Discordant sex in monozygotic XXY/XX twins: a case report.

We report a case of discordant phenotypic sex in monozygotic twins mosaic 47,XXY/46,XX: monozygotic heterokaryotypic twins. The twins presented with cognitive and comprehension delay, behavioural and language disorders, all symptoms frequently reported in Klinefelter syndrome. Molecular zygosity analysis with several markers confirmed that the twins are in effect monozygotic (MZ). Array comparative genomic hybridization found no evidence for the implication of copy number variation in the phenotypes. Ultrasound scans of the reproductive organs revealed no abnormalities. Endocrine tests showed a low testosterone level in Twin 1 (male phenotype) and a low gonadotrophin level in Twin 2 (female phenotype) which, combined with the results from ultrasound examination, provided useful information for potentially predicting the future fertility potential of the twins. Blood karyotypes revealed the presence of a normal 46,XX cell line and an aneuploïd 47,XXY cell line in both patients. Examination of the chromosome constitutions of various tissues such as blood, buccal smear and urinary sediment not surprisingly showed different proportions for the 46,XX and 47,XXY cell lines, which most likely explains the discordant phenotypic sex and mild Klinefelter features. The most plausible underlying biological mechanism is a post-zygotic loss of the Y chromosome in an initially 47,XXY zygote. This would result in an embryo with both 46,XX and 47,XXY cells lines which could subsequently divide into two monozygotic embryos through a twinning process. The two cell lines would then be distributed differently between tissues which could result in phenotypic discordances in the twins. These observations emphasize the importance of regular paediatric evaluations to determine the optimal timing for fertility preservation measures and to detect new Klinefelter features which could appear throughout childhood in the two subjects.

Distribution of olfactory receptor genes in the human genome.

We demonstrate that members of the olfactory receptor (OR) gene family are distributed on all but a few human chromosomes. Through FISH analysis, we show that OR sequences reside at more than 25 locations in the human genome. Their distribution is biased for terminal bands. Flow-sorted chromosomes were used to isolate 87 OR sequences derived from 16 chromosomes. Their sequence-relationships are indicative of the inter- and intrachromosomal duplications responsible for OR family expansion. The human genome has accumulated a striking number of dysfunctional copies: 72% of the sequences are pseudogenes. ORF-containing sequences predominate on chromosomes 7, 16 and 17.

The human necdin gene, NDN, is maternally imprinted and located in the Prader-Willi syndrome chromosomal region.

Prader-Willi syndrome (PWS) is a neurogenetic disorder that results from the absence of a normal paternal contribution to the 15q11-13 region. The clinical manifestations of PWS are a transient severe hypotonia in the newborn period, with mental retardation, hypogonadism and obesity observed later in development. Five transcripts with exclusive expression from the paternal allele have been isolated, but none of these has been shown to be involved in PWS. In this study, we report the isolation and characterization of NDN, a new human imprinted gene. NDN is exclusively expressed from the paternal allele in the tissues analysed and is located in the PWS region. It encodes a putative protein homologous to the mouse brain-specific NECDIN protein, NDN; as in mouse, expression in brain is restricted to post-mitotic neurons. NDN displays several characteristics of an imprinted locus, including allelic DNA methylation and asynchronous DNA replication. A complete lack of NDN expression in PWS brain and fibroblasts indicates that the gene is expressed exclusively from the paternal allele in these tissues and suggests a possible role of this new gene in PWS.

[Revelation of an acute lymphoblastic leukemia in the delivery room]. / Leucémie aiguë lymphoblastique néonatale: une affection rare à révélation immédiate en salle de naissance.

Acute leukemia is uncommon in neonates and has a much poorer prognosis than in older children. We report on a case of acute lymphoblastic leukemia observed in a neonate who had bleeding and hepatosplenomegaly at birth, which justified intensive care during the first postnatal week. Despite early appropriate treatment, the patient died at 7 months of age. We present here physical and laboratory findings, which indicate a grim prognosis. These criteria should be considered carefully in order to ensure a realistic information for the parents and appropriate decisions.

The human growth factor-inducible immediate early gene, CYR61, maps to chromosome 1p.

Complementary DNA encoding the human CYR61 protein was isolated from human embryonic tissues and mapped to chromosome 1p22-p31. We show that CYR61 encodes a 381 amino acid protein rich in cysteine and proline residues that is strongly conserved with the mouse homologue. Sequence analysis reveals the presence of several distinct protein domains which confer a mosaic structure to this protein and makes human CYR61 a member of a recently described growth regulator family that includes several proto-oncogene products. From our results we hypothesize that this new immediate early gene may play a role in cell commitment during embryogenesis and more generally in the control of cell proliferation.

