Sulfotransferase (ST) enzymes catalyze the sulfate conjugation of many hormones, neurotransmitters, drugs, and xenobiotic compounds. These reactions result in enhanced renal excretion of the sulfate-conjugated reaction products, but they can also lead to the formation of "bioactivated" metabolites. ST enzymes are members of an emerging gene superfamily that presently includes phenol ST (PST), hydroxysteroid ST (HSST), and, in plants, flavonol ST (FST) "families," members of which share at least 45% amino acid sequence identity. These families can be further subdivided into "subfamilies" that are at least 60% identical in amino acid sequence. For example, the PST family includes both PST and estrogen ST (EST) subfamilies. Amino acid sequence motifs exist within ST enzymes that are conserved throughout phylogeny. These signature sequences may be involved in the binding of 3'-phosphoadenosine-5 '-phosphosulfate, the cosubstrate for the sulfonation reaction. There are presently five known human cytosolic ST enzymes: an EST, an HSST, and three PSTs. cDNAs and genes for all of these enzymes have been cloned, and chromosomal localizations have been reported for all five genes. Genes for these human enzymes, as well as those of other mammalian cytosolic ST enzymes that have been cloned, show a high degree of structural homology, with conservation of the locations of most intron/exon splice junctions. Human ST enzyme expression varies among individuals. Functionally significant genetic polymorphisms for ST enzymes in humans have been reported, and other molecular genetic mechanisms that might be involved in the regulation of the expression of these enzymes are being explored. Knowledge of the molecular biology of cytosolic ST enzymes, when placed within a context provided by decades of biochemical research, promises to significantly enhance our understanding of the regulation of the sulfate conjugation of hormones, neurotransmitters, and drugs.
Thiopurine methyltransferase (TPMT) catalyzes the S-methylation of thiopurine drugs. Individual variation in the toxicity and therapeutic efficacy of these drugs is associated with a common genetic polymorphism that controls levels of TPMT activity and immunoreactive protein in human tissues. Because of the clinical significance of the "pharmacogenetic" regulation of this enzyme, it would be important to clone the gene for TPMT in humans and to study the molecular basis for the genetic polymorphism. As a first step toward cloning the gene for TPMT, we used the rapid amplification of genomic DNA ends to obtain a TPMT-specific intron sequence. That DNA sequence was used to design primers for the polymerase chain reaction (PCR), which made it possible to determine that the active gene for TPMT is located on human chromosome 6. A TPMT-positive cosmid clone was then isolated from a human chromosome 6-specific genomic DNA library, and the gene was sublocalized to chromosome band 6p22.3 by fluorescence in situ hybridization. The gene for TPMT was found to be approximately 34 kb in length and consisted of 10 exons and 9 introns. On the basis of the results of 5'-rapid amplification of cDNA ends, transcription initiation occurred at or near a point 89 nucleotides upstream from the translation initiation codon of previously reported TPMT cDNAs. Once the structure of the TPMT gene had been determined, it was possible to perform the PCR with primers complementary to the sequences of introns flanking each exon that encodes enzyme protein with template DNA obtained from subjects with known phenotypes for the TPMT genetic polymorphism. This DNA was isolated from blood samples from 4 unrelated subjects with genetically low TPMT activity and 4 unrelated subjects with high TPMT activity. All subjects with low TPMT activity were homozygous for two point mutations--a G-->A transition at nucleotide 460 in exon 7 and an A-->G transition at nucleotide 719 in exon 10. Both mutations resulted in alterations in amino acid sequence, with Ala-154-->Thr and Tyr-240-->Cys, respectively. All DNA samples isolated from the blood of subjects with high TPMT activity contained "wild-type" sequence. Results obtained with these blood samples were confirmed when DNA from four human liver samples with high TPMT activity were found to have wild-type sequence at nucleotides 460 and 719, while three liver samples with intermediate enzyme activity (i.e., samples presumed to be heterozygous for the polymorphism) were heterozygous for the exon 7 and exon 10 mutations present in the blood samples of homozygous low subjects. Transient expression in COS-1 cells of TPMT expression constructs that contained both of the mutations in exons 7 and 10, as well as each independently, demonstrated that each mutation, as well as both together, resulted in decreased expression of TPMT enzymatic activity and immunoreactive protein. Molecular cloning and structural characterization of the TPMT gene as well as elucidation of the molecular basis for a common TPMT genetic poly...
Meiotic silencing of sex chromosomes may cause their depletion of meiosis-specific genes during evolution. Here, we challenge this hypothesis by reporting the identification of TEX11 as the first X-encoded meiosis-specific factor in mice. TEX11 forms discrete foci on synapsed regions of meiotic chromosomes and appears to be a novel constituent of meiotic nodules involved in recombination. Loss of TEX11 function causes chromosomal asynapsis and reduced crossover formation, leading to elimination of spermatocytes, respectively, at the pachytene and anaphase I stages. Specifically, TEX11-deficient spermatocytes with asynapsed autosomes undergo apoptosis at the pachytene stage, while those with only asynapsed sex chromosomes progress. However, cells that survive the pachytene stage display chromosome nondisjunction at the first meiotic division, resulting in cell death and male infertility. TEX11 interacts with SYCP2, which is an integral component of the synaptonemal complex lateral elements. Thus, TEX11 promotes initiation and/or maintenance of synapsis and formation of crossovers, and may provide a physical link between these two meiotic processes.[Keywords: TEX11; male infertility; meiosis; synapsis; meiotic recombination; X chromosome] Supplemental material is available at http://www.genesdev.org.
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