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PubMed2004

A mutation in a functional Sp1 binding site of the telomerase RNA gene (hTERC) promoter in a patient with Paroxysmal Nocturnal Haemoglobinuria.

BACKGROUND: Mutations in the gene coding for the RNA component of telomerase, hTERC, have been found in autosomal dominant dyskeratosis congenita (DC) and aplastic anemia. Paroxysmal nocturnal hemoglobinuria (PNH) is a clonal blood disorder associated with aplastic anemia and characterized by the presence of one or more clones of blood cells lacking glycosylphosphatidylinositol (GPI) anchored proteins due to a somatic mutation in the PIGA gene. METHODS: We searched for mutations in DNA extracted from PNH patients by amplification of the hTERC gene and denaturing high performance liquid chromatography (dHPLC). After a mutation was found in a potential transcription factor binding site in one patient electrophoretic mobility shift assays were used to detect binding of transcription factors to that site. The effect of the mutation on the function of the promoter was tested by transient transfection constructs in which the promoter is used to drive a reporter gene. RESULTS: Here we report the finding of a novel promoter mutation (-99C->G) in the hTERC gene in a patient with PNH. The mutation disrupts an Sp1 binding site and destroys its ability to bind Sp1. Transient transfection assays show that mutations in this hTERC site including C-99G cause either up- or down-regulation of promoter activity and suggest that the site regulates core promoter activity in a context dependent manner in cancer cells. CONCLUSIONS: These data are the first report of an hTERC promoter mutation from a patient sample which can modulate core promoter activity in vitro, raising the possibility that the mutation may affect the transcription of the gene in hematopoietic stem cells in vivo, and that dysregulation of telomerase may play a role in the development of bone marrow failure and the evolution of PNH clones.

Journal Article↗

PubMed1988

Mutation of Ki-ras and N-ras oncogenes in myelodysplastic syndromes.

Somatic mutation of the N-ras oncogene occurs frequently in de novo acute myeloid leukemia (AML). By virtue of their relation to AML, myelodysplastic syndromes (MDS) provide an in vivo model of human leukemogenesis. By using a strategy for analysis of gene mutation based on in vitro amplification of target sequences by the polymerase chain reaction (PCR) and selective oligonucleotide hybridization we analyzed the mutational status of codons 12, 13, and 61 of Ha-ras, K-ras, and N-ras in peripheral blood (PB) and/or bone marrow (BM) in 34 cases of primary MDS. Mutations at codon 12 of Ki-ras or N-ras were detected in three cases (9%): one of six cases of refractory anemia with excess blasts (RAEB) and two of nine cases of chronic myelomonocytic leukemia (CMML). The nucleotide substitution differed in each. In all cases the mutant allele was detectable in PB cells. A sustained hematologic remission was achieved after low-dose cytarabine therapy in the case of RAEB. Neither case of CMML exhibited signs of disease progression during follow-up at 7 and 12 months. In contrast, four of 31 patients without the ras mutation underwent transformation to AML within 12 months of genetic analysis. We conclude that ras mutations in MDS are heterogeneous and may develop at an early stage during the evolution of MDS. Their detection in PB cells illustrates the potential utility of ras mutation as a clonal marker in myeloid malignancy.

Genes, ras↗

PubMed2004

The phylogenetic origins of the antigen-binding receptors and somatic diversification mechanisms.

The adaptive immune system arose in ancestors of the jawed vertebrates approximately 500 million years ago. Homologs of immunoglobulins (Igs), T-cell antigen receptors (TCRs), major histocompatibility complex I (MHC I) and MHC II, and the recombination-activating genes (RAGs) have been identified in all extant classes of jawed vertebrates; however, no definitive homolog of any of these genes has been identified in jawless vertebrates or invertebrates. RAG-mediated recombination and associated junctional diversification of both Ig and TCR genes occurs in all jawed vertebrates. In the case of Igs, somatic variation is expanded further through class switching, gene conversion, and somatic hypermutation. Although the identity of the 'primordial' receptor that was interrupted by the recombination mechanism in jawed vertebrates may never be established, many different families of genes that exhibit predicted characteristics of such a receptor have been described both within and outside the jawed vertebrates. Recent data from various model systems point toward a continuum of immune receptor diversity, encompassing many different families of recognition molecules whose functions are integrated in an organism's response to pathogenic invasion. Various approaches, including both genomic and protein-functional analyses, currently are being applied in jawless vertebrates, protochordates, and other invertebrate deuterostome systems and may yield definitive evidence regarding the presence or absence of adaptive immune homologs in species lacking adaptive immune systems. Such studies have the potential for uncovering previously unknown mechanisms of generating receptor diversity.

