Isolation and characterization of complementary DNAs encoding human pregnancy-specific beta 1-glycoprotein

Pregnancy-specific beta 1-glycoprotein (PS beta G) isolated from human placenta consists of a set of at least three glycoproteins with apparent molecular masses of 72, 64, and 54 kDa, respectively. This heterogeneity is confirmed by the detection of three nonglycosylated polypeptides of 50, 48, and 36 kDa, which can be immunoprecipitated by antiserum to placental PS beta G obtained by in vitro translation of placental poly(A)+ RNA. To examine the structural relationships between these proteins, two cDNA clones of 1912 base pairs (PSG16) and 2131 base pairs (PSG93) encoding human PS beta Gs were isolated from a human placental lambda gt11 cDNA library. The sequenced portions of these two cDNAs are identical with the exception that clone PSG93 contains an additional 86 base pairs at the end of the common 3'-coding region. This insertion could result in the generation of a PS beta G species of 419 amino acid residues instead of the 417 amino acid residues predicted by the sequence of clone PSG16. The calculated molecular masses of the two polypeptides encoded by PSG16 and PSG93 are 46.9 and 47.2 kDa, close to the size of the major nonglycosylated PS beta G of 48 kDa. The identity of proteins coded for by these cDNA clones was confirmed by comparing the predicted amino acid sequences to sequences determined from endoproteinase Lys-C peptides obtained from human placental PS beta G. Two placental PS beta G mRNAs of 2200 bases (major) and 1700 bases (minor) have been detected by Northern hybridization analysis. Primer extension and S1 nuclease mapping experiments demonstrated that PS beta G mRNAs have heterogeneous 5' termini.     

Sequence-specific recognition of DNA: NMR studies of the imino protons of a synthetic RNA polymerase promoter

We have synthesized both strands of a DNA duplex containing the consensus Pribnow promoter sequence TATAATG , flanked by GC base pairs to stabilize the ends of the helix. The stability of this duplex has been studied by using 1H nuclear magnetic resonance. The imino protons have been assigned by using the sequential nuclear Overhauser effect approach. Exchange rates have been monitored by using selective inversion recovery measurements. The helix is relatively unstable in the center of the AT-rich region even when surrounded by GC base pairs, and there is considerable asymmetry in the melting of the helix.     

The high maltose-producing α-amylase of Penicillium expansum

Summary An -amylase capable of producing exceptionally high levels of maltose (74%) from starch has been identified from a strain of Penicillium expansum. The enzyme is produced extracellularly and was purified to homogeneity by starch adsorption and Sephadex gel filtration chromatography. P. expansum -amylase has a pH optimum of 4.5 and is stable in the pH range of 3.6–6.0. Other properties include a temperature optimum of 60° C, a molecular weight of 69 000 and an isoelectric point of 3.9. The most outstanding feature of the P. expansum enzyme is its ability to yield 14% more maltose and 17.1% less maltotriose than a currently used commercial enzyme. This may be partly explained by the greater affinity of this new enzyme for maltotriose (K _m=0.76 mM) relative to the commerical enzyme, Fungamyl (K _m=2.9 mM). The enzyme reported here is unique among fungal -amylases in being able to produce such high levels of maltose and its physicochemical properties suggest that it has potential for commercial development.     

Low-frequency vibrations of DNA molecules.

A model for calculating the low-frequency modes in DNA molecules is presented. The present model is associated with the 'breathing' of a DNA molecule as well as its complementary hydrogen bonds. The calculated results show excellent agreement with the observed low-frequency wavenumber (30 cm-1). Consequently, such an internal motion as reflected in the proposed model might be the origin of the observed low-frequency vibration in DNA molecules. This is helpful for investigating the relevant biological functions, which so far have been discussed by many scientists.     

Removal of Transmissible Spongiform Encephalopathy Prion from Large Volumes of Cell Culture Media Supplemented with Fetal Bovine Serum by Using Hollow Fiber Anion-Exchange Membrane Chromatography

Cases of variant Creutzfeldt-Jakob disease in people who had consumed contaminated meat products from cattle with bovine spongiform encephalopathy emphasize the need for measures aimed at preventing the transmission of the pathogenic prion protein (PrP^Sc) from materials derived from cattle. Highly stringent scrutiny is required for fetal bovine serum (FBS), a growth-medium supplement used in the production of parenteral vaccines and therapeutic recombinant proteins and in the ex vivo expansion of stem cells for transplantation. One such approach is the implementation of manufacturing steps dedicated to removing PrP^Sc from materials containing FBS. We evaluated the use of the QyuSpeed D (QSD) adsorbent hollow-fiber anion-exchange chromatographic column (Asahi Kasei Medical, Tokyo, Japan) for the removal of PrP^Sc from cell culture media supplemented with FBS. We first established that QSD filtration had no adverse effect on the chemical composition of various types of culture media supplemented with 10% FBS or the growth and viability characteristics of human embryonic kidney (HEK293) cells, baby hamster kidney (BHK-21) cells, African green monkey kidney (Vero) cells, and Chinese hamster ovary (CHO-k1) cells propagated in the various culture-medium filtrates. We used a 0.6-mL QSD column for removing PrP^Sc from up to 1000 mL of Dulbecco’s modified Eagle’s medium containing 10% FBS previously spiked with the 263K strain of hamster-adapted scrapie. The Western blot analysis, validated alongside an infectivity assay, revealed that the level of PrP^Sc in the initial 200mL flow-through was reduced by 2.5 to > 3 log₁₀, compared with that of the starting material. These results indicate that QSD filtration removes PrP^Sc from cell culture media containing 10% FBS, and demonstrate the ease with which QSD filtration can be implemented in at industrial-scale to improve the safety of vaccines, therapeutic recombinant proteins, and ex vivo expanded stem cells produced using growth media supplemented with FBS.     