Cloning, expression and subcellular localization of the human homolog of p40MO15 catalytic subunit of cdk-activating kinase.

Transitions of the cell cycle are controlled by cyclin-dependent protein kinases (cdks) whose phosphorylation on the Thr residue included in the conserved sequence YTHEVV dramatically increases the activity. A kinase responsible for this specific phosphorylation, called CAK for cdk-activating kinase, has been recently purified from starfish and Xenopus oocytes and shown to contain the MO15 gene product as a catalytic subunit. In the present paper, we have cloned the human homolog of Xenopus p40MO15 by probing a HeLa cell cDNA library with degenerate oligonucleotides deduced from Xenopus and starfish MO15 sequences. Human and Xenopus MO15 displayed a strong homology showing 86% identity with regard to amino acid sequences. Northern blot analysis of RNA extracts from a series of human tissues as well as from cultured rodent fibroblasts revealed a unique 1.4 kb MO15 mRNA. No variation in the amount of MO15 transcript or protein was found along the entire course of the fibroblast cell cycle. Fluorescence in situ hybridization on human lymphocyte metaphases showed two distinct chromosomal locations of human MO15 gene at 5q12-q13 and 2q22-q24. By using gene tagging and mammalian cell transfection, we demonstrate that the KRKR motif located at the carboxy terminal end of MO15 is required for nuclear targeting of the protein. Mutation of KRKR to NGER retains MO15 in the cytoplasmic compartment, whilst the wild-type protein is detected exclusively in the nucleus. Interestingly, we demonstrate that the nuclear targeting of MO15 is necessary to confer the protein its CAK activity. In contrast to the wild-type, the NLS-mutated MO15 expressed in Xenopus oocytes is unable to generate CAK as long as the nuclear envelope is not broken. The nuclear localization of both the MO15 gene product and CAK activity may imply that cdks activation primarily occurs in the cell nucleus.

t(5;14)/HOX11L2-positive T-cell acute lymphoblastic leukemia. A collaborative study of the Groupe Français de Cytogénétique Hématologique (GFCH).

To accurately estimate the incidence of HOX11L2 expression, and determine the associated cytogenetic features, in T-cell acute lymphoblastic leukemia (T-ALL), the Groupe Français de Cytogénétique Hématologique (GFCH) carried out a retrospective study of both childhood and adult patients. In total, 364 patients were included (211 children

Heterotaxia syndrome and autosomal dominant inheritance.

Previous familial cases of recurrent heterotaxia have suggested an autosomal recessive or exceptionally X-linked or dominant inheritance. Here, we report six families including 18 affected members, consistent with autosomal dominant inheritance. Among these, four families have more than one case of heterotaxia. The other two families have one member with heterotaxia and at least one other affected member with an "isolated" heart malformation, which could be considered as a mild form of heterotaxia. In five families, the disorder is transmitted through two or three generations. In one family, the patients are of the same generation but are linked to each other by obligate carriers. We suggest a rule to classify these families with heart malformations, according to the etiologic factor involved (rule of precocity). This rule might be useful to other disruptions of morphogenetic processes.

5-HT4 receptors: cloning and expression of new splice variants.

On the basis of differences in the potencies and intrinsic activity of 5-HT4 receptor agonists in different biological models it has been suggested that there is heterogeneity among 5-HT4 receptors. Here, we report the molecular cloning of several 5-HT4 receptor splice variants in mouse, rat, and human brain. Our data suggest that the differences in efficacy of 5-HT4 ligands on 5-HT4 receptor-mediated responses in several tissues is due to differences in coupling efficiency rather than to the presence of different 5-HT4 receptor isoforms.

Heterogeneous patterns of amplification of the NUP214-ABL1 fusion gene in T-cell acute lymphoblastic leukemia.

