Hotspot for deletions in the CGG repeat region of FMR1 in fragile X patients

The fragile X syndrome is the most frequent cause of inheritedmental retardation. The molecular mechanism of the disorderis based on the expansion of a CGG repeat in the 5' UTR of theFMR1 gene In the majority of fragile X patients. The instabilityof this CGG repeat containing region is not restricted to theCGG repeat Itself but expands to the flanking region as well.We describe four unrelated fragile X patients that are mosaicfor both a full mutation and a small deletion in the CGG repeatcontaining region. Sequence analysis of the regions surroundingthe deletions showed that both the (CGG)_n repeat and some flankingsequences were missing in all four patients. The 5' breakpointsof the deletions were found to be located between 75–53bp proximal to the CGG repeat. This suggests the presence ofa hot spot region for deletions in the CGG repeat region ofthe FMR1 gene and emphasizes the instability of this regionIn the presence of an expanded CGG repeat.     

Visualization of erythrocyte membrane antigen by means of fluorescent microspheres coupled to monoclonal mouse auto-antibody: Distribution of this antigen among species

Mouse peritoneal cells (PC) in culture produce auto-antibodies lyzing bromelain-treated mouse erythrocytes (MRBCbr). These auto-antibodies have been obtained in an homogeneous form in substantial amounts after cell fusion of PC with myeloma X63.Ag8. They have been identified as the anti-Hb-auto-antibodies described by us in 1980. Once coupled to fluorescent microspheres (Ms), they were used to detect the corresponding antigen. It was found that the specific antigen was not only present on the surface of all MRBCbr but also, in a much smaller proportion, on some normal MRBC. Its distribution is not restricted to the mouse: pigeon RBC is stained heavily; human red cells give, more or less, positive reaction, according to their blood group. Some species, as horse RBC, are consistently negative. The opportunity offered by the fluorescent microspheres technique to trace the antigen recognized by the Hb-auto-antibody in the mouse tissues and on cells from other species should lead to a better understanding of the cross-antigenicity of many RBC and of the peculiar auto-immune process involving MRBCbr in the mouse.     

ThinPrep evaluation of fluid samples aspirated from cystic ovarian masses

The cytological evaluation of ovarian cystic fluid using ThinPrep has not been reported. To determine the diagnostic accuracy of ThinPrep cytology in distinguishing between benign and nonbenign ovarian cystic lesions, we examined 65 fluid samples aspirated during intraoperative consultation with subsequent histologic correlation. One ThinPrep slide was prepared from each sample aspirated from surgically removed ovarian cystic masses and reviewed blindly by a panel of three cytopathologists. The parameters used in cytological evaluation were cellularity, cell types, cellular arrangement, and background. Four samples were acellular and excluded from the study. The consensus cytologic diagnoses were compiled for 61 cases which were assigned to one of the following diagnostic categories: negative for malignant cells (40 cases), atypical cytology (13 cases), and suspicious or positive for malignancy (8 cases). Histologic correlation of the cytological benign/negative cases showed that 26/40 (65%) were histologically benign and 14/40 were false-negative (35%, 5 carcinomas and 9 borderline tumors) with 10 of these cases being mucinous tumors. Most false-negative cytologic samples (11/14 or 79%) did not have an epithelial component. Of the 21 cytological nonbenign diagnoses (atypical/suspicious/positive), 15 (71%) were confirmed on histology (10 carcinomas and 5 borderline tumors). However, a nonbenign cytologic diagnosis was rendered in 6 histologically benign cases, including 2 serous cystadenomas, 1 mucinous cystadenoma, 1 serous cystadenofibroma, 1 endometriosis, and 1 corpus luteal cyst. The diagnostic sensitivity by ThinPrep evaluation of ovarian cystic masses is 81% (26/32) for benign and 52% (15/29) for nonbenign lesions. Our results concluded that ThinPrep examination of ovarian cystic fluid is not accurate in distinguishing benign from malignant cysts, given the significant number of false-negative diagnoses. Major contributing factors include sparse cellularity of the fluid samples and mucinous differentiation of the tumors.     

Structural and functional characterization of mutants of the bovine leukemia virus transactivator protein p34

Mutants of the bovine leukemia virus (BLV) transactivator protein (tat, tax, p34, the XLOR gene product) were constructed by site-directed deletions, in-phase linker insertions, or fragment replacements (swapping) among BLV variants. The mutant constructs were transfected into cos cells and transiently expressed. Western blot analysis using a mixture of monoclonal antibodies to wild-type p34 revealed the presence of mutated XLOR gene products in all the mutants tested. The transactivating activity of 11 tax mutants containing site-directed deletions and in-phase linker insertions was completely abolished. Only the swapping mutant tested, a hybrid between two BLV variants, transactivated LTR-directed gene expression at wild-type levels. These data illustrate the narrow range of structural variations that allow full activity of the BLV tax product and suggest that the present molecular structure of the transactivator protein results from heavy evolutionary constraints.     

