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.     

Gonadal agenesis in a 46,XY female with multiple malformations and positive testing for the sex-determining region of the Y chromosome.

A full-term 46,XY female newborn presented with respiratory failure due to a right-sided diaphragmatic hernia. During surgical repair, exploration revealed isolated dextrocardia and hypoplasia of the right lung. Neither gonads nor wolffian or müllerian structures could be palpated. Cardiac catheterization demonstrated defects of the ventricular septum, hypoplasia of the right pulmonary artery, persistence of the left vena cava superior and a patent ductus arteriosus. Anthropometric data were normal at birth, but fell below the 3rd percentile during follow-up. Body proportions displayed a predominance of the upper compared to the lower segment. Endocrine studies indicated no defect of steroid biosynthesis and no functional gonadal tissue. Using genetic analyses of various loci within the testis-determining region of the Y chromosome, a mutation could not be detected. The patient died from pneumonia at the age of 19 months. Postmortem examination confirmed the diagnosis of gonadal agenesis.     

The sex-determining region Y (SRY) gene is mapped to p12-p13 of the Y chromosome in pig (Sus scrofa domestica) by in situ hybridization

The sex-determining region Y is a gene located in the distal portion of the short arm of human (SRY) and mouse (Sry) Y chromosomes and considered to be the best candidate for the testis determining factor (TDF/Tdy). The gene is believed to be the key factor in sex differentiation in mammals and is conserved across mammalian species. We report herein that the SRY/Sry gene has been assigned to pi 2-p13 on the short arm of the Y chromosome in pig by in situ hybridization. The result confirms interspecies conservation of this chromosomal segment in the evolution of mammalian chromosomes, and suggests further use of this gene probe in genomic studies in other mammals. The assignment of the Sry gene is the second physical gene mapping data available for the Y chromosome in pigs. Such data can be used in the effort of constructing the pig gene map and for further establishment of a comparison of sex chromosome morphology in different mammalian species concerning sex-specific and pseudoautosomal regions.     

Caenorhabditis elegans lin-13, a member of the LIN-35 Rb class of genes involved in vulval development, encodes a protein with zinc fingers and an LXCXE motif

The SynMuv genes appear to be involved in providing a signal that inhibits vulval precursor cells from adopting vulval fates in Caenorhabditis elegans. One group of SynMuv genes, termed class B, includes genes encoding proteins related to the tumor suppressor Rb and RbAp48, a protein that binds Rb. Here, we provide genetic evidence that lin-13 behaves as a class B SynMuv gene. We show that null alleles of lin-13 are temperature sensitive and maternally rescued, resulting in phenotypes ranging in severity from L2 arrest (when both maternal and zygotic activities are removed at 25 degrees ), to sterile Multivulva (when only zygotic activity is removed at 25 degrees ), to sterile non-Multivulva (when both maternal and zygotic activities are removed at 15 degrees ), to wild-type/class B SynMuv (when only zygotic activity is removed at 15 degrees ). We also show that LIN-13 is a nuclear protein that contains multiple zinc fingers and a motif, LXCXE, that has been implicated in Rb binding. These results together suggest a role for LIN-13 in Rb-mediated repression of vulval fates.     

Defective calmodulin-mediated nuclear transport of the sex-determining region of the Y chromosome (SRY) in XY sex reversal

The sex-determining region of the Y chromosome (SRY) plays a key role in human sex determination, as mutations in SRY can cause XY sex reversal. Although some SRY missense mutations affect DNA binding and bending activities, it is unclear how others contribute to disease. The high mobility group domain of SRY has two nuclear localization signals (NLS). Sex-reversing mutations in the NLSs affect nuclear import in some patients, associated with defective importin-beta binding to the C-terminal NLS (c-NLS), whereas in others, importin-beta recognition is normal, suggesting the existence of an importin-beta-independent nuclear import pathway. The SRY N-terminal NLS (n-NLS) binds calmodulin (CaM) in vitro, and here we show that this protein interaction is reduced in vivo by calmidazolium, a CaM antagonist. In calmidazolium-treated cells, the dramatic reduction in nuclear entry of SRY and an SRY-c-NLS mutant was not observed for two SRY-n-NLS mutants. Fluorescence spectroscopy studies reveal an unusual conformation of SRY.CaM complexes formed by the two n-NLS mutants. Thus, CaM may be involved directly in SRY nuclear import during gonadal development, and disruption of SRY.CaM recognition could underlie XY sex reversal. Given that the CaM-binding region of SRY is well-conserved among high mobility group box proteins, CaM-dependent nuclear import may underlie additional disease states.     

