A de novo satellited short arm of the Y chromosome possibly resulting from an unstable translocation

A satellited long arm of the Y chromosome (Yqs) is considered a normal variation, whereas the presence of a satellite on the short arm of the Y (Yps) has never been described in the literature. A Yps chromosome could be clinically significant if the translocation resulting in Yps has relocated the testis-determining gene, SRY, to another chromosome. A carrier of such a translocation would therefore be at increased risk for having XX male and XY female offspring. Here we describe the first reported case of de novo Yps present in a phenotypically normal male. This Yps chromosome was positive for C-banding and nucleolus organizer region (NOR) staining and showed a hybridization signal for the beta-satellite sequence. Fluorescence in situ hybridization (FISH) analysis indicated that SRY was retained on the Yps and the translocation breakpoint on Yps was distal to the pseudoautosomal region. At prenatal diagnosis, a normal appearing Y chromosome was found in his son, and thus the satellite on Yps was lost during meiotic Xp-Yp pairing. This Yps chromosome was likely the product of an "unstable" translocation.     

Abnormal XY interchange between a novel isolated protein kinase gene, PRKY, and its homologue, PRKX, accounts for one third of all (Y+)XX males and (Y-)XY females

XX males and XY females have a sex reversal disorder which can be caused by an abnormal interchange between the X and the Y chromosomes. We have isolated and characterized a novel gene on the Y chromosome, PRKY. This gene is highly homologous to a previously isolated gene from Xp22.3, PRKX, and represents a member of the cAMP-dependent serine threonine protein kinase gene family. Abnormal interchange can occur anywhere on Xp/Yp proximal to SRY. We can show that abnormal interchange happens particularly frequently between PRKX and PRKY. In a collection of 26 XX males and four XY females, between 27 and 35% of the interchanges take place between PRK homologues but at different sites within the gene. PRKY and PRKX are located far from the pseudoautosomal region where XY exchange normally takes place. The unprecedented high sequence identity and identical orientation of PRKY to its homologous partner on the X chromosome, PRKX, explains the high frequency of abnormal pairing and subsequent ectopic recombination, leading to XX males and XY females and to the highest rate of recombination outside the pseudoautosomal region.     

A sex reversal infant with XX karyotype and complete male external genitalia

The unusual case of a Japanese newborn XX male is presented. Examination of chromosomes in amniotic fluid cells had shown a normal female karyotype (46,XX), but ultrasonography revealed a penis and a scrotum. The neonate had normal male external genitalia, and serum levels of luteinizing hormone, follicle stimulating hormone, and testosterone were all within the normal range. High resonance chromosome analysis revealed an excess portion on the short arm of one of the X chromosomes. We examined his genomic DNA by polymerase chain reaction (PCR) and detected two Y specific regions in his genomic DNA, the sex-determining region Y (SRY) and pseudoautosomal boundary Y. Nucleotide sequencing of the PCR products of SRY indicated no mutation. These findings suggested that the translocation or insertion of an SRY region on the X chromosome led to the development of testicles and a male phenotype.     

Receptor-activated Ca2+ influx via human Trp3 stably expressed in human embryonic kidney (HEK)293 cells. Evidence for a non-capacitative Ca2+ entry

Gonadal differentiation involves a complex interplay of developmental pathways. The sex determining region Y (SRY) gene plays a key role in testis determination, but its interaction with other genes is less well understood. Abnormalities of gonadal differentiation result in a range of clinical problems. 46,XY complete gonadal dysgenesis is defined by an absence of testis determination. Subjects have female external genitalia and come to clinical attention because of delayed puberty. Individuals with 46,XY partial gonadal dysgenesis usually present in the newborn period for the valuation of ambiguous genitalia. Gonadal histology always shows an abnormality of seminiferous tubule formation. A diagnosis of 46,XY true hermaphroditism is made if the gonads contain well-formed testicular and ovarian elements. Despite the pivotal role of the SRY gene in testis development, mutations of SRY are unusual in subjects with a 46,XY karyotype and abnormal gonadal development. 46,XX maleness is defined by testis determination in an individual with a 46,XX karyotype. Most affected individuals have a phenotype similar to that of Klinefelter syndrome. In contrast, subjects with 46,XX true hermaphroditism usually present with ambiguous genitalia. The majority of subjects with 46,XX maleness have Y sequences including SRY in genomic DNA. However, only rare subjects with 46,XX true hermaphroditism have translocated sequences encoding SRY. Mosaicism and chimaerism involving the Y chromosome can also be associated with abnormal gonadal development. However, the vast majority of subjects with 45,X/46,XY mosaicism have normal testes and normal male external genitalia.     

