High-resolution mapping of genotype-phenotype relationships in cri du chat syndrome using array comparative genomic hybridization

We have used array comparative genomic hybridization to map DNA copy-number changes in 94 patients with cri du chat syndrome who had been carefully evaluated for the presence of the characteristic cry, speech delay, facial dysmorphology, and level of mental retardation (MR). Most subjects had simple deletions involving 5p (67 terminal and 12 interstitial). Genotype-phenotype correlations localized the region associated with the cry to 1.5 Mb in distal 5p15.31, between bacterial artificial chromosomes (BACs) containing markers D5S2054 and D5S676; speech delay to 3.2 Mb in 5p15.32-15.33, between BACs containing D5S417 and D5S635; and the region associated with facial dysmorphology to 2.4 Mb in 5p15.2-15.31, between BACs containing D5S208 and D5S2887. These results overlap and refine those reported in previous publications. MR depended approximately on the 5p deletion size and location, but there were many cases in which the retardation was disproportionately severe, given the 5p deletion. All 15 of these cases, approximately two-thirds of the severely retarded patients, were found to have copy-number aberrations in addition to the 5p deletion. Restriction of consideration to patients with only 5p deletions clarified the effect of such deletions and suggested the presence of three regions, MRI-III, with differing effect on retardation. Deletions including MRI, a 1.2-Mb region overlapping the previously defined cri du chat critical region but not including MRII and MRIII, produced a moderate level of retardation. Deletions restricted to MRII, located just proximal to MRI, produced a milder level of retardation, whereas deletions restricted to the still-more proximal MRIII produced no discernible phenotype. However, MR increased as deletions that included MRI extended progressively into MRII and MRIII, and MR became profound when all three regions were deleted.     

A novel proline-rich motif present in ActA of Listeria monocytogenes and cytoskeletal proteins is the ligand for the EVH1 domain, a protein module present in the Ena/VASP family.

The ActA protein of the intracellular pathogen Listeria monocytogenes induces a dramatic reorganization of the actin-based cytoskeleton. Two profilin binding proteins, VASP and Mena, are the only cellular proteins known so far to bind directly to ActA. This interaction is mediated by a conserved module, the EVH1 domain. We identify E/DFPPPPXD/E, a motif repeated 4-fold within the primary sequence of ActA, as the core of the consensus ligand for EVH1 domains. This motif is also present and functional in at least two cellular proteins, zyxin and vinculin, which are in this respect major eukaryotic analogs of ActA. The functional importance of the novel protein-protein interaction was examined in the Listeria system. Removal of EVH1 binding sites on ActA reduces bacterial motility and strongly attenuates Listeria virulence. Taken together we demonstrate that ActA-EVH1 binding is a paradigm for a novel class of eukaryotic protein-protein interactions involving a proline-rich ligand that is clearly different from those described for SH3 and WW/WWP domains. This class of interactions appears to be of general importance for processes dependent on rapid actin remodeling.     

On the possible localization of a gene for triosephosphate isomerase on the short arm of human chromosome 5

Summary Chromosome 5 was examined by fluoresence microscopy in 13 individuals whose karyotype was 5p-. Great variation was observed in length (30–80% of the total short arm length), in type (translocation, interstitial and terminal deletions), and in localization of the deleted segment. There was no evidence of correlation between length, type or localization of deletion and triosephosphate isomerase activity. Triosephosphate isomerase activity of the 13 cases of karyotype 5p- was 2,74 moles/min/mg hemoglobin (95% confidence limits 2.49–3.32). Identical values were obtained in 16 normal individuals; 2.44 moles/min/mg hemoglobin (95% confidence limits 2.16–2.72). The present investigation does not confirm the postulated localization of the gene for triosephosphate isomerase on the short arm of human chromosome 5.

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Zusammenfassung 13 Patienten mit einem 5p—-Karyotyp wurden durch Fluorescenzuntersuchungen identifiziert und das Ausmaß der Deletion bestimmt. Triosephosphatisomeraseaktivitäten dieser 13 Fälle waren 2,74 mol/min/mg Hämoglobin, 95%-Vertrauensgrenzen 2,49–3,32.Identische Werte wurden bei 16 Normalpersonen gefunden (2,44 mol/min/mg Hämoglobin, 95%-Vertrauensgrenzen 2,16–2,72).Diese Befunde lassen sich dahingehend interpretieren, daß keine Assoziation zwischen Triosephosphatisomerase und Chromosom 5 besteht.

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Supported by the Research Committee of the Danish Mental Retardation Service (Project No. 106).     

