Outer Dense Fiber 2 Is a Widespread Centrosome Scaffold Component Preferentially Associated with Mother Centrioles: Its Identification from Isolated Centrosomes

Because centrosomes were enriched in the bile canaliculi fraction from the chicken liver through their association with apical membranes, we developed a procedure for isolation of centrosomes from this fraction. With the use of the centrosomes, we generated centrosome-specific monoclonal antibodies. Three of the monoclonal antibodies recognized an antigen of ∼90 kDa. Cloning of its cDNA identified this antigen as a chicken homologue of outer dense fiber 2 protein (Odf2), which was initially identified as a sperm outer dense fiber-specific component. Exogenously expressed and endogenous Odf2 were shown to be concentrated at the centrosomes in a microtubule-independent manner in various types of cells at both light and electron microscopic levels. Odf2 exhibited a cell cycle-dependent pattern of localization and was preferentially associated with the mother centrioles in G0/G1-phase. Toward G1/S-phase before centrosome duplication, it became detectable in both mother and daughter centrioles. In the isolated bile canaliculi and centrosomes, Odf2, in contrast to other centrosomal components, was highly resistant to KI extraction. These findings indicate that Odf2 is a widespread KI-insoluble scaffold component of the centrosome matrix, which may be involved in the maturation event of daughter centrioles.

Duplication and functional divergence in the chalcone synthase gene family of Asteraceae: evolution with substrate change and catalytic simplification.

Plant-specific polyketide synthase genes constitute a gene superfamily, including universal chalcone synthase [CHS; malonyl-CoA:4-coumaroyl-CoA malonyltransferase (cyclizing) (EC 2.3.1.74)] genes, sporadically distributed stilbene synthase (SS) genes, and atypical, as-yet-uncharacterized CHS-like genes. We have recently isolated from Gerbera hybrida (Asteraceae) an unusual CHS-like gene, GCHS2, which codes for an enzyme with structural and enzymatic properties as well as ontogenetic distribution distinct from both CHS and SS. Here, we show that the GCHS2-like function is encoded in the Gerbera genome by a family of at least three transcriptionally active genes. Conservation within the GCHS2 family was exploited with selective PCR to study the occurrence of GCHS2-like genes in other Asteraceae. Parsimony analysis of the amplified sequences together with CHS-like genes isolated from other taxa of angiosperm subclass Asteridae suggests that GCHS2 has evolved from CHS via a gene duplication event that occurred before the diversification of the Asteraceae. Enzyme activity analysis of proteins produced in vitro indicates that the GCHS2 reaction is a non-SS variant of the CHS reaction, with both different substrate specificity (to benzoyl-CoA) and a truncated catalytic profile. Together with the recent results of Durbin et al. [Durbin, M. L., Learn, G. H., Jr., Huttley, G. A. & Clegg, M. T. (1995) Proc. Natl. Acad. Sci. USA 92, 3338-3342], our study confirms a gene duplication-based model that explains how various related functions have arisen from CHS during plant evolution.

Chromosomal localization of the proteasome Z subunit gene reveals an ancient chromosomal duplication involving the major histocompatibility complex.

Proteasomes are the multi-subunit protease thought to play a key role in the generation of peptides presented by major histocompatibility complex (MHC) class I molecules. When cells are stimulated with interferon gamma, two MHC-encoded subunits, low molecular mass polypeptide (LMP) 2 and LMP7, and the MECL1 subunit encoded outside the MHC are incorporated into the proteasomal complex, presumably by displacing the housekeeping subunits designated Y, X, and Z, respectively. These changes in the subunit composition appear to facilitate class I-mediated antigen presentation, presumably by altering the cleavage specificities of the proteasome. Here we show that the mouse gene encoding the Z subunit (Psmb7) maps to the paracentromeric region of chromosome 2. Inspection of the mouse loci adjacent to the Psmb7 locus provides evidence that the paracentromeric region of chromosome 2 and the MHC region on chromosome 17 most likely arose as a result of a duplication that took place at an early stage of vertebrate evolution. The traces of this duplication are also evident in the homologous human chromosome regions (6p21.3 and 9q33-q34). These observations have implications in understanding the genomic organization of the present-day MHC and offer insights into the origin of the MHC.

Minimal definition of the imprinting center and fixation of chromosome 15q11-q13 epigenotype by imprinting mutations.