Wilms' tumor suppressor gene (WT1) as a target gene of SRY function in a mouse ES cell line transfected with SRY

With the aim of identifying the gene(s) located downstream from SRY, we transfected an ES cell line with XX karyotype, TMA-18, with a Sry DNA construct and established cell lines, TS18-1 and TS18-2, where the transfected Sry was expressed in the functional linear mRNA form. Among the five potential SRY-target genes examined, i.e., MIS, SF1, P450arom, Sox9 and WT1, only the expression of WT1 was induced de novo by the unscheduled expression of Sry in the transfected cell lines. No clear indication of Sry-induced enhancement of Sox9 expression was obtained in the present series of experiments. Function of a yet unidentified gene(s) located on the Y chromosome might be needed for the up-regulation of Sox 9 expression which takes place during the development of male gonads. Quantitative RT-PCR analysis of the patterns of WT1 expression in developing fetal gonads revealed that although both male and female fetal gonads express WT1, male gonads invariably expressed WT1 mRNA at higher levels than female ones after the Sry expression. Immunohistochemical analysis of the male fetal gonads between 10.5 and 13.5 dpc demonstrated the presence of strong WT1 immunoreactivity in Sertoli cells of the primordial testes. Suggestions were made in the past indicating that both SF1 and WT1 proteins might be active in a common pathway upstream from Sry. Our results showed that WT1 is located downstream, rather than upstream from Sry and behaves independently from SF1. Analysis using an appropriate in vitro system will be essential to understand the molecular mechanisms of SRY action within cells.     

Prenatal identification of mos 45,X/46,X,+mar in a normal male baby by cytogenetic and molecular analysis

We report a case of mos 45,X/46,X,+mar, diagnosed prenatally by amniocentesis, whose physical examination, including external and internal organs, along with serum testosterone values were normal five years after delivery. The mosaic karyotype was seen in 146 of 240 cells examined (amniotic fluid cells, 110/65; placental chorionic villi: 5/4; cord blood, 21/81; cultured skin fibroblasts, 10/90) from 386 metaphases, and the marker chromosome appeared as a small non-fluorescent acrocentric chromosome. All autosomes appeared normal, and no normal Y chromosome could be demonstrated. Analysis of 26 Y-chromosome loci by molecular techniques such as PCR, Southern analysis using multiple Y-specific DNA probes, and Hae III restriction endonuclease assessment of male-specific repeated DNA in the heterochromatic region of the Y chromosome, and fluorescence in situ hybridization (FISH), revealed the marker was derived from a Y chromosome including p terminal to q11.23, and paracentric inversion in the remaining Y long arm. The formation of testes can be considered as existence of SRY (sex-determining region of Y) as a testis-determining factor. The present report illustrates the importance of FISH and molecular techniques as a complement to cytogenetic methods for accurate identification and characterization of chromosome rearrangements in prenatal diagnosis.     

Development in a 46 XX boy with positive SRY gene

We present the case of an 11 year-old boy, who asked for medical attention due to obesity and assumed underdeveloped external genitalia. He did not have genital anomalies, penile length was 5.3 cm, testicular volume 2 ml and pubic hair Tanner stage 1. His bone age was normal for chronological age. Endocrinological study showed normal results for his age. Karyotype revealed a 46 XX pattern. MRI of external genitalia showed bilateral scrotal testes which were normal in diameter for his age. The check of his historical growth chart and follow-up revealed normal growth with spontaneous pubertal development. However, hormonal studies showed progressive increase of FSH levels, indicative of failure of germinal epithelium. The presence of Y sequences, including SRY gene, was demonstrated by PCR. Our observation is in agreement with the view that 46 XX male subjects diagnosed at peripubertal age with the SRY gene in the genome have a good prognosis regarding growth and development, but the principal problem of these patients is infertility.     