Animals↗

PubMed1996

Localization of the homolog of a mouse craniofacial mutant to human chromosome 18q11 and evaluation of linkage to human CLP and CPO.

The transgene-induced mutation 9257 and the spontaneous mutation twirler cause craniofacial and inner ear malformations and are located on mouse chromosome 18 near the ataxia locus ax. To map the human homolog of 9257, a probe from the transgene insertion site was used to screen a human genomic library. Analysis of a cross-hybridizing human clone identified a 3-kb conserved sequence block that does not appear to contain protein coding sequence. Analysis of somatic cell hybrid panels assigned the human locus to 18q11. The polymorphic microsatellite markers D18S1001 and D18S1002 were isolated from the human locus and mapped by linkage analysis using the CEPH pedigrees. The 9257 locus maps close to the centromeres of human chromosome 18q and mouse chromosome 18 at the proximal end of a conserved linkage group. To evaluate the role of this locus in human craniofacial disorders, linkage to D18S1002 was tested in 11 families with autosomal dominant nonsyndromic cleft lip and palate and 3 families with autosomal dominant cleft palate only. Obligatory recombinants were observed in 8 of the families, and negative lod scores from the other families indicated that these disorders are not linked to the chromosome 18 loci.

Alleles↗

PubMed1995

Somatic diversification and selection of immunoglobulin heavy and light chain variable region genes in IgG+ CD5+ chronic lymphocytic leukemia B cells.

Chronic lymphocytic leukemia (CLL) is characterized by the clonal expansion of CD5-expressing B lymphocytes. Most studies have found that these leukemic CD5+ B cells, like their normal counterparts, use immunoglobulin (Ig) variable (V) region genes that exhibit minimal, if any, somatic diversity. These and other observations have suggested that CD5+ B cells may be incapable of generating Ig V gene diversity, and therefore may not be able to develop higher affinity binding sites that could be selected by antigen. However, most of the studies of CLL and normal CD5+ B cells have focused on IgM-producing cells. Since somatic mutations are most often seen in B cells that have undergone an isotype class switch, we analyzed the Ig heavy (H) and light (L) chain variable region genes of seven IgG+CD5+ CLL B cells to determine if somatic diversification and antigen selection had occurred. The data derived provide evidence for skewed use, somatic diversification, and antigenic selection of the Ig V region genes. Nonrandom use of both H and L chain V region genes was manifested by an overrepresentation of VH4 and VKI family genes and the underrepresentation of the JH4 gene segment. Furthermore, VH4 gene use was restricted to only two family members (4.21 and 4.18). In four of the seven cases, the VH and VL genes displayed > or = 5% difference from the most homologous known germline counterparts. Polymerase chain reaction and Southern blot analyses performed in two of these patients demonstrated that their unique VH CDR2 and adjacent sequences were not present in their germline DNA. In addition, a significant level of diversity was seen in the rearranged DJH segments and at the VL-JL junctions of every patient that occurred both at the time of recombination and subsequently. The localization of replacement changes to complementarity determining regions of some patients suggested that antigen selection had occurred. Furthermore, the mutations identified in the VH and VL genes of each individual patient were strikingly similar, both in number and location. Collectively, the data indicate that a subset of CD5+ CLL B cells can display Ig V region gene mutations. In addition, they are consistent with the notions that in some cases antigen selection of these mutations may have occurred, and that antigen stimulation may be a promoting factor in the evolution of certain CLL clones.