Episomes with the NUP214-ABL1 fusion gene have been observed in 6% of T-ALL. In this multicentric study we collected 27 cases of NUP214-ABL1-positive T-ALL. Median age was 15 years with male predominance. Outcome was poor in 12 patients. An associated abnormality involving TLX1 or TLX3 was found in all investigated cases. Fluorescent in situ hybridization revealed a heterogeneous pattern of NUP214-ABL1 amplification. Multiple episomes carrying the fusion were detected in 24 patients. Episomes were observed in a significant number of nuclei in 18 cases, but in only 1-5% of nuclei in 6. In addition, intrachromosomal amplification (small hsr) was identified either as the only change or in association with episomes in four cases and two T-ALL cell lines (PEER and ALL-SIL). One case showed insertion of apparently non-amplified NUP214-ABL1 sequences at 14q12. The amplified sequences were analyzed using array-based CGH.These findings confirm that the NUP214-ABL1 gene requires amplification for oncogenicity; it is part of a multistep process of leukemogenesis; and it can be a late event present only in subpopulations. Data also provide in vivo evidence for a model of episome formation, amplification and optional reintegration into the genome. Implications for the use of kinase inhibitors are discussed.

Expression and human chromosomal localization to 17q25 of the growth-regulated gene encoding the mitochondrial ribosomal protein MRPL12.

Mitochondrial activity requires the expression of nuclear genes, whose products are part of multiproteic complexes leading to ATP production and delivery. We recently characterized a growth-activated mRNA encoding the human mitochondrial ribosomal MRPL12 protein, which is thought to act as a translational regulator of mitochondrial mRNAs. We show here that MRPL12 mRNA expression is enhanced in growth-stimulated cells as a result of transcriptional activation, a feature lost in transformed cell lines. MRPL12 mRNA is highly expressed in the colon, in which a reduction in mitochondrial activity was shown to be associated with tumor formation. The human MRPL12 protein is encoded by a unique gene located on chromosome 17 (q25-qter). As no predisposition to colon cancer linked to this chromosomal region was hitherto reported, the MRPL12 gene might be involved in the process of differentiation of colonic epithelial cells.

Localization of human cell cycle regulatory genes CDC25C to 5q31 and WEE1 to 11p15.3-11p15.1 by fluorescence in situ hybridization.

The cell cycle control genes are highly conserved during evolution since they play a key role in the regulation of cell division. We have localized CDC25C and WEE1 respectively at 5q31 and 11p15.3-11p15.1 using fluorescent in situ hybridization of cDNA probes on human chromosomes. This shows that genes acting through a regulatory phosphorylation cascade are not clustered on the same chromosome. Furthermore, they appear to map on chromosomal regions involved in tumorigenesis. The 5q23-q31 region of chromosome 5 is deleted in some hematologic disorders, and the p15 region of chromosome 11 is involved in development of embryonic tumors.

[Diagnosis of minor chromosome modifications by molecular cytogenetics]. / Diagnostic de remaniements chromosomiques de petite taille par la cytogénétique moléculaire.

Chromosomal in situ hybridization with radioactive probes allows the detection of single copy DNA segments of very small size. In two phenotypically normal individuals, classical chromosomal analysis revealed a small partial deletion of chromosome 18 short arm. In situ hybridization of probe D18S3, located at 18p13, showed in both instances a balanced reciprocal translocation: 46,XY,t(5;18) (p153;p112) and 46,XX,t(18;22)(p111;p11), respectively.

Interstitial deletion of chromosome 1 del (1) (q32 q42): case report and review of the literature.

We describe the case of a female infant with multiple congenital anomalies who was found to have a de novo distal intestinal del (1) (q32 q42). The clinical features of other reports of similar deletions are briefly reviewed. No characteristic phenotype seems to be as yet definable due to the limited number of cases published.

Assignment of human genes for beta 2 and beta 4 subunits of voltage-dependent Ca2+ channels to chromosomes 10p12 and 2q22-q23.

We have used human beta 2 and beta 4 cDNA probes to map the genes encoding two isoforms of the regulatory beta subunit of voltage-activated Ca2+ channels, viz. CACNB2 (beta 2) and CACNB4 (beta 4), to human chromosomes 10p12 and 2q22-q23, respectively, by fluorescence in situ hybridization. The gene encoding the beta 2 protein, first described as a Lambert-Eaton myasthenic syndrome (LEMS) antigen in humans, is found close to a region that undergoes chromosome rearrangements in small cell lung cancer, which occurs in association with LEMS. CACNB2 (beta 2) and CACNB4 (beta 4) genes are members of the ion-channel gene superfamily and it should now be possible to examine their loci by linkage analysis of ion-channel-related disorders. To date, no such disease-related gene has been assigned to 10p12 and 2q22-q23.

Cloning, chromosomal location and functional expression of the human voltage-dependent calcium-channel beta 3 subunit.

