Regional assignment of the human gene for platelet-type phosphofructokinase (PFKP) to chromosome 10p: novel use of polyspecific rodent antisera to localize human enzyme genes

Human phosphofructokinase (PFK; EC 2.7.1.11) is under the control of three structural loci which encode muscle-type (M), liver-type (L), and platelet or fibroblast-type (P) subunits; human diploid fibroblasts and leukocytes express all three loci. In order to assign the human PFKP locus to a specific human chromosome, in this study, we have examined ten human X rodent somatic cell hybrids for the expression of human P subunits using a mouse anti-human P subunit-specific antiserum in an active-enzyme-immunoprecipitation technique. In nine of ten hybrids studied, the expression of the PFKP locus segregated concordantly with chromosome 10 and none other, indicating that PFKP is located on chromosome 10; the discordancy rates for all the other chromosomes were 0.2 or greater. In the one discordant hybrid, only the long arm of chromosome 10 was retained and PFKP was not expressed. Human fibroblasts from a patient with duplication of the short arm of chromosome 10 consistently exhibited PFK activity values 180% of normal. These data indicate that human PFKP is located on the short arm of chromosome 10, and that a gene dosage effect is demonstrable in fibroblasts with a duplication of 10p. The use of rodent antihuman antibody combined with immunoprecipitation aided by staphylococci-bearing protein A may find general application in mapping human enzyme genes, when human and rodent gene-products are not distinguishable by other means.     

Studies on immobilized fungal spores for microbial transformation of steroids: 11 alpha-Hydroxylation of progesterone with immobilized spores of Aspergillus ochraceus G8 on polyacrylamide gel and other matrices

Hydroxylation in the 11 alpha-position in the progesterone molecule employing immobilized spores of Aspergillus ochraceus strain No. G8 (CDRI catalogue No.) was achieved. For immobilization the activity of the spores was evaluated on a variety of matrices such as alginate beads, epoxy resin beads, polyacrylamide gel, and collagen. Spores entrapped in polyacrylamide gel were found to be the most active. Studies of various parameters, e.g. monomer content, cell loading capacity, optimum pH, temperature, and substrate concentration, were carried out on polyacrylamide gel. In polyacrylamide, the entrapped spores normal decay pattern, as indicated by loss of activity, was observed after four uses. At the end of 15 cycles, the residual activity was found to be 18% of the original. It was possible to regenerate the activity by incubating the preparation in a nutrient medium. The regenerated spores showed increasing rate of loss of activity upon recycling.     

Site-specific nicking at the avian retrovirus LTR circle junction by the viral pp32 DNA endonuclease.

The avian retrovirus pp32 DNA endonuclease prefers to nick supercoiled DNA containing long terminal repeat (LTR) circle junction sequences at one or the other of two sites, each which mapped two nucleotides back from the circle junction. The sequence at the sites of nicking was (sequence: see text) where increases indicates the positions of the two alternative nicked sites. This site-specific nicking was observed when the circle junction LTR DNA was present in supercoiled form, the divalent metal ion was Mg2+ and the molar ratio of protein to DNA was low. The majority of other LTR DNA sites nicked by pp32 in the presence of Mg2+ were adjacent to or within the dinucleotide CA.     

Beta-defensin-2 expression is regulated by TLR signaling in intestinal epithelial cells

The intestinal epithelium serves as a barrier to the intestinal flora. In response to pathogens, intestinal epithelial cells (IEC) secrete proinflammatory cytokines. To aid in defense against bacteria, IEC also secrete antimicrobial peptides, termed defensins. The aim of our studies was to understand the role of TLR signaling in regulation of beta-defensin expression by IEC. The effect of LPS and peptidoglycan on beta-defensin-2 expression was examined in IEC lines constitutively or transgenically expressing TLRs. Regulation of beta-defensin-2 was assessed using promoter-reporter constructs of the human beta-defensin-2 gene. LPS and peptidoglycan stimulated beta-defensin-2 promoter activation in a TLR4- and TLR2-dependent manner, respectively. A mutation in the NF-kappaB or AP-1 site within the beta-defensin-2 promoter abrogated this response. In addition, inhibition of Jun kinase prevents up-regulation of beta-defensin-2 protein expression in response to LPS. IEC respond to pathogen-associated molecular patterns with expression of the antimicrobial peptide beta-defensin-2. This mechanism may protect the intestinal epithelium from pathogen invasion and from potential invaders among the commensal flora.     

Regional assignment of human liver-type 6-phosphofructokinase to chromosome 21q22.3 by using somatic cell hybrids and a monoclonal anti-L antibody

Summary The three structural loci encoding human phosphofructokinase, a key regulatory enzyme of glycolysis, are located on separate chromosomes. The gene coding for the liver-type subunit PFKL has previously been assigned to chromosome 21. We have used a subunit- and human-specific monoclonal antibody to liver PFK to detect the expression of human PFKL in hamster x human hybrid cell lines. A cell line carrying an 8;21 translocation which contains all of chromosome 21 except the band 21q22.3 was negative for the expression of PFKL whereas cell lines carrying the reciprocal 8;21 translocation were positive. In addition, a cell line with a ring chromosome 21 containing a breakpoint which excluded the distal part of the q22.3 band was negative for expression of PFKL. These results indicate that human PFKL is located on chromosome 21q22.3.     

Metabolic studies with neurospora: Action of 5-bromo- and 5-nitrouracils

Summary There is no growth inhibition of Neurospora crassa even in presence of high levels of 5-bromouracil. Although there is no significant change in the DNA and RNA contents, the analogue is utilized by the organism, a part of it being incorporated in DNA. There is an increase in total folate and citrovorum factor activities with 5-bromouracil.5-Nitrouracil causes a significant growth inhibition at higher concentration which is accompanied by a decrease in pteroylglutamic acid and citrovorum factor levels in the organism.Pteroylglutamic acid and thymidine cause a partial reversal of growth inhibition by 5-nitrouracil and complete reversal when the two metabolites are added simultaneously.     

Experimental Sociality and Gestational Surrogacy in the Indian ART Clinic

Abstract

This article marks experimental modes of sociality in a transnational Indian assisted reproductive technology (ART) clinic as a contact zone between elite doctors, gestational surrogates, and transnational commissioning parents. It examines efforts within one ART clinic to separate social relationships from reproductive bodies in its surrogacy arrangements as well as novel social formations occurring both because of and despite these efforts. Draft regulative legislation in India marks a shift in the distribution of risk among actors in the clinic that parallels a shift in medical practice away from a technique of caring for the body to producing bodies as instruments of contracted service. The clinic provides an opportunity to observe forms of sociality that emerge as experiments with modernities, with different relationships to the body and the social meaning of medicalized biological reproduction, and with understanding the role of the market and altruism in the practice of gestational surrogacy.