Amino Acid Sequence↗

PubMed1994

Tactile discrimination capacity in relation to size and organization of somatic sensory cortex in primates: I. Old-World prosimian, Galago; II. New-World anthropoids, Saimiri and Cebus.

Living primates vary considerably in brain organization, in sensorimotor and cognitive abilities, and in natural behavioral repertoires. Comparative studies of primary (SI) and secondary (SII) somatic sensory cortex of primates reveal major differences in the size and in the complexity of topographic projection patterns. The separate projections of the glabrous hand to SI cytoarchitectonic areas 3b and 1 described in the Old World (OW) anthropoid Macaca and in New World (NW) anthropoids Cebus, Saimiri, and Aotus are lacking in NW Saguinus and in the prosimian Galago. The relationship between the size and complexity of SI organization and tactile abilities is explored in this study of four species of primates--Galago, Macaca, Cebus, and Saimiri. These species were trained to discriminate between pairs of objects differing either in cross-sectional diameter (size) or surface roughness (texture). The course of acquisition of such tactile discrimination in normal Macaca and the nature of deficits following SI or SII removals are known. Selective lesions of either cytoarchitectonic area 1 or 2 in Macaca affect only texture or size discriminations, respectively. Removal of area 3b in SI, or of SII, in Macaca affects both size and texture capacities. The single projection of the glabrous hand to area 3b-1 of Galago led to our expectation that the capacity of Galago to discriminate textures would be more similar to an area 1-lesioned than to a normal Macaca. The substantial and persistent differences between Macaca and Galago on texture, but not size, tasks lend support to the view that the evolution of a second projection of the glabrous hand to area 1 in Macaca contributes to increased texture discrimination capacity. The similarity in multiple projection patterns of the glabrous hand to areas 3b and 1 in Macaca, Saimiri, and Cebus led us to expect greater correspondence in texture discrimination capacity between these three anthropoids than to Galago. Contrary to expectations, Saimiri and Cebus showed a tactile capacity more similar to Galago than to Macaca. Furthermore, the texture discrimination capacity of Cebus actually improved substantially after removal of area 1. This provides further evidence, together with the single SI hand area in NW Saguinus and Galago, that the separate cutaneous projections to area 1 in OW and NW primates are not homologous but evolved independently and possibly serve different tactile functions.

Animals↗

PubMed1989

A fine-structure deletion map of human chromosome 11p: analysis of J1 series hybrids.

Deletion analysis offers a powerful alternative to linkage and karyotypic approaches for human chromosome mapping. A panel of deletion hybrids has been derived by mutagenizing J1, a hamster cell line that stably retains chromosome 11 as its only human DNA, and selecting for loss of MIC1, a surface antigen encoded by a gene in band 11p13. A unique, self-consistent map was constructed by analyzing the pattern of marker segregation in 22 derivative cells lines; these carry overlapping deletions of 11p13, but selectively retain a segment near the 11p telomere. The map orders 35 breakpoints and 36 genetic markers, including 3 antigens, 2 isozymes, 12 cloned genes, and 19 anonymous DNA probes. The deletions span the entire short arm, dividing it into more than 20 segments and define a set of reagents that can be used to rapidly locate any newly identified marker on 11p, with greatest resolution in the region surrounding MIC1. The approach we demonstrate can be applied to map any mammalian chromosome. To test the gene order, we examined somatic cell hybrids from five patients, whose reciprocal translocations bisect band 11p13; these include two translocations associated with familial aniridia and two with acute T-cell leukemia. In each patient, the markers segregate in telomeric and centromeric groups as predicted by the deletion map. These data locate the aniridia gene (AN2) and a recurrent T-cell leukemia breakpoint (TCL2) in the marker sequence, on opposite sides of MIC1. To provide additional support, we have characterized the dosage of DNA markers in a patient with Beckwith-Wiedemann syndrome and an 11p15-11pter duplication. Our findings suggest the following gene order: TEL - (HRAS1, MER2, CTSD, TH/INS/IGF2, H19, D11S32) - (RRM1, D11S1, D11S25, D11S26) - D11S12 - (HBBC, D11S30) - D11S20 - (PTH, CALC) - (LDHA, SAA, TRPH, D11S18, D11S21) - D11S31 - D11S17 - HBVS1 - (FSHB, D11S16) - AN2 - MIC1 - TCL2 - delta J - CAT - MIC4 - D11S9 - D11S14 - ACP2 - (D11S33, 14L) - CEN. We have used the deletion map to show the distribution on 11p of two centromeric repetitive elements and the low-order interspersed repeat A36Fc. Finally, we provide evidence for an allelic segregation event in the hamster genome that underlies the stability of chromosome 11 in J1. The deletion map provides a basis to position hereditary disease loci on 11p, to distinguish the pattern of recessive mutations in different forms of cancer and, since many of these genes have been mapped in other mammalian species, to study the evolution of a conserved syntenic group.