The role of the sex-determining region of the Y chromosome (SRY) in the etiology of 46,XX true hermaphroditism

Summary The syndrome of 46,XX true hermaphroditism is a clinical condition in which both ovarian and testicular tissue are found in one individual. Both Mullerian and Wolffian structures are usually present, and external genitalia are often ambiguous. Two alternative mechanisms have been proposed to explain the development of testicular tissue in these subjects: (1) translocation of chromosomal material encoding the testicular determination factor (TDF) from the Y to the X chromosome or to an autosome, or (2) an autosomal dominant mutation that permits testicular determination in the absence of TDF. We have investigated five subjects with 46,XX true hermaphroditism. Four individuals had a normal 46,XX karyotype; one subject (307) had an apparent terminal deletion of the short arm of one X chromosome. Genomic DNA was isolated from these individuals and subjected to Southern blot analysis. Only subject 307 had Y chromosomal sequences that included the pseudoautosomal boundary, SRY (sex-determining region of Y), ZFY (Y gene encoding a zinc finger protein), and DXYS5 (an anonymous locus on the distal short arm of Y) but lacked sequences for DYZ5 (proximal short arm of Y) and for the long arm probes DYZ1 and DYZ2. The genomic DNA of the other four subjects lacked detectable Y chromosomal sequences when assayed either by Southern blotting or after polymerase chain reaction amplification. Our data demonstrate that 46,XX true hermaphroditism is a genetically heterogeneous condition, some subjects having TDF sequences but most not. The 46,XX subjects without SRY may have a mutation of an autosomal gene that permits testicular determination in the absence of TDF.     

A mouse brain cDNA encodes a novel protein with the protein kinase C phosphorylation site domain common to MARCKS

We have isolated a mouse brain cDNA clone encoding a protein of 200 amino acids (Mr 20,165) with partial homology with MARCKS (myristoylated alanine-rich C-kinase substrate). Two regions show similarity with MARCKS, one is the kinase C phosphorylation site domain which is supposed to bind calmodulin, and the other is the region near to the N-terminus, including the consensus sequence of myristoylation. It has a similar amino acid composition to MARCKS, but the content of alanine is not as high. It is distributed throughout the mouse brain, but the pattern is not identical with that of MARCKS. Both proteins may be members of a new protein family involved in coupling the protein kinase C and calmodulin signal transduction systems.     

The X chromosome of marsupials shares a highly conserved region with eutherians

Ten genes, located on the long arm of the human X chromosome, were mapped in several marsupial species by somatic cell analysis and in situ hybridization. All were located on the X chromosome in each species. We conclude that the long arm of the human X chromosome represents a highly conserved region that formed part of the X chromosome in a therian ancestor 120-150 million years ago, before the mammalian infraclasses diverged.     

A structural analysis of the Sxr region of the mouse Y chromosome.

Three genetic functions have been mapped to the minute Sxr (sex-reversed) region of the mouse Y chromosome. These are Tdy, the primary testis determinant; Hya, the locus (either structural or regulatory) controlling the expression of the male-specific minor histocompatibility antigen H-Y; and Spy, a spermatogenic gene. Hya and Spy map to DNA deleted from the Sxr region in the deletion variant Sxrb (the delta Sxrb DNA). With the object of cloning Hya and Spy, we initiated chromosome walking in the delta Sxrb DNA. From three independent loci--Sx1, Zf2, and T5--we have isolated approximately 270 kb of delta Sxrb DNA lying in three contigs of 145, 60, and 65 kb, respectively. Within 17 kb of the 3' end of the Zfy-2 gene, lowcopy repeat elements were found in a region that extends for approximately 35 kb. Probes isolated from this region detect multiple Sxr loci, some of which map to the delta Sxrb DNA present in the T5 contig DNA. Three of these multicopy probes detect delta Sxrb loci not represented in our three contigs, which means that six distinct delta Sxrb loci have now been identified. Here we present a preliminary model of the molecular structure of the DNA in this unique region.     

The mouse Y* chromosome involves a complex rearrangement, including interstitial positioning of the pseudoautosomal region.

Cytological analysis of the mouse Y* chromosome revealed a complex rearrangement involving acquisition of a functional centromere and centromeric heterochromatin and attachment of this chromosomal segment to the distal end of a normal Y* chromosome. This rearrangement positioned the Y* short-arm region at the distal end of the Y* chromosome and the pseudoautosomal region interstitially, just distal to the newly acquired centromere. In addition, the majority of the pseudoautosomal region was inverted. Recombination between the X and the Y* chromosomes generates two new sex chromosomes: (1) a large chromosome comprised of the X chromosome attached at its distal end to all of the Y* chromosome but missing the centromeric region (XY*) and (2) a small chromosome containing the centromeric portion of the Y* chromosome attached to G-band-negative material from the X chromosome (YX). Mice that inherit the XY* chromosome develop as sterile males, whereas mice that inherit the Y*X chromosome develop as fertile females. Recovery of equal numbers of recombinant and nonrecombinant offspring from XY* males supports the hypothesis that recombination between the mammalian X and Y chromosomes is necessary for primary spermatocytes to successfully complete spermatogenesis and form functional sperm.