Two SRY-negative XX male brothers without genital ambiguity

We report a Mexican family in which two sibs were identified as "classic" XX males without genital ambiguities. Molecular studies revealed that both patients were negative for several Y sequences, including SRY. A review of familial cases disclosed that this is the first family where a complete male phenotype was observed in Y-negative XX male non-twin brothers. These data suggest that an inherited loss-of-function mutation, in a gene participating in the sex-determining cascade, can induce normal male sexual differentiation in the absence of SRY.     

Hormonal, genetic and clinical findings in an XX male

A 17-year-old XX male with constitutional delay of growth and development and genetic short stature is described. Testosterone levels were normal but luteinizing-hormone-releasing-hormone-stimulated gonadotropin concentrations were increased. Testicular biopsy showed atrophic tubuli seminiferi and hyperplasia of the Leydig cells, and the spermiogram indicated azoospermia. Molecular analysis demonstrated the SRY gene close to the centromere of the paternally derived X chromosome. Clinical data in addition to the cytogenetic and molecular aspects are discussed.     

Preoperative and intraoperative radioimmunodetection of cancer pretargeted by biotinylated monoclonal antibodies

OBJECTIVES: Two cases of 46,XX true hermaphroditism were analyzed for two Y-DNA sequences, including the recently cloned gene for male testis determination, the sex-determining region of the Y chromosome (SRY). METHODS: Polymerase chain reaction was performed to amplify the SRY. DNA was prepared from peripheral blood lymphocytes as well as from gonadal tissue preserved in a paraffin block. RESULTS: One hermaphrodite contained the SRY sequences in peripheral blood lymphocytes and the testicular part of ovotestis tissue preserved in a paraffin block, while in the second patient these sequences were not detected. CONCLUSIONS: The SRY positive subject resulted from occult Y mosaicism rather than from X-Y translocation. Testis differentiation in the SRY negative subject may have been caused by mutation of a gene on the X chromosome or, alternatively, on an autosome.     

Dynamic three-dimensional echocardiographic reconstruction of congenital cardiac septation defects

DNA and FISH (fluorescence in situ hybridization) analysis were carried out in 12 patients with stigmata of Turner syndrome to determine whether the Supernumerary Marker Chromosome (SMC) found cytogenetically in each of these patients was derived from the Y chromosome. The presence of a Y chromosome in these patients may predispose them to develop gonadoblastoma. PCR-Southern blot analysis, followed by FISH, was used to detect the presence of Y chromosome material. The Sex determining Region Y (SRY), Testis Specific Protein Y-encoded (TSPY) and Y-chromosome RNA Recognition Motif (YRRM) genes, which map at Yp11.31, Yp11.1-11.2 and Yp11.2/Yq11.21-11.23, respectively, were selected as markers, because they span the whole Y chromosome, and more importantly, they are considered to be involved in the development of gonadoblastoma. It was shown that in 12 patients, all of whom had an SMC, the SMC of 11 was derived from the Y chromosome. Furthermore, the presence of the SRY, TSPY and YRRM gene sequences was determined and FISH analysis confirmed the Y origin of the SMCs. The methodology described in this report is a rapid, reliable and sensitive approach which may be easily applied to determine the Y origin of an SMC carried in Turner syndrome. The identification of an SMC is important for the clinical management and prognostic counseling of these patients with Turner syndrome.     

