Protein Coding Region Of Gene Research Articles

Genome-wide DNA methylation patterns in LSH mutant reveals de-repression of repeat elements and redundant epigenetic silencing pathways.

Cytosine methylation is critical in mammalian development and plays a role in diverse biologic processes such as genomic imprinting, X chromosome inactivation, and silencing of repeat elements. Several factors regulate DNA methylation in early embryogenesis, but their precise role in the establishment of DNA methylation at a given site remains unclear. We have generated a comprehensive methylation map in fibroblasts derived from the murine DNA methylation mutant Hells−/− (helicase, lymphoid specific, also known as LSH). It has been previously shown that HELLS can influence de novo methylation of retroviral sequences and endogenous genes. Here, we describe that HELLS controls cytosine methylation in a nuclear compartment that is in part defined by lamin B1 attachment regions. Despite widespread loss of cytosine methylation at regulatory sequences, including promoter regions of protein-coding genes and noncoding RNA genes, overall relative transcript abundance levels in the absence of HELLS are similar to those in wild-type cells. A subset of promoter regions shows increases of the histone modification H3K27me3, suggesting redundancy of epigenetic silencing mechanisms. Furthermore, HELLS modulates CG methylation at all classes of repeat elements and is critical for repression of a subset of repeat elements. Overall, we provide a detailed analysis of gene expression changes in relation to DNA methylation alterations, which contributes to our understanding of the biological role of cytosine methylation.

LIN-42, the Caenorhabditis elegans PERIOD homolog, negatively regulates microRNA transcription.

During C. elegans development, microRNAs (miRNAs) function as molecular switches that define temporal gene expression and cell lineage patterns in a dosage-dependent manner. It is critical, therefore, that the expression of miRNAs be tightly regulated so that target mRNA expression is properly controlled. The molecular mechanisms that function to optimize or control miRNA levels during development are unknown. Here we find that mutations in lin-42, the C. elegans homolog of the circadian-related period gene, suppress multiple dosage-dependent miRNA phenotypes including those involved in developmental timing and neuronal cell fate determination. Analysis of mature miRNA levels in lin-42 mutants indicates that lin-42 functions to attenuate miRNA expression. Through the analysis of transcriptional reporters, we show that the upstream cis-acting regulatory regions of several miRNA genes are sufficient to promote highly dynamic transcription that is coupled to the molting cycles of post-embryonic development. Immunoprecipitation of LIN-42 complexes indicates that LIN-42 binds the putative cis-regulatory regions of both non-coding and protein-coding genes and likely plays a role in regulating their transcription. Consistent with this hypothesis, analysis of miRNA transcriptional reporters in lin-42 mutants indicates that lin-42 regulates miRNA transcription. Surprisingly, strong loss-of-function mutations in lin-42 do not abolish the oscillatory expression patterns of lin-4 and let-7 transcription but lead to increased expression of these genes. We propose that lin-42 functions to negatively regulate the transcriptional output of multiple miRNAs and mRNAs and therefore coordinates the expression levels of genes that dictate temporal cell fate with other regulatory programs that promote rhythmic gene expression.

Identification of miRNAs that specifically target tumor suppressive KLF6-FL rather than oncogenic KLF6-SV1 isoform

The Krüppel like factor 6 (KLF6) gene encodes multiple protein isoforms derived from alternative mRNA splicing, most of which are intimately involved in hepatocarcinogenesis and tumor progression. Recent bioinformatics analysis shows that alternative mRNA splicing of the KLF6 gene produces around 16 alternatively spliced variants with divergent or even opposing functions. Intriguingly, the full-length KLF6 (KLF6-FL) is a tumor suppressor gene frequently inactivated in liver cancer, whereas KLF6 splice variant 1 (KLF6-SV1) is an oncogenic isoform with antagonistic function against KLF6-FL. Compelling evidence indicates that miRNA, the small endogenous non-coding RNA (ncRNA), acts as a vital player in modulating a variety of cellular biological processes through targeting different mRNA regions of protein-coding genes. To identify the potential miRNAs specifically targeting KLF6-FL, we utilized bioinformatics analysis in combination with the luciferase reporter assays and screened out two miRNAs, namely miR-210 and miR-1301, specifically targeted the tumor suppressive KLF6-FL rather than the oncogenic KLF6-SV1. Our in vitro experiments demonstrated that stable expression of KLF6-FL inhibited cell proliferation, migration and angiogenesis while overexpression of miR-1301 promoted cell migration and angiogenesis. Further experiments demonstrated that miR-1301 was highly expressed in liver cancer cell lines as well as clinical specimens and we also identified the potential methylation and histone acetylation for miR-1301 gene. To sum up, our findings unveiled a novel molecular mechanism that specific miRNAs promoted tumorigenesis by targeting the tumor suppressive isoform KLF6-FL rather than its oncogenic isoform KLF6-SV1.

The wide expansion of hepatitis delta virus-like ribozymes throughout trypanosomatid genomes is linked to the spreading of L1Tc/ingi clade mobile elements.