Do individual allocyclic chromosomes in metaphase reflect their interphase domains?

Summary Individual S phase allocyclic chromosomes have been analyzed in Bloom syndrome lymphocytes, in cells with an r(9), and in hypotetraploid Ehrlich mouse ascites cells treated with 1-methyl-2-benzyl hydrazine. On the basis of the following observations, we conclude that such chromosomes more or less reflect their domains in interphase: (1) The S phase allocyclic chromosomes have the same structure as S phase prematurely condensed chromatin (PCC) in fused cells; in other words they form limited areas of chromatin dots; (2) the allocyclic chromosome is the only chromosome in a metaphase plate which synthesizes DNA simultanneously with interphase nuclei; (3) the size of the allocyclic chromosomes is related to the size of the corresponding metaphase chromosome; and (4) the S phase allocyclic chromosomes resemble closely the chromosome domains in interphase made visible with biotinylated human DNA. A variety of evidence shows that most allocyclic chromosomes are simply left behind in their cycle, which presumably is caused by a deletion or inactivation of a hypothetical coiling center situated on each chromosome arm.     

CD200R1 Supports HSV-1 Viral Replication and Licenses Pro-Inflammatory Signaling Functions of TLR2

The CD200R1:CD200 axis is traditionally considered to limit tissue inflammation by down-regulating pro-inflammatory signaling in myeloid cells bearing the receptor. We generated CD200R1^−/− mice and employed them to explore both the role of CD200R1 in regulating macrophage signaling via TLR2 as well as the host response to an in vivo, TLR2-dependent model, herpes simplex virus 1 (HSV-1) infection. CD200R1^−/− peritoneal macrophages demonstrated a 70–75% decrease in the generation of IL-6 and CCL5 (Rantes) in response to the TLR2 agonist Pam₂CSK₄ and to HSV-1. CD200R1^−/− macrophages could neither up-regulate the expression of TLR2, nor assemble a functional inflammasome in response to HSV-1. CD200R1^−/− mice were protected from HSV-1 infection and exhibited dysfunctional TLR2 signaling. Finally, both CD200R1^−/− mice and CD200R1^−/− fibroblasts and macrophages showed a markedly reduced ability to support HSV-1 replication. In summary, our data demonstrate an unanticipated and novel requirement for CD200R1 in “licensing” pro-inflammatory functions of TLR2 and in limiting viral replication that are supported by ex vivo and in vivo evidence.     

Ultraviolet light-induced responses of an mfd mutant of escherichia coli B/r having a slow rate of dimer excision

A mutant of Eschirichia coli B/r designated mfd has drastically reduced ability to exhibit “mutation frequency decline” (MFD) the irreversible loss of potential suppressor mutations which occurs when protein synthesis is briefly inhibited after irradiation with U.V. We have found that the initial rate of thymine dimer excision in the mfd mutant is only about one-third that of its mfd⁺ parent strain after a UV dose of 400 erg/mm². The yield of UV-induced Tyr⁺ revertants is 4–10 times higher in the mfd strain than in the mfd⁺ strain. This is comparable to the level of UV-mutability in the mfd⁺ strain in the presence of caffeine, an inhibitor of dimer excision. UV-mutability, prophage induction and Weigle reactivation of irradiated λ phage occur to a greater extent at low UV doses (10–50 erg/mm²) in the mfd strain compared to the mfd⁺ strain. We propose that the slow excision repair in the mfd mutant results in a shift in the induction threshold for these UV-inducible functions toward lower UV doses.     

Smoking during Pregnancy: Findings from the 2009–2010 Canadian Community Health Survey

Objectives

Smoking during pregnancy may cause many health problems for pregnant women and their newborns. However, there is a paucity of research that has examined the predictors of smoking during pregnancy in Canada. This study used data from the 2009–2010 Canadian Community Health Survey (CCHS) to estimate the prevalence of smoking during pregnancy and examine the demographic, socioeconomic, health-related and behavioral determinants of this behavior.

Methods and Findings

The data were obtained from the 2009–2010 CCHS master data file. Weighted estimates of the prevalence were calculated. Multivariable logistic regression was used to determine demographic, socioeconomic, health related and behavioral characteristics associated with smoking behavior during pregnancy. Women living in the Northern Territories had a high rate of smoking during pregnancy (59.3%). The prevalence of smoking during pregnancy was also high among women under 25 years old, of low socioeconomic status, who reported not having a regular medical doctor, being fair to poor in self-perceived health, having at least one chronic disease, having at least one mental illness, being heavy smokers, and being regular alcohol drinkers. Results from multivariable logistic regression revealed that the odds of smoking during pregnancy were decreased with increasing age (odds ratio [OR], 0.95; 95% confidence interval [CI], 0.91–0.99), having a regular family doctor [OR, 0.24; 95% CI, 0.11–0.52], having highest level of family income [OR, 0.09; 95% CI, 0.03–0.29]. Mothers who reported poor or fair self-perceived health [OR, 2.13; 95% CI, 0.96–4.71] and those who had at least one mental illness [OR, 1.81; 95% CI, 1.00–3.28] had greater odds of smoking during pregnancy.

Conclusions

There are a number of demographic, socio-economic, health-related and behavioral characteristics that should be considered in developing and implementing effective population health promotional strategies to prevent smoking during pregnancy, promoting health and well-being of pregnant women and their newborns.