A novel human-voltage-dependent-calcium-channel (VDCC) beta subunit was isolated from a 9-week-old human total-embryo cDNA library. Of the four genes encoding beta-subunit isoforms that have been identified in animal species, this isoform shares strong similarity with the rat and rabbit beta 3-related gene product and is referred to here as H beta 3 subunit. The H beta 3 isoform is the second beta subunit identified in human. Its open reading frame encodes a 482-amino-acid protein with a predicted molecular mass of 54.571 kDa. The H beta 3 mRNA is expressed mostly in brain, smooth muscle and ovary. The gene for the human H beta 3 was specifically localized on chromosome 12q13. The cloned H beta 3 subunit was further expressed in Xenopus oocytes to demonstrate its ability to modulate VDCC activity.

Human cardiac troponin T: cloning and expression of new isoforms in the normal and failing heart.

Troponin T, like many myofibrillar proteins, exists as multiple isoforms encoded by distinct genes or generated by splicing of the same primary RNA transcript. We have previously cloned the first human cardiac troponin T (cTnT) cDNA and showed the differential expression of cTnT in cardiac and skeletal muscle during ontogenic development. In this work we located the human cTnT gene by means of fluorescent in situ hybridization to 1q32 and, by sequencing thirteen cDNAs isolated from a human fetal heart cDNA library, identified three new isoforms resulting from specific combinations of three variable regions in human cTnT cDNA. The first variable region is a 30-bp box located at the 5' end of the cDNA, which can be excised either totally or only from the first 3 bp onwards; the second is a codon which can be completely excised; and the third is a 9-bp box in the 3' half of the cDNA, which can also be excised either totally or only from the first 3 bp. The existence of the corresponding RNAs in fetal and adult ventricles was confirmed by RNase protection studies. No accumulation of the fetal isoforms was found in failing ventricles compared with controls.

Characterization of the cDNA and pattern of expression of a new gene over-expressed in human hepatomas and colonic tumors.

A cDNA clone of 6.449 kb ch-TOG (for colonic and hepatic tumor over-expressed gene) initially selected from various human libraries and completed by 5' rapid amplification of cDNA ends (RACE) PCR is described. The original cDNA clone was extracted from an expression library constructed from a human tumoral brain. This library was screened with an antibody raised against the cytochrome P450tu that was shown to be over-expressed in chemically induced mouse hepatic tumors. Using this cDNA as a probe, a full-length cDNA was characterized. Its nucleotide sequence shows no significant similarity with any of the gene sequences collected in the various DNA data bases. The translation of the larger open reading frame leads to a putative protein of 1972 amino acids (molecular mass = 218453 Da). Hybridization analyses on Southern blot and on metaphase chromosomes indicate that this gene is present as a single copy in the genome and is localized on the short arm of chromosome 11. ch-TOG transcripts are present in several human tissues. Over-expression of ch-TOG in neoplastic liver and colon compared with the corresponding normal corresponding tissues is demonstrated. The level of the expression of ch-TOG transcripts was also studied in the various differentiation stages of the human colonic adenocarcinoma cell line Caco-2.

Molecular analysis of the sex-determining region from the Y chromosome in two patients with Frasier syndrome.

In the Frasier syndrome there is an association between XY gonadal dysgenesis and chronic renal failure. Owing to an observed sex reversal, the Y chromosomes of two girls with this syndrome have been analyzed. Using molecular-biology techniques, no major alterations of the known sex-determining area of the Y chromosome were found. Furthermore, the sequence did not reveal impairment of the recently described testis-determining factor SRY. These data suggest that in the Frasier syndrome, XY sex reversal and renal failure could be the result of either faulty gene(s) located downstream in the sex differentiation pathway during embryogenesis, or impaired SRY regulation. Preliminary results on the Wilms' tumor suppressor gene WT1, a candidate for acting downstream to SRY, are also provided.

SOX22 is a new member of the SOX gene family, mainly expressed in human nervous tissue.

SOX (SRY box-containing) genes share a particular DNA-binding domain, called HMG, with the mammalian testis-determining gene SRY Several SOX genes have already been shown to be transcription factors involved in the decision of important cell fates during development. Here we report the cloning of a new human member of the SOX gene family, SOX22. The corresponding protein contains several domains that are also present in other paralogous SOX proteins. The SOX22 gene maps to chromosome 20 on band p13 and does not appear to be clustered with any other SOX gene mapped to date. SOX22 mRNA is expressed in various fetal and adult organs and tissues, suggesting that this gene plays roles in both differentiation and maintenance of several cell types.