Animals↗

PubMed2026

A duplicated female pathway gene figla-like evolves as the male sex-determining gene in tilapia.

As the largest group of vertebrates, fish exhibit frequent turnover of sex-determining (SD) genes. Here, we assemble a chromosome-level YY red tilapia genome and identify figla-like (figlal) as the SD gene on tilapia linkage group (LG) 1. Integrative phylogenetic and genomic evidence suggests that figlal originated from a tilapia-specific duplication and transposition of the ancestral bHLH family gene figla from LG12 to LG1. Fluorescence in situ hybridization reveals expression divergence between figla and figlal, with figla expressed in female oocytes and figlal expressed in male gonadal somatic cells during early gonadal differentiation. The shift in expression after duplication might be driven by the insertion of cis-regulatory elements mediated by transposable elements. Knockout of figlal in XY fish results in male-to-female sex reversal as indicated by ovarian morphology, down-regulation of the male pathway gene dmrt1, and up-regulation of the female pathway gene cyp19a1a in the gonads. In contrast, overexpression of figlal in XX fish induces female-to-male sex reversal. These findings implicate figlal as an SD gene on tilapia LG1 and reveal the history of a unique evolutionary innovation in which a female oocyte gene evolved into a male SD gene via duplication, transposition, and cis-regulatory rewiring.

Animals↗

PubMed2005

New mammalian cellular systems to study mutations introduced at the break site by non-homologous end-joining.

The non-homologous end-joining (NHEJ) pathway is a mechanism to repair DNA double strand breaks, which can introduce mutations at repair sites. We constructed new cellular systems to specifically analyze sequence modifications occurring at the repair site. In particular, we looked for the presence of telomeric repeats at the repair junctions, since our previous work indicated that telomeric sequences could be inserted at break sites in germ-line cells during primate evolution. To induce specific DNA breaks, we used the I-SceI system of Saccharomyces cerevisiae or digestion with restriction enzymes. We isolated human and hamster cell lines containing the I-SceI target site integrated in a single chromosomal locus and we exposed the cells to a continuous expression of the I-SceI endonuclease gene. Additionally, we isolated human cell lines that expressed constitutively the I-SceI endonuclease and we introduced the target site on an episomal plasmid stably transfected into the cells. These strategies allowed us to recover repair junctions in which the I-SceI target site was modified at high frequency (100% in hamster cells and about 70% in human cells). Finally, we analyzed junctions produced on an episomal plasmid linearized by restriction enzymes. In all the systems studied, sequence analysis of individual repair junctions showed that deletions were the most frequent modifications, being present in more than 80% of the junctions. On the episomal plasmids, the average deletion length was greater than at intrachromosomal sites. Insertions of nucleotides or deletions associated with insertions were rare events. Junction organization suggested different mechanisms of formation. To check for the insertion of telomeric sequences, we screened plasmid libraries representing about 3.5 x 10(5) junctions with a telomeric repeat probe. No positive clones were detected, suggesting that the addition of telomeric sequences during double strand break repair in somatic cells in culture is either a very rare event or does not occur at all.

Animals↗

PubMedDate not supplied

An update on the oxygen stress-mitochondrial mutation theory of aging: genetic and evolutionary implications.