BackgroundHepatitis Delta Virus (HDV)-like ribozymes have recently been found in many mobile elements in which they take part in a mechanism that releases intermediate RNAs from cellular co-transcripts. L1Tc in Trypanosoma cruzi is one of the elements in which such a ribozyme is located. It lies in the so-called Pr77-hallmark, a conserved region shared by retrotransposons belonging to the trypanosomatid L1Tc/ingi clade. The wide distribution of the Pr77-hallmark detected in trypanosomatid retrotransposons renders the potential catalytic activity of these elements worthy of study: their distribution might contribute to host genetic regulation at the mRNA level. Indeed, in Leishmania spp, the pervasive presence of these HDV-like ribozyme-containing mobile elements in certain 3′-untranslated regions of protein-coding genes has been linked to mRNA downregulation.ResultsIntensive screening of publicly available trypanosomatid genomes, combined with manual folding analyses, allowed the isolation of putatively Pr77-hallmarks with HDV-like ribozyme activity. This work describes the conservation of an HDV-like ribozyme structure in the Pr77 sequence of retrotransposons in a wide range of trypanosomatids, the catalytic function of which is maintained in the majority.These results are consistent with the previously suggested common phylogenetic origin of the elements that belong to this clade, although in some cases loss of functionality appears to have occurred and/or perhaps molecular domestication by the host.ConclusionsThese HDV-like ribozymes are widely distributed within retrotransposons across trypanosomatid genomes. This type of ribozyme was once thought to be rare in nature, but in fact it would seem to be abundant in trypanosomatid transcripts. It can even form part of the pool of mRNA 3′-untranslated regions, particularly in Leishmania spp. Its putative regulatory role in host genetic expression is discussed.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-340) contains supplementary material, which is available to authorized users.

Evolutionary conservation in genes underlying human psychiatric disorders.

Many psychiatric diseases observed in humans have tenuous or absent analogs in other species. Most notable among these are schizophrenia and autism. One hypothesis has posited that these diseases have arisen as a consequence of human brain evolution, for example, that the same processes that led to advances in cognition, language, and executive function also resulted in novel diseases in humans when dysfunctional. Here, the molecular evolution of the protein-coding regions of genes associated with these and other psychiatric disorders are compared among species. Genes associated with psychiatric disorders are drawn from the literature and orthologous sequences are collected from eleven primate species (human, chimpanzee, bonobo, gorilla, orangutan, gibbon, macaque, baboon, marmoset, squirrel monkey, and galago) and 34 non-primate mammalian species. Evolutionary parameters, including dN/dS, are calculated for each gene and compared between disease classes and among species, focusing on humans and primates compared to other mammals, and on large-brained taxa (cetaceans, rhinoceros, walrus, bear, and elephant) compared to their small-brained sister species. Evidence of differential selection in humans to the exclusion of non-human primates was absent, however elevated dN/dS was detected in catarrhines as a whole, as well as in cetaceans, possibly as part of a more general trend. Although this may suggest that protein changes associated with schizophrenia and autism are not a cost of the higher brain function found in humans, it may also point to insufficiencies in the study of these diseases including incomplete or inaccurate gene association lists and/or a greater role of regulatory changes or copy number variation. Through this work a better understanding of the molecular evolution of the human brain, the pathophysiology of disease, and the genetic basis of human psychiatric disease is gained.

Efficient purging of deleterious mutations in plants with haploid selfing.

In diploid organisms, selfing reduces the efficiency of selection in removing deleterious mutations from a population. This need not be the case for all organisms. Some plants, for example, undergo an extreme form of selfing known as intragametophytic selfing, which immediately exposes all recessive deleterious mutations in a parental genome to selective purging. Here, we ask how effectively deleterious mutations are removed from such plants. Specifically, we study the extent to which deleterious mutations accumulate in a predominantly selfing and a predominantly outcrossing pair of moss species, using genome-wide transcriptome data. We find that the selfing species purge significantly more nonsynonymous mutations, as well as a greater proportion of radical amino acid changes which alter physicochemical properties of amino acids. Moreover, their purging of deleterious mutation is especially strong in conserved regions of protein-coding genes. Our observations show that selfing need not impede but can even accelerate the removal of deleterious mutations, and do so on a genome-wide scale.

Prediction of Translation Initiation Site in Bacterial and Archaeal Genomes

Abstract: Driven by the rapid growth of the complete genome sequences, it is accepted that genome annotation has been resorted mostly to automatic methodology. For computational annotation to bacterial and archaeal genomes, accurate prediction of translation initiation sites (TISs) is essential to locate protein coding regions of genes. Therefore, TIS prediction has been a challenge to a number of gene finders and TIS processors, leading to recent studies of TIS prediction or correction in prokaryotic genome annotation as well as of the mechanism of translation initiation. It is time for the research community to review the available mathematical models of TIS of prokaryotic gene, and the resultant algorithms for a series of current TIS processors and TIS prediction modules in gene finders. In fact, the TIS models have been improved along with the knowledge of the mechanism of translation initiation. Several studies of the mechanism of translation initiation in prokaryotic genomes have been summarized. With a few of published data sets widely-used in evaluation of TIS identification, the performances of the existing methods are assessed and discussed in this article. It is also interesting to discuss the relation between the algorithms and the understanding of prokaryotic translation initiation mechanism, which can enlighten us on the state-of-the-art studies of TIS prediction in bacterial and archaeal genomes.

