Mapping of the structural gene for S-adenosyl homocysteine hydrolase to mouse Chromosome 2, and related sequences to Chromosomes 8 and X

Comparative mapping studies in human and mouse have shown that, to date, human Chromosome (Chr) 20 is completely syntenic with distal mouse Chr 2. The structural locus for S-adenosyl-l-homocysteine hydrolase (EC 3.3.1.1) in human, AHCY, maps to 20 qterq13.1, and we report here that the homologous locus in the mouse, Ahcy, maps to distal mouse Chr 2 with gene order Pcna-Ahcy-Ada. Analysis of 123 progeny of an interspecific backcross between a laboratory stock, AN, and Mus spretus using a rat cDNA probe revealed the presence of at least two other Ahcy-related sequences segregating independently in the mouse genome. One, Ahcy-rs1, was mapped to Chr 8 in the BXH recombinant inbred strains, and the other, Ahcy-rs2, shows a pattern of inheritance consistent with X-linkage.     

Epithelio--mesenchymal interactions are critical for Quox 7 expression and membrane bone differentiation in the neural crest derived mandibular mesenchyme.

In higher vertebrates, branchial arch mesenchyme (ectomesenchyme) is derived from the cephalic neural crest. The ectomesenchyme of the mandibular arch yields the Meckel's cartilage and several membrane bones. We previously reported the isolation of a quail homeobox gene, Quox 7. In common with its mouse counterpart Hox 7, Quox 7 is highly expressed in the medioventral part of the mandibular arch and later in the precursor cells of the membrane bones. Since bone differentiation from ectomesenchyme is strictly dependent upon a signal provided by the mandibular epithelium, we decided to see whether the regulation of Quox 7 gene activity might be correlated with epithelio--mesenchymal interactions. Quox 7 expression was studied in E3 mandibular ectomesenchyme cultured in vitro or grafted on the chick chorioallantoic membrane either alone or recombined with the homotopic and heterotopic epithelia. We found that Quox 7 mRNA was undetectable after 48 h in cultures of mesenchyme alone while it remained abundant in non-cartilaginous tissue of the mandibular arch ectomesenchyme recombined with its own epithelium. The signal provided by the mandibular epithelium for Quox 7 expression can also arise from various heterotopic epithelia, e.g. of dorsal or ventral body wall and of limb bud. Thus the effect of the epithelium on Quox 7 expression in mesenchymal cells strictly parallels that on bone formation. These results strongly suggest that the epithelio-mesenchymal interactions have an essential role on the regulation of Quox 7 gene, the product of which seems to be, in turn, necessary for the execution of the skeletal developmental program in the facial area.     

Solution conformation of an oligonucleotide containing a G.G mismatch determined by nuclear magnetic resonance and molecular mechanics.

We have determined by two-dimensional nuclear magnetic resonance studies and molecular mechanics calculations the three dimensional solution structure of the non-selfcomplementary oligonucleotide, d(GAGGAGGCACG). d(CGTGCGTCCTC) in which the central base pair is G.G. This is the first structural determination of a G.G mismatch in a oligonucleotide. Two dimensional nuclear magnetic resonance spectra show that the bases of the mismatched pair are stacked into the helix and that the helix adopts a classical B-DNA form. Spectra of the exchangeable protons show that the two guanosines are base paired via their imino protons. For the non-exchangeable protons and for some of the exchangeable protons nuclear Overhauser enhancement build up curves at short mixing times have been measured. These give 84 proton-proton distances which are sensitive to the helix conformation. One of the guanosines adopts a normal anti conformation while the other is syn or close to syn. All non-terminal sugars are C2' endo. These data sets were incorporated into the refinement of the oligonucleotide structure by molecular mechanics calculations. The G.G mismatch shows a symmetrical base pairing structure. Although the mismatch is very bulky many of its features are close to that of normal B-DNA. The mismatch induces a small lateral shift in the helix axis and the sum of the helical twist above and below the mismatch is close to that of B-DNA.     

The solution structure of a DNA hairpin containing a loop of three thymidines determined by nuclear magnetic resonance and molecular mechanics.

We have determined by two-dimensional nuclear magnetic resonance studies and molecular mechanics calculations the three-dimensional solution structure of a 21 residue oligonucleotide capable of forming a hairpin structure with a loop of three thymidine residues. This structure is in equilibrium with a duplex form. At 33 degrees C, low ionic strength and in the presence of MgCl2 the hairpin form dominates in solution. Six Watson-Crick base pairs are formed topped by the loop structure. The residues 1-3 and 18-21 are not complementary and form dangling ends. Distance constraints have been derived from nuclear Overhauser enhancement measurements. These, together with molecular mechanics calculations, have been used to determine the structure. We do not observe stacking of thymidine residues either over the 3' or the 5' end of the stem.     

Use of a simple method for the Epstein-Barr virus transformation of lymphocytes from members of large families of Réunion Island

A simple method for the preparation of lymphoblastoid cell lines from small amounts (100 microliter) of frozen whole blood is described. A success score greater than 90% was obtained for EBV transformations using blood samples which had been collected several months before the infection. Due to the simplicity of the technique, up to 80 samples could be processed per day. This technique was used to prepared 242 permanent cell lines from 13 large families from Réunion Island showing blood group H deficiency. These cell lines are now available for genetic studies.     

