthesis and Glycosylation

We have identified at least six early polypeptides induced by Epstein-Barr virus in cells or under conditions which are nonpermissive for Epstein-Barr virus DNA replication ranging in molecular weight from 140,000 to 26,000.     

Immunochemical detection and characterisation of osteocalcin from moa bone

Osteocalcin (the 6,000 dalton Mr gamma-carboxyglutamate-containing protein of bone) has been detected in acid extracts of bones of the extinct class of New Zealand ratite birds, the moas, using a radioimmunoassay for sheep osteocalcin. The immunoreactive osteocalcin of the extracts of two of these bones (the fibulae from two specimens of Pachyornis elephantopus found in South Island swamps) has been fractionated by gel filtration chromatography and reversed-phase high performance liquid chromatography, and behaves in a manner characteristic of osteocalcin from modern bones. Carbon-14 dating of bones and gizzard contents found in association with these specimens indicates approximate ages of 3,600 and 7,400 years respectively.     

Monoclonal antibody specific for α2→8-linked oligo deaminated neuraminic acid (KDN) sequences in glycoproteins. Preparation and characterization of a monoclonal antibody and its application in immunohistochemistry

Two particular types of sialoglycoproteins have been detected in fish: polysialoglycoproteins containing 28-linked polysialic acid (8Neu5Gc2) _n present in unfertilized Salmonidae fish eggs, and glycoproteins bearing oligo/polymers of deaminated neuraminic acids (KDN) found in the vitelline envelope of the eggs and ovarian fluid. We report the preparation and characterization of a monoclonal antibody specifically recognizing oligo/polymers of KDN sequences in glycoproteins and its application in immunohistochemistry. Fusion of spleen cells from a BALB/c mouse immunized with a KDN-rich glycoprotein (KDN-gp) containing (8KDN2) _n 6(KDN23Gal13GlNAc13) GalNAc1 residues, with mouse myeloma cells yielded a hybrid cell line producing a monoclonal antibody that bound to KDN-gp, but not to KDN-gp depleted of KDN residues. The specificity of the monoclonal antibody, designated mAb.kdn8kdn, was determined by an enzyme-linked immunosorbent assay using KDN-gp samples that varied in KDN content. These antigens were prepared by the selective removal of KDN residues from the native KDN-gp. The mAb.kdn8kdn reacted most strongly with the intact KDN-gp and less strongly with KDN-gp samples containing decreased numbers of KDN residues. The mAb.kdn8kdn was shown specifically to recognize the 28-linked oligo/polyKDN sequences, (8KDN2) _n , and to be able to distinguish specifically (8KDN2) _n chains from (8Neu5Ac2) _n and (8Neu5Gc2) _n chains. The antibody was used successfully for the immunohistochemical detection of reactive KDN epitopes in sections of paraffin embedded rat pancreas. Several controls verified the specificity of the immunohistochemical staining, thus providing the first demonstration of (8KDN2) _n sequences in a mammalian tissue. The mAb.kdn8kdn can now be used to search further for glycoconjugates containing (8KDN2) _n chains and will facilitate studies on their biosynthesis, intracellular localization and function.     

Humpback whale abundance in the North Pacific estimated by photographic capture‐recapture with bias correction from simulation studies

We estimated the abundance of humpback whales in the North Pacific by capture‐recapture methods using over 18,000 fluke identification photographs collected in 2004–2006. Our best estimate of abundance was 21,808 (CV = 0.04). We estimated the biases in this value using a simulation model. Births and deaths, which violate the assumption of a closed population, resulted in a bias of +5.2%, exclusion of calves in samples resulted in a bias of ?10.5%, failure to achieve random geographic sampling resulted in a bias of ?0.4%, and missed matches resulted in a bias of +9.3%. Known sex‐biased sampling favoring males in breeding areas did not add significant bias if both sexes are proportionately sampled in the feeding areas. Our best estimate of abundance was 21,063 after accounting for a net bias of +3.5%. This estimate is likely to be lower than the true abundance due to two additional sources of bias: individual heterogeneity in the probability of being sampled (unquantified) and the likely existence of an unknown and unsampled breeding area (?8.7%). Results confirm that the overall humpback whale population in the North Pacific has continued to increase and is now greater than some prior estimates of prewhaling abundance.     

Genomic regions associated with microdeletion/microduplication syndromes exhibit extreme diversity of structural variation

Segmental duplications (SDs) are a class of long, repetitive DNA elements whose paralogs share a high level of sequence similarity with each other. SDs mediate chromosomal rearrangements that lead to structural variation in the general population as well as genomic disorders associated with multiple congenital anomalies, including the 7q11.23 (Williams–Beuren Syndrome, WBS), 15q13.3, and 16p12.2 microdeletion syndromes. Population-level characterization of SDs has generally been lacking because most techniques used for analyzing these complex regions are both labor and cost intensive. In this study, we have used a high-throughput technique to genotype complex structural variation with a single molecule, long-range optical mapping approach. We characterized SDs and identified novel structural variants (SVs) at 7q11.23, 15q13.3, and 16p12.2 using optical mapping data from 154 phenotypically normal individuals from 26 populations comprising five super-populations. We detected several novel SVs for each locus, some of which had significantly different prevalence between populations. Additionally, we localized the microdeletion breakpoints to specific paralogous duplicons located within complex SDs in two patients with WBS, one patient with 15q13.3, and one patient with 16p12.2 microdeletion syndromes. The population-level data presented here highlights the extreme diversity of large and complex SVs within SD-containing regions. The approach we outline will greatly facilitate the investigation of the role of inter-SD structural variation as a driver of chromosomal rearrangements and genomic disorders.     

ELECTRON-OPAQUE BODIES AND FAT DROPLETS IN MOUSE LIVER AFTER FASTING OR GLUCOSE INJECTION

Fasting produces an increased mobilization of lipid from adipose tissue to the liver and a decreased hepatic lipogenesis, but the administration of glucose stimulates lipid synthesis by the liver. After fasting of C3H mice numerous electron-opaque bodies and large lipid droplets were present in the liver. In the liver of untreated controls only a few small electron-opaque bodies and an occasional fat droplet were observed. After glucose injection the number of electron-opaque bodies in the liver was no greater than that observed in livers of saline-injected controls. In the livers of all groups these bodies were located intracellularly within cytoplasmic vesicles; those in extracellular locations were not membrane bounded and were located at indented and thickened hepatocyte plasma membranes or within the space of Disse. In fasted liver the dense bodies were often associated with large fat droplets.