Prenatal in situ hybridization test for deleted steroid sulfatase gene

X-linked ichthyosis results from steroid sulfatase (STS) deficiency; 90% of affected patients have a complete deletion of the entire 146 kb STS gene on the distal X chromosome short arm (Xp22.3). In these families prenatal diagnosis and carrier testing can be completed in 2 days by hybridizing simultaneously 2 different cosmid probes labeled with fluorescein or Texas red and counterstaining interphase nuclear DNA with DAPI. An STS gene probe labeled with Texas red hybridizes specifically to the steroid sulfatase gene on the X chromosome. A second flanking probe labeled with fluorescein hybridizes to both the normal Y chromosome and normal and STS deleted X chromosomes. In this fashion the interphase nuclei of normal males, affected males, normal females, and carrier females can be distinguished unambiguously. Because normal males and carrier females each show two yellow-green fluorescein spots and one Texas red STS spot, use of this test prenatally requires determining fetal sex independently with repetitive X and Y chromosome-specific probes. This procedure can be used with lymphocytes, direct and cultured chorionic villus cells, direct and cultured amniocytes, and fibroblasts. Similar methods are anticipated to be useful for rapid diagnostic assessment of other aneuploid gene disorders.     

A gene involved in XY sex reversal is located on chromosome 9, distal to marker D9S1779

The genetic mechanisms involved in sex differentiation are poorly understood, and progress in identification of the genes involved has been slow. The fortuitous finding of chromosomal rearrangements in association with a sex-reversed phenotype has led to the isolation of SRY and SOX9, both shown to be involved in the sex-determining pathway. In addition, duplications of the X chromosome, deletions of chromosomes 9 and 10, and translocations involving chromosome 17 have been reported to be associated with abnormal testicular differentiation, leading to male-to-female sex reversal in 46,XY individuals. We present the cytogenetic and molecular analyses of four sex-reversed XY females, each with gonadal dysgenesis and other variable malformations, and with terminal deletions of distal chromosome 9p, resulting from unbalanced autosomal translocations. PCR amplification and DNA sequence analysis of SRY revealed no mutations in the high-mobility-group domain (i.e., HMG box) in any of the four patients. Conventional and molecular cytogenetic analyses of metaphase chromosomes from each patient suggest that the smallest region of overlap (SRO) of deletions involves a very small region of distal band 9p24. Loss-of-heterozygosity studies using 17 highly polymorphic microsatellite markers, as well as FISH using YAC clones corresponding to the most distal markers on 9p, showed that the SRO lies distal to marker D9S1779. These results significantly narrow the putative sex-determining gene to the very terminal region of the short arm of chromosome 9.     

Mammalian sex determination: from gonads to brain

In mammals, sex is determined by the Y chromosome, which encodes a testis-determining factor (TDF). This factor causes the undifferentiated embryonic gonads to develop as testes rather than ovaries. The testes subsequently produce the male sex hormones that are responsible for all male sexual characteristics. In 1990, the sex-determining gene, TDF, was identified and termed SRY in humans (Sry in mice). It encodes a protein containing a high mobility group (HMG) motif, which confers the ability to bind and to bend DNA. Genetic evidence supporting SRY as TDF came from the observation of a male phenotype in XX mice transgenic for a small genomic fragment containing Sry, and from the study of XY sex-reversed individuals who harbor de novo mutations in the SRY coding sequence. Other non-Y-linked genes involved in sex determination were subsequently found by genetic analysis of XY sex-reversed patients not explained by mutations in SRY. These genes are WT1, SF1, DAX1, and SOX9. A regulatory cascade hypothesis for mammalian sex determination, proposing that SRY represses a negative regulator of male development, was recently supported by observation of mice that expressed a DAX1 transgene and developed as XY sex-reversed females. The role of some sex-determining genes, such as DAX1 and SF1, in the development of the entire reproductive axis, a functionally integrated endocrine axis, leads to a new concept. Normal sexual development may result from the functional and developmental integration of a number of different genes that play roles in sex determination, sexual differentiation, and sexual behavior.     

Primary infertility in a phenotypic male with 46XX chromosomal constitution

The case of a 32 year old male with normal male adrenarchal hair pattern, bilateral gynaecomastia, a small phallus, hypospadias and bilateral poorly developed testes presenting with primary infertility secondary to azoospermia and a pelvic cyst is described. Repeated chromosomal analysis showed 46XX chromosomal constitution. Laparotomy revealed a simple cyst between the urinary bladder and the rectum. XX male syndrome is a rare cause of male infertility. The majority of cases is due to interchange of a fragment of the short arm of the Y chromosome containing the region that encodes the testes determining factor with the X chromosome. The presence of a simple cyst in the anatomical location of the uterus to our knowledge has not been reported in the literature.     