CFC syndrome: a syndrome distinct from Noonan syndrome

We report two children with a common pattern of birth defects. Both have very sparse, curly hair, nystagmus and mental retardation. The first one has Noonan syndrome habitus associated with keratosis plantaris and nystagmus; the second one has a slightly Noonan-like face, macrocephaly, keratosis pilaris, and hypertrophic cardiomyopathy. They represent the extreme of a spectrum of congenital defects recently reported independently as CFC syndrome by Reynolds and as "Noonan-like short stature syndrome with sparse hair" by Baraitser and Patton. The clinical features are reviewed and the autonomy of the syndrome with regards to Noonan syndrome, is disputed, since every sign seems to occur independently in Noonan syndrome. The father of the second case probably has a minor syndrome expression, pointing to probable autosomal dominant inheritance.     

Evidence for heterogeneity of low-density lipoprotein metabolism in the cynomolgus monkey

Preliminary studies were performed to establish whether there was kinetic heterogeneity in the metabolism of subclasses of low-density lipoproteins (LDL) in the cynomolgus monkey. Previous studies of the effects of inhibition of hepatic triglyceride lipase in this species had shown an increase in the mass of lighter LDL (Sf greater than 9) and a decrease in the mass of denser LDL. LDL (1.019 less than d less than 1.063) were subdivided into two subfractions LDL1 (1.019 less than d less than 1.035) and LDL2 (1.035 less than d less than 1.063) by ultracentrifugation. The lipoproteins in these two fractions could be shown to have different flotation by analytic and isopycnic ultracentrifugation. When tracer amounts of homologous 125I-labeled very-low-density lipoproteins (VLDL) were injected into chow-fed cynomolgus monkeys, apoB radioactivity appeared in LDL1 prior to its appearance in LDL2. [125I]LDL1 injected into the monkey was removed from the LDL1 density subclass with a half-life of 5.5-10.3 h. Much of the radioactivity injected as LDL1 was converted to denser LDL (LDL2). Labeled LDL2 injected into the monkey was not converted to LDL1. Thus, at least two kinetically distinct subpopulations of LDL circulate in the plasma of this species. The lighter LDL is to a large extent a metabolic precursor of the more dense LDL (LDL2).     

Antigenic, functional, and molecular genetic studies of human natural killer cells and cytotoxic T lymphocytes not restricted by the major histocompatibility complex

Cytotoxicity not restricted by the major histocompatibility complex (MHC) is mediated by two distinct types of lymphocyte: natural killer (NK) cells and non-MHC-restricted cytotoxic T lymphocytes (CTL). These two types of cytotoxic lymphocytes can be distinguished by antigenic phenotype, function, and molecular genetic studies. In human peripheral blood, NK cells are identified by expression of the Leu-19 and/or CD16 cell surface antigens, and lack of CD3/T cell antigen receptor (Ti) complex expression (i.e., CD3-,Leu-19+). Peripheral blood non-MHC-restricted CTL express both CD3 and Leu-19 (i.e., CD3+, Leu-19+, referred to as Leu-19+ T cells). Both Leu-19+ T cells and NK cells lyse "NK-sensitive" hematopoietic tumor cell targets, such as K562, without deliberate immunization of the host. However, most "NK activity" in peripheral blood is mediated by NK cells, because they are usually more abundant and more efficient cytotoxic effectors than Leu-19+ T cells. The cytolytic activity of both NK cells and Leu-19+ T cells against hematopoietic targets was enhanced by recombinant interleukin 2 (rIL 2). NK cells, but not peripheral blood Leu-19+ T cells, were also capable of lysing solid tumor cell targets after short-term culture in rIL 2. Southern blot analysis of NK cells revealed that both the T cell antigen receptor beta-chain genes and the T cell-associated gamma genes were not rearranged, but were in germ-line configuration. These findings indicate that NK cells are distinct in lineage from T lymphocytes and do not use the T cell antigen receptor genes for target recognition.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modifications of sphingomyelin and phosphatidylcholine metabolism by tricyclic antidepressants and phenothiazines

Phenothiazines and tricyclic antidepressants, when added to culture medium, gave rise in several types of cells (C6 rat glioma cells and human fibroblasts), to a decrease in lysosomal sphingomyelinase activity. The effect of chlorpromazine and desipramine was dose dependent, and was observed after 3 hours of incubation with the drugs at concentrations ranging between 1 and 10 microM. In C6 glioma cell cultures, the decrease in sphingomyelinase activity was related to the clinical effectiveness of phenothiazines, tricyclic antidepressants and derivatives. Incorporation of (choline-14C) sphingomyelin showed that the metabolic pathway implying the synthesis of phosphatidylcholine from the hydrolysis of sphingomyelin and/or transfer of phosphorylcholine to phosphatidylcholine was also partially reduced.