Patients with disorders involving imprinted genes such as Angelman syndrome (AS) and Prader-Willi syndrome (PWS) can have a mutation in the imprinting mechanism. Previously, we identified an imprinting center (IC) within chromosome 15q11-ql3 and proposed that IC mutations block resetting of the imprint, fixing on that chromosome the parental imprint (epigenotype) on which the mutation arose. We now describe four new microdeletions of the IC, the smallest (6 kb) of which currently defines the minimal region sufficient to confer an AS imprinting mutation. The AS deletions all overlap this minimal region, centromeric to the PWS microdeletions, which include the first exon of the SNRPN gene. None of five genes or transcripts in the 1.0 Mb vicinity of the IC (ZNF127, SNRPN, PAR-5, IPW, and PAR-1), each normally expressed only from the paternal allele, was expressed in cells from PWS imprinting mutation patients. In contrast, AS imprinting mutation patients show biparental expression of SNRPN and IPW but must lack expression of the putative AS gene 250-1000 kb distal of the IC. These data strongly support a model in which the paternal chromosome of these PWS patients carries an ancestral maternal epigenotype, and the maternal chromosome of these AS patients carries an ancestral paternal epigenotype. The IC therefore functions to reset the maternal and paternal imprints throughout a 2-Mb imprinted domain within human chromosome 15q11-q13 during gametogenesis.

A serine proteinase inhibitor locus at 18q21.3 contains a tandem duplication of the human squamous cell carcinoma antigen gene.

The squamous cell carcinoma antigen (SCCA) is a member of the ovalbumin family of serine proteinase inhibitors (serpins). A neutral form of the protein is found in normal and some malignant squamous cells, whereas an acidic form is detected exclusively in tumor cells and in the circulation of patients with squamous cell tumors. In this report, we describe the cloning of the SCCA gene from normal genomic DNA. Surprisingly, two genes were found. They were tandemly arrayed and flanked by two other closely related serpins, plasminogen activator inhibitor type 2 (PAI2) and maspin at 18q21.3. The genomic structure of the two genes, SCCA1 and SCCA2, was highly conserved. The predicted amino acid sequences were 92% identical and suggested that the neutral form of the protein was encoded by SCCA1 and the acidic form was encoded by SCCA2. Further characterization of the region should determine whether the differential expression of the SCCA genes plays a causal role in development of more aggressive squamous cell carcinomas.

Large national series of patients with Xq28 duplication involving MECP2: Delineation of brain MRI abnormalities in 30 affected patients.

Xq28 duplications encompassing MECP2 have been described in male patients with a severe neurodevelopmental disorder associated with hypotonia and spasticity, severe learning disability, stereotyped movements, and recurrent pulmonary infections. We report on standardized brain magnetic resonance imaging (MRI) data of 30 affected patients carrying an Xq28 duplication involving MECP2 of various sizes (228 kb to 11.7 Mb). The aim of this study was to seek recurrent malformations and attempt to determine whether variations in imaging features could be explained by differences in the size of the duplications. We showed that 93% of patients had brain MRI abnormalities such as corpus callosum abnormalities (n = 20), reduced volume of the white matter (WM) (n = 12), ventricular dilatation (n = 9), abnormal increased hyperintensities on T2-weighted images involving posterior periventricular WM (n = 6), and vermis hypoplasia (n = 5). The occipitofrontal circumference varied considerably between >+2SD in five patients and <-2SD in four patients. Among the nine patients with dilatation of the lateral ventricles, six had a duplication involving L1CAM. The only patient harboring bilateral posterior subependymal nodular heterotopia also carried an FLNA gene duplication. We could not demonstrate a correlation between periventricular WM hyperintensities/delayed myelination and duplication of the IKBKG gene. We thus conclude that patients with an Xq28 duplication involving MECP2 share some similar but non-specific brain abnormalities. These imaging features, therefore, could not constitute a diagnostic clue. The genotype-phenotype correlation failed to demonstrate a relationship between the presence of nodular heterotopia, ventricular dilatation, WM abnormalities, and the presence of FLNA, L1CAM, or IKBKG, respectively, in the duplicated segment.

Complete Urethral Duplication in Children: A Case Report

Introduction: Urethral duplication (UD) is a rare congenital anomaly with multiple anatomical variants. Case Presentation: In this article we present a four year-old child with complete UD. The patient was admitted for hypospadias repair, in evaluation we found type IIA1 UD according to Effmann classification. Patient underwent hypospadias repair saving complete UD. Conclusions: After one year follow-up he has normal and continent urination.