Identification of loci determining susceptibility to the lethal effects of amyloid precursor protein transgene overexpression

Phenotypes produced by expression of human amyloid precursor protein (APP) transgenes vary depending on the genetic background of the mouse. FVB/N mice overexpressing human APP695 develop a central nervous system disorder and die prematurely, precluding development of Abeta peptide amyloid plaques. 129S6 mice are resistant to the lethal effects of APP overexpression, allowing sufficient levels of Abeta expression for the development of amyloid plaques and age-dependent memory deficits. To identify the genes that determine susceptibility or resistance to APP we analyzed crosses involving FVB/NCr and 129S6.Tg2576 mice that overexpress 'Swedish' mutant (K670N, M671L) APP695. APP transgene-positive FVB129S6F1 (F1) mice are resistant to the lethal effects of APP overexpression, so FVBxF1 backcross and F2 intercross offspring were produced. Analysis of age of death as a quantitative trait revealed significant linkage to loci on proximal chromosome 14 and on chromosome 9; 129S6 alleles protect against the lethal effects of APP. Within the chromosome 14 interval are segments homologous to regions on human chromosome 10 that have been linked to late onset Alzheimer's disease or to levels of Abeta peptide in plasma. However, analysis of plasma Abeta peptide concentrations at 6 weeks in backcross offspring produced no significant linkage. Similarly, elevation of human Abeta peptide concentrations by expression of mutant presenilin transgenes did not increase the proportion of mice dying prematurely, suggesting that early death reflects effects of APP or fragments other than Abeta.     

Autosomal dominant inheritance of left ventricular outflow tract obstruction

Most nonsyndromic congenital heart malformations (CHMs) in humans are multifactorial in origin, although an increasing number of monogenic cases have been reported recently. We describe here four new families with presumed autosomal dominant inheritance of left ventricular outflow tract obstruction (LVOTO), consisting of hypoplastic left heart (HLHS) or left ventricle (HLV), aortic valve stenosis (AS) and bicuspid aortic valve (BAV), hypoplastic aortic arch (HAA), and coarctation of the aorta (CoA). LVOTO in these families shows a wide clinical spectrum with some family members having severe anomalies such as hypoplastic left heart, and others only minor anomalies such as mild aortic valve stenosis. This supports the suggestion that all anomalies of the LVOTO spectrum are developmentally related and can be caused by a single gene defect.     

L1 knockout mice show dilated ventricles, vermis hypoplasia and impaired exploration patterns

L1 is a neural cell adhesion molecule mainly involved in axon guidance and neuronal migration during brain development. Mutations in the human L1 gene give rise to a complex clinical picture, with mental retardation, neurologic abnormalities and a variable degree of hydrocephalus. Recently, a transgenic mouse model with a targeted null mutation in the L1 gene was generated. These knockout (KO) mice show hypoplasia of the corticospinal tract. Here we have performed further studies of these KO mice including magnetic resonance imaging of the brain, neuropathological analysis and behavioral testing. The ventricular system was shown to be abnormal with dilatation of the lateral ventricles and the 4th ventricle, and an altered shape of the Sylvius aqueduct. Additionally, the cerebellar vermis of the KO mice is hypoplastic. Their exploratory behavior is characterized by stereotype peripheral circling reminiscent of that of rodents with induced cerebellar lesions.      

Low Spatial Frequency Sensitivity and Emotional Face Processing in Adolescents: An Event-related Potential Study

ABSTRACT

Slow maturation of visual pathways transmitting low spatial frequency (LSF) information may contribute to inaccurate facial emotion recognition in adolescence. We recorded ERPs from adolescents and adults to upright and inverted happy faces, fearful faces, and chairs, which were unfiltered, contained only LSFs, or only high spatial frequencies. P100s and N170s were larger for adolescents than adults, with the greatest effect size for LSF stimuli. For LSFs only, adolescents showed a larger N170 inversion effect for happy than for fearful faces, but adults showed the opposite response. Thus, immaturities in LSF pathways appear to impact facial expression processing in adolescents.     

Transient requirement of the PrrA-PrrB two-component system for early intracellular multiplication of Mycobacterium tuberculosis

Adaptive regulation of gene expression in response to environmental changes is a general property of bacterial pathogens. By screening an ordered transposon mutagenesis library of Mycobacterium tuberculosis, we have identified three mutants containing a transposon in the coding sequence or in the 5' regions of genes coding for two-component signal transduction systems (trcS, regX3, prrA). The intracellular multiplication capacity of the three mutants was investigated in mouse bone marrow-derived macrophages. Only the prrA mutant showed a defect in intracellular growth during the early phase of infection, and this defect was fully reverted when the mutant was complemented with prrA-prrB wild-type copies. The mutant phenotype was transient, as after 1 week this strain recovered full growth capacity to reach levels similar to that of the wild type at day 9. Moreover, a transient induction of prrA promoter activity was observed during the initial phase of macrophage infection, as shown by a prrA promoter-gfp fusion in M. bovis BCG infecting the mouse macrophages. The concordant transience of the prrA mutant phenotype and prrA promoter activity indicates that the PrrA-PrrB two-component system is involved in the environmental adaptation of M. tuberculosis, specifically in an early phase of the intracellular growth, and that, similar to other facultative intracellular parasites, M. tuberculosis can use genes temporarily required at different stages in the course of macrophage infection.