Exploring the Regulation Mechanism of miRNAs Transcription Using Genomic and Epigenetic Functional Annotations

Uncovering the transcriptional regulation mechanism of mammalian miRNAs is crucial to understand the role of these tiny regulators in cellular processes. Based on 1,030 genomic and epigenetic functional annotations, we compared these features of regulatory regions between miRNAs and protein-coding genes. Support vector machine (SVM) was used to quantify the contribution of each annotation group or group combination in distinguishing miRNA regulatory regions. We observed fewer repetitive DNA elements and SNPs, but more intensive DNA methylation in the miRNA regulatory regions. On the contrary, there are more predicted CpG islands and higher H3K9me1 levels in the regulatory regions of protein-coding genes. This analysis indicated that epigenetic factors such as DNA methylation and specific histone marks are the most informative groups, suggesting epigenetic events may play a more important role in miRNA transcription than previously thought. Furthermore, these results also revealed the interactive effects among various genomic or epigenetic factors involved in miRNA transcription.

Long non-coding rnas in osteoarthritis and chondrogenesis

Long non-coding rnas in osteoarthritis and chondrogenesis

Intraspecific comparison of complete mitogenome sequences from two Asian raccoon dogs (Canidae: Nyctereutes procyonoides)

We determined the complete mitochondrial genome (GenBank accession number: KF709435) of the Korean raccoon dog Nyctereutes procyonoides koreensis and compared it with a previously published mitogenome (GenBank accession number: GU256221) of a Chinese raccoon dog. The total length of N. p. koreensis mitogenome is 16,802 bp, with a base composition of 32.1% A, 26.9% T, 26.8% C and 14.2% G. High similarity of 98.7% was found between the complete mitogenome sequences of Korean and Chinese raccoon dogs. Sequence similarity of the two mitogenomes was 99.3% in the other gene regions except for D-loop. The sequence similarity of 99.1% was found in the 13 protein-coding gene regions, whereas 99.6% was identical in mtDNA regions covering all the 22 tRNA genes. There was no variation between 12S rRNAs, whereas 0.5% difference was found between 16S rRNAs.

Effects of regulatory evolution on morphological diversity

An important task in evolutionary biology is to understand the reason for and mechanisms of morphological diversification. Studies in evolutionary developmental biology have revealed that, rather than being invented repeatedly from scratch, many complex morphological structures have evolved by modification of ancient regulatory networks. In other words, morphological diversity is not always produced by changes in the protein-coding region of regulatory genes; rather, it largely depends on the evolution of gene regulation. As the main components of the regulatory regions of a gene, cis regulatory elements bind to specific trans factors and determine the precise expression of the gene in time, place and amount. As a result, gain, loss, change or modification of cis regulatory elements may lead to shifts in gene expression, which, in turn, generate morphological diversity. Here, by reviewing recent progress in this and related fields, we summarize the basic features of gene regulation in eukaryotes, elucidating its fundamental evolutionary pattern and revealing its importance in generating morphological diversity.

MiRIAD-integrating microRNA inter- and intragenic data.

MicroRNAs (miRNAs) are a class of small (∼22 nucleotides) non-coding RNAs that post-transcriptionally regulate gene expression by interacting with target mRNAs. A majority of miRNAs is located within intronic or exonic regions of protein-coding genes (host genes), and increasing evidence suggests a functional relationship between these miRNAs and their host genes. Here, we introduce miRIAD, a web-service to facilitate the analysis of genomic and structural features of intragenic miRNAs and their host genes for five species (human, rhesus monkey, mouse, chicken and opossum). miRIAD contains the genomic classification of all miRNAs (inter- and intragenic), as well as classification of all protein-coding genes into host or non-host genes (depending on whether they contain an intragenic miRNA or not). We collected and processed public data from several sources to provide a clear visualization of relevant knowledge related to intragenic miRNAs, such as host gene function, genomic context, names of and references to intragenic miRNAs, miRNA binding sites, clusters of intragenic miRNAs, miRNA and host gene expression across different tissues and expression correlation for intragenic miRNAs and their host genes. Protein–protein interaction data are also presented for functional network analysis of host genes. In summary, miRIAD was designed to help the research community to explore, in a user-friendly environment, intragenic miRNAs, their host genes and functional annotations with minimal effort, facilitating hypothesis generation and in-silico validations.Database URL: http://www.miriad-database.org

Evaluating summary statistics used to