Polymorphismus der menschlichen Erythrocyten-Glutamat-Pyruvat-Transminase

Zusammenfassung Bei einer Stichprobe nichtverwandter Personen aus der Bevölkerung von Berlin (West) wurden folgende GPT-Frequenzen gefunden: GPT¹ 0,512, GPT² 0,484, GPT³ 0,004. Dieses Ergebnis stimmt mit anderen Untersuchungen aus Deutschland gut überein.

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Polymorphism of the human red cell glutamate-pyruvate transaminase

Summary In a random population sample in Berlin (West) the following GPT-Frequencies were found: GPT¹ 0.512, GPT² 0.484, GPT³ 0.004. This is consistent with other results obtained in German populations.

     

The Cenomanian of northern Germany: facies analysis of a transgressive biosedimentary system

A facies analysis of the epicontinental marine Cenomanian sediments of northern Germany shows the presence of 17 facies types (FTs, including several subtypes) which can be assigned to three facies associations: 1) an inner shelf facies association (FT 1–8) with high amounts of terrigenous material and/or high-energy depositional features, 2) a middle shelf facies association (FT 9–15) of predominantly calcareous sediments with moderate amounts of generally fine siliciclastics, and 3) an outer shelf facies association (FT 16–17) of low-energy, fine-grained, pure limestones. These three facies associations roughly correspond to the well-known lithological units of the Cenomanian of northern Germany, i.e., the Essen Greensand/Cenomanian Marls complex, the Pläner Limestones, and the Poor rhotomagense Limestones. The sediments were deposited on a northward-dipping homoclinal ramp with more-or-less shoreline-parallel facies belts. The sediment composition on this ramp-like shelf was a function of the varying importance of three different sediment sources: 1) terrigenous input from the south (Rhenobohemia), generally fining/decreasing in a proximal–distal (i.e., S–N) direction; 2) production of skeletal grains, mainly by macrobenthic organisms; and 3) settling of planktic carbonate (mainly calcispheres and calcareous nannofossils). In response to decreasing water energy with increasing water depth, the seaward decreasing terrigenous influence, and increasing planktic carbonate production, increasingly finer and more calcareous sediments were deposited in a proximal–distal transect. This rather straightforward picture was slightly modified by highest carbonate accumulation rates (planktic and benthic) on the middle shelf, forming a mid-shelf depocenter (fossiliferous, calcisphere-rich Pläner Limestones). Time-transgressive, southward-directed onlap of this biosedimentary system during the Cenomanian caused a significant retreat of the coastline towards the south and a retrogradational stacking of facies belts, explaining the broadly similar facies development and lithology of Cenomanian successions across northern Germany. The boundaries of the lithological units, however, tend to be considerably diachronous in a distal–proximal transect. In the late Middle and early Late Cenomanian, a final drowning and facies levelling (“oceanization”) is indicated by the widespread deposition of uniform calcareous nannofossil mudstones (Poor rhotomagense Limestones).     

In vitro evaluation of native entomopathogenic fungi and neem (Azadiractha indica) extracts on Spodoptera frugiperda

The control of Spodoptera frugiperda is based on synthetic insecticides, so some alternatives are the use of entomopathogenic fungi (EF) and neem extract. The objective of the study was to evaluate in vitro effectiveness of native EF and neem extracts on S. frugiperda larvae. Six EF were identified by DNA sequencing of ITS regions from three EF (Fusarium solani, Metarrhizium robertsii, Nigrospora spherica and Penicillium citrinum). They were evaluated in concentrations of 1 × 10⁸ spores/ mL. In addition, a second bioassay was carried out evaluating only F. solani, M. robertsii and N. sphaerica and the addition of vegetable oil. On the other hand, extraction of secondary metabolites from neem seed (Azadirachta indica) was carried out by performing, mass (g) and solvent volume (mL ethanol and water) combinations, which were subjected to microwaves and ultrasound. Subsequently, these extracts were evaluated in concentrations of 3%, 4% and 5%. A survival analysis was performed for each of the bioassays. With respect to the results of the first bioassay, F. solani obtained a probability of survival of 0.476 on the seventh day, while in the second bioassay, M. robertsii obtained 0.488 survival probability. This suggests that the expected percentage of larvae that stay alive on the sixth day is 48.8%. However, in the evaluation of the neem extract the combination 1:12/70% to 4% caused 84% mortality of larvae. The use of native HE and neem extracts has potential for the control of S. frugiperda.     