The acceleration of fixed-postmitotic cell aging by a high metabolic rate and the age related loss of mitochondria found in that cell type led us to propose an oxygen stress-mitochondrial mutation theory of aging, according to which senescence may be linked to mutations of the mitochondrial genome (mtDNA) of the irreversibly differentiated cells. This extranuclear somatic gene mutation concept of aging is supported by the fact that mtDNA synthesis takes place at the inner mitochondrial membrane near the sites of formation of highly reactive oxygen species. Mitochondrial DNA may be unable to prevent the intrinsic mutagenesis caused by those byproducts of respiration because, in contrast to the nuclear genome, it lacks excision and recombination repair. The resulting mitochondrial impairment and concomitant cell bioenergetic decline may cause the senescent loss of physiological performance and may play a key role in the pathogenesis of many age-related degenerative diseases. These concepts are integrated with classic and contemporary hypotheses in a unitary theory that reconciles programmed and stochastic concepts of aging. Thus, it is suggested that cells are programmed to differentiate, and then they accumulate mitochondrial-genetic damage because of their high levels of oxyradical stress and the loss of the organelle rejuvenating power of mitosis.

Aging↗

PubMed1997

Imprinting is also a mechanism for immediate or delayed hemizygous expression of several uniparental haplotypes selected from the genome of each sex.

A peculiar and interesting aspect of monoallelic or hemizygous expression, resulting from genomic imprinting, should be a likeness or resemblance for some phenotypic traits between relatives inheriting identical active genes or domains. Although the word "likon," a neologism, is reminiscent of the above implication, it is here proposed for use in a broader sense, namely, to designate a haplotype or part of a haplotype of an imprinted domain. As learned from earlier studies of imprinting and uniparental disomies, haplotypes at loci of such domains may be expressed (E) or unexpressed (U) in somatic cells; they may also be transmitted to be expressed or not in the next generation by germ cells "acting" (A) or marked to be "resting" (R) for such loci. Thus the soma/germinal status of "likons" might for each genitor be abbreviated as EA, UA, ER, and UR. In an evolutionary sense the assumption is that the same monoallelically expressed loci and domains when carried by two or more relatives should be the source of identical transcripts contributing to a closely similar phenotype. If so, the overall phenotype would be distinct if arising from some 10 to 20 imprinted genes or domains potentially gaining expression from the germ cells of either one or the other sex in humans. The result may have evolutionary implications by narrowing the scope of random individual variation and by strengthening assortative and associative values (physical, behavioral, and instinctual) in one's own lineage and species.

Chromosome Aberrations↗

PubMed1989

The IL-4 and IL-5 genes are closely linked and are part of a cytokine gene cluster on mouse chromosome 11.

The murine IL-4 and IL-5 genes encode hemopoietic growth factors involved in the stimulation, proliferation, and differentiation of cells of the T lymphocyte, B lymphocyte, and granulocyte lineages. We have mapped the Il-4 and Il-5 loci representing the structural genes for IL-4 and IL-5, respectively, to mouse chromosome 11 using Chinese hamster x mouse and rat x mouse somatic cell hybrids. Physical linkage studies of the IL-4 and IL-5 genes by pulsed field gel electrophoresis have shown that they are closely linked, being 110-180 kb apart. Since the Il-5 locus maps to the interface of bands A5 and B1 in the same location as the genes for IL-3 and GM-CSF, this places these three cytokine genes, as well as the IL-4 gene, within a region of about 5000-10,000 kb. The present physical linkage studies indicate that the IL-4 and IL-5 genes are a minimum of 600 kb apart from the closely linked IL-3 and GM-CSF genes. The gene clustering, together with similarities in gene structure, regulation, and biological function, raises the possibility that the four genes may be part of a distantly related cytokine gene family.

Animals↗

PubMed1985

On the diversity of sperm histones in the vertebrates: IV. Cytochemical and amino acid analysis in Anura.