SRY and karyotypic status of one abnormal and two intersexual marsupials

An intersexual agile wallaby (Macropus agilis) with a penis, a pouch and four teats had a sex-chromosome constitution of XXY in lymphocytes and cultured fibroblasts; the sex-determining region Y (SRY) gene was present, consistent with the presence of a testis. An intersexual eastern grey kangaroo (Macropus giganteus) with a small empty scrotum and no penis, and an abnormal red kangaroo (Macropus rufus) with no penis, pouch or teats, both had XX sex-chromosome complements; the SRY gene was not present, consistent with testis absence. The agile wallaby and grey kangaroo described here provide further evidence that scrotal development in marsupials is independent of the Y chromosome. The cause of the abnormalities in the XX individuals cannot be determined until candidate genes are identified. These animals provide a basis for further genetic studies into marsupial intersexuality and sex differentiation.     

Ovarian Sertoli-Leydig cell tumor: a SRY gene-independent pathway of pseudomale gonadal differentiation

Sertoli-Leydig cell tumors (SLCT) are rare sex-cord stromal tumors of the ovary composed of undifferentiated gonadal stromal cells, Leydig cells (LC), and Sertoli cells (SC), with the latter forming structures resembling fetal testicular tubules. The histogenetic basis of morphological male differentiation patterns in females is controversial. Here, we report a SLCT with intermediate differentiation in a 23-year-old woman investigated by light microscopy, immunohistochemistry for intermediate filaments, and sex steroid hormone receptors (SSHR), as well as by polymerase chain reaction (PCR) for the presence of the sex-determining region Y gene (SRY). Our investigation shows that the SCs of SLCT express progesterone and androgen receptors as well as cytokeratins and vimentin. By PCR, SLCT-derived genomic DNA lacked the SRY gene, indicating that the SLCT results from a SRY gene-independent pathway of pseudomale gonadal differentiation. The expression of progesterone receptors (PRs) in the SCs of the SLCT is in contrast to their absence in testicular SCs, but in line with their presence in ovarian granulosa and surface epithelial cells. Thus, our results provide strong evidence for a close histogenetic relationship between the SLCT and the female gonocyte-supporting cell, the granulosa cell (GC).     

The effect of indolinic and quinolinic nitroxide radicals on trout erythrocytes exposed to oxidative stress

We have undertaken a clinical and molecular study of 25 females with deletions of the short arm of the X chromosome. We have determined the deletion breakpoints, the parental origin and the activation status of the deleted X chromosomes. Genotype-phenotype correlations suggest that the presence of a single copy of the DFFRX gene, previously postulated as a gene involved in the ovarian failure seen in Turner syndrome, may be compatible with normal ovarian function, and that there may be a gene for Turner-like features located in distal Xp22.3.     

Vectorette PCR isolation of microsatellite repeat sequences using anchored dinucleotide repeat primers

We have developed a vectorette PCR approach to provide an improved method for isolation of microsatellite repeats. The modified procedure relies on PCR amplification using a vectorette-specific primer in combination with one of a panel of anchored dinucleotide repeat primers. The target DNA to be screened for microsatellite sequences can be from YAC, P1, cosmid, bacteriophage or plasmid clones. We have used this technique to isolate novel, polymorphic microsatellite repeats from clones containing the amelogenin gene (AMGX) located on human chromosome Xp22.3.     

Therapeutic trials for promotion of faecal excretion of PCDFs by the administration of rice bran fiber and cholestyramine in Yusho patients

Two girls with Swyer syndrome (SS) were described. Diagnosis was established according to clinical data and ultrasound, laparoscope, histopathological, hormonal and cytogenetical examinations. One presents diagnostic possibilities followed advanced methods in genetics. The GTG and RBG high resolution bounding technique and replication analysis of short arms (Xp and Yp) were employed. Normal structure of end segments of X and Y chromosomes was mentioned. Molecular DNA analysis by polymerase chain reaction (PCR) did not find any mutation in SRY gene. Normal structure of this gene does not exclude possibility of SS existence. Our data implicates on the other mechanism of these disturbances.