Physical mapping of three fruit ripening genes：Endopolygalacturonase,ACC oxidase and ACC synthase from apple（Malus x domestica） in an apple rootstock A106（Malus sieboldii

The apple rootstock,A106(Malus sieboldii),had 17 bivalents in pollen mother cells at meiotic metaphase 1,and 17 chromosomes in a haploid pollen cell.Karyotypes were prepared from root-tip cells with 2n=34 chromosomes,Seven out of 82 karyotypes(8.5%) showed one pari of satellites at the end of the short arm of chromosome 3.C-bands were shown on 6 pairs of chromosomes 2,4,6,8,14,and 16 near the telomeric regions of short arms.Probes for three ripening-related genes from Malus x domestica:endopolygalacturonase(EPG,0.6kb),ACC oxidase(1.2kb),and ACC synthase(2kb)were hybridized in situ to metaphase chromosomes of A106.Hybridization sites for the EPG gene were observed on the long arm of chromosome 14 in 15 out of 16 replicate spreads and proximal to the centromere of chromosomes 6 and 11.For the ACC oxidase gene,hylridization sites were observed in the telomeric region of the short arm of chromosomes 5 and 11 in 87% and 81% of 16 spreads respectively,proxiaml to the centromere of chromosome 1 in 81% of the spreads,and on the long arm of chromosome 13 in 50% of the spreads. Physical mapping of three fruit ripening genes in an apple rootstock A106.Twenty five spreads were studied for the ACC synthase gene and hybridization sites were observed in the telomeric region of the short arm of chromosome 12 in 96% of the spreads.chromosomes 9 and 10 in 76% of the spreads,and chromosome 17 in 56% of the spreads.     

The cytokineplast: purified, stable, and functional motile machinery from human blood polymorphonuclear leukocytes

We examined the formation of motile, chemotactically active, anucleate fragments from human blood polymorphonuclear leukocytes (PMN, granulocytes), induced by the brief application of heat. These granule-poor fragments are former protopods (leading fronts, lamellipodia) that become uncoupled from the main body of the cell and leave it, at first with a connecting filament that breaks and seals itself. The usual random orientation of such filaments can be controlled by preorientation of cells in a gradient of the chemotactic peptide, N-formylmethionylleucylphenylalanine (F-Met-Leu-Phe) (2x10(-9) M- 1x10(-8)). Cytochalsin B, 2.5-5 μg/ml, prevents fragment formation; colchicine, 10(-5) M, does not. In scanning electron micrographs, fragments are ruffled and the cell body rounded up and rather smooth. In transmission electron micrographs, fragments contain microfilaments but lack centrioles and microtubules. Like intact cells, both bound and free fragments can respond chemotactically to an erythrocyte destroyed by laser microirradiation (necrotaxis); the free, anucleate fragments may do so repeatedly, even after having been held overnight at ambient temperatures. We propse the name cytokineplast for the result of this self-purification of motile apparatus. The exodus of the motile machinery from the granulocyte requires anchoring of the bulk of the cell to glass and uncoupling, which may involve heat-induced dysfunction of the centrosome. In ultrastructural studies of the centrosomal region after heat, centriolar structure remains intact, but pericentriolar osmiophilic material appears condensed, and microtubules are sparse. These changes are found in all three blood cell types examined: PMN, eosinophil, and monocyte. Of these, the first two make fragments under our conditions; the more sluggish monocyte does not. Uncoupling is further linked to centrosomal dysfunction by the observation that colchicines-treated granulocytes (10(-5)M, to destroy the centrosome’s efferent arm) make fragments after less heat than controls. If motive force and orientation are specified mainly from the organelle-excluding leading front, then endoplasmic streaming in PMN is a catch-up phenomenon, and microtubules do not provide the vector of locomotion but rather stabilize and orient the “baggage” (nucleus, granuloplasm)—i.e., they prevent fishtailing. Moreover, constraints emanating from the centrosome may now be extended to include, maintenance of the motile machinery as an integral part of the cell.     

Unusual findings by fluorescence microscopy of a t(13q14q)

Summary Fluorescence studies in a 13q14q translocation seem to indicate, that the centromeric region of both chromosomes has been preserved. It is assumed, that the 13 centromere, not visible in Giemsa staining, is suppressed by the active 14 centromere. It is emphasized that Robertsonian translocations in man might be due to breaks in the short arms of the involved chromosomes.

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Zusammenfassung Die Färbung einer 13q14q-Translokation mit Quinacrin-Lost scheint zu indicieren, daß die Zentromerregionen beider Chromosomen erhalten sind. Das Zentromer des Chromosoms Nr. 13 ist nicht sichtbar bei Giemsafärbung. Es ist möglich, daß es durch das aktive 14-Zentromer unterdrückt worden ist. Zentrische Fusionen beim Menschen können somit wahrscheinlich durch Brüche an beiden kurzen Armen der implizierten Chromosomen entstehen.