The variability of sperm histones in frogs has been studied by cytochemical and amino acid analyses. Cytochemically, Rana sperm proteins fall into Bloch's ('69, '76) type 4 somatic-like histone category, while Xenopus and Bufo have type 3 intermediate sperm histones. Extractability in 5% trichloroacetic acid (TCA) at different temperatures splits this type 3 category into two groups: type 3B intermediate sperm histones of Bufo are extractable at 85-90 degrees C, while Xenopus intermediate type 3A sperm histones require temperatures of 95-100 degrees C for extraction. Amino acid analysis confirms that Rana sperm histones are of the nucleosomal type, with a testis-specific, very lysine-rich H1 histone. The sperm protein in Bufo is richer in arginine than the proteins in Xenopus. Both of these genera contain lysine and histidine as well as arginine in their sperm proteins. These results confirm earlier electrophoretic data (Kasinsky et al., '78) and indicate that sperm histones in the order Anura can vary markedly between different genera.

Amino Acids↗

PubMed1991

Follicular lymphoma: a model of lymphoid tumor progression in man.

Human follicular lymphoma can be viewed as a malignancy in evolution. Since this disease is composed of a clonal population of B lymphocytes all expressing a given immunoglobulin light chain and heavy chain, it seems possible that the initial transforming event, the t(14; 18) chromosomal translocation, occurs in a cell already committed to the expression of a particular VH and VL gene. A panel of antibodies has been assembled which define a set of idiotypes expressed repeatedly by B-cell lymphomas. Nonetheless, VH gene usage in follicular lymphoma tumors appears to reflect the normal B-cell repertoire. Growth of follicular lymphoma appears to be partially under normal regulatory control. The expanding malignant B-cell clone grows in follicles with particular apposition to follicular dendritic cells and heavy infiltration with CD4+ T cells. Interaction with T cells can induce the proliferation of follicular lymphoma cells. This tumor eventually evolves into a diffuse large-cell lymphoma which is highly aggressive and lethal. It is now clear that the malignant progression occurs from a single cell within the expanding follicular lymphoma clone. A panel of monoclonal antibodies to cell surface molecules has been generated that inhibit proliferation of diffuse lymphoma cell lines, and some of the target molecules have been partially characterized. Therapeutic application of anti-idiotype monoclonal antibodies has shown a high degree of tumor responsiveness, but ultimately escape of idiotype-negative variant cells occurs. These variants arise as a result of extensive somatic point mutation in the VH and VL genes of follicular lymphoma. Active immunization can result in an immune response by patients directed against the idiotype expressed on their own B-cell tumors.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Communication↗

PubMed1995

Understanding the biology of aging: the key to prevention and therapy.

OBJECTIVE: To review recent progress and consider future approaches for basic research on aging with clinical applicability. DATA SOURCES: Peer-reviewed publications on experimental gerontology and geriatrics. STUDY SELECTION AND DATA EXTRACTION: Studies were selected that described experimental approaches in gerontology and geriatrics, starting with the evolutionary basis of aging, through theories trying to explain its major causes, to novel experimental approaches, e.g., computer informatics, protein chemistry and genetics. DATA SYNTHESIS: Our increased understanding of the evolutionary basis of aging has made it possible to consider a number of experimental strategies more rationally. Most theories on the causes of aging involve some kind of somatic damage that accumulates with age, the rate of which is determined by environmental, genetic, and behavioral factors. The recent emergence of more powerful methodology offers new possibilities for identifying basic mechanisms of aging, which would increase our understanding of biologically based susceptibility to age-related health problems. CONCLUSIONS: There is a growing awareness that age-related deterioration will affect an ever growing number of people, in both absolute and relative terms. It can be expected that this will further increase the resources that will be made available for research on aging. Although ultimately unavoidable, aging is a process that appears to be experimentally accessible. Therefore, the mechanisms of senescence and death may eventually be more completely understood, with the promise of preventing and/or delaying many of the adverse effects associated with aging, including most of the common diseases, and possibly also of extending lifespan.

Aged↗

PubMed1999

Intercellular bridges between granulosa cells and the oocyte in the elasmobranch Raya asterias.

In the present ultrastructural study intercellular bridges, connecting somatic granulosa cells to oocyte, have been detected for the first time and their modifications have been followed during Raja oogenesis. Intercellular bridges make their first appearance in small previtellogenic follicles as connecting devices between small cells and the oocyte. Later on, when the follicular epithelium becomes polymorphic and multilayered, for the presence of small, large, and pyriform-like cells, intercellular bridges link the oocyte and the different granulosa cells. Intercellular bridges contain ribosomes, whorl of membranes, mitochondria and vacuoles. Such cytoplasmic components are present also in the cell apex of large and pyriform-like cells thus suggesting, in agreement with other species (Motta et al. J. Exp. Zool., 1996;276:223-241) they may flow toward the oocyte. In this regard the presence of intercellular bridges during the oogenesis of cartilagineous fish may represent a crucial event of the active cooperation between granulosa cells and the oocyte.

Animals↗

PubMed2002

The significance of telomerase activation and cellular immortalization in human cancer.

Repression of telomerase in the somatic tissues of humans, and probably other long-lived mammals, appears to have evolved as a powerful protective barrier against cancer. Immortalization in vitro of normal human cells that lack telomerase involves the reactivation of telomerase or, rarely, an alternative (ALT) mechanism for maintaining telomeres. Inactivation of the effectors of replicative senescence, i.e. genes encoding one or more elements of the p16/pRB and/or ARF/p53/p21 anti-proliferative pathways, is required for telomerase depression leading to immortalization. Regulation of telomerase in normal human cells is mediated primarily by transcriptional repression of hTERT, the gene encoding the catalytic subunit of telomerase. Rodent cells do not possess stringent controls on telomerase activity in the soma and this explains why they are so readily immortalized and transformed in culture compared with their human counterparts. Because active telomerase has been found to exist in the proliferative compartments of self-renewing tissues, it is not yet clear whether the telomerase present in 90% of human cancers exists as a consequence of selection of pre-existing telomerase-positive cells during carcinogenesis or through induction of hTERT expression in cells in which it is normally tightly repressed. In support of the latter, chromosome transfer techniques have revealed the presence of genes on normal human chromosomes that are able to extinguish hTERT transcription in cancer cells and induce them to undergo senescence. It is clear that telomerase is obligatory for continuous tumour cell proliferation, clonal evolution and malignant progression. Telomerase therefore represents an attractive target at which to aim new anti-cancer drugs. Results with a variety of telomerase inhibitory strategies in human cancer cells have confirmed that its functional inactivation results in progressive telomere shortening, leading to growth arrest and/or cell death through apoptosis. Promising candidate small molecule inhibitors are beginning to emerge that will form the basis for anti-telomerase drug development.

Animals↗

PubMed2000

Functional conservation of the active sites of human and Drosophila angiotensin I-converting enzyme.

Human somatic angiotensin I-converting enzyme (sACE) has two active sites present in two homologous protein domains, resulting from a tandem gene duplication. It has been proposed that the N- and C-terminal active sites can have specific in vivo roles. In Drosophila melanogaster, Ance and Acercode for two ACE-like single-domain proteins, also predicted to have distinct physiological roles. We have investigated the relationship of Ance and Acer to the N- and C-domains of human sACE by genomic sequence analysis and by using domain-selective inhibitors, including RXP 407, a selective inhibitor of the human N-domain. These phosphinic peptides were potent inhibitors of Acer, but not of Ance. We conclude that the active sites of the N-domain and of Acer share structural features that permit the binding of the unusual RXP407 inhibitor and the hydrolysis of a broader range of peptide structures. In comparison, Ance, like the human C-domain of ACE, displays greater inhibitor selectivity. From the analysis of the published sequence of the Adh region of Drosophila chromosome 2, which carries Ance, Acer, and four additional ACE-like genes, we also suggest that this functional conservation is reflected in an ancestral gene structure identifiable in both protostome and deuterostome lineages and that the duplication seen in vertebrate genomes predates the divergence of these lineages. The conservation of ACE enzymes with distinct active sites in the evolution of both vertebrate and invertebrate species provides further evidence that these two kinds of active sites have different physiological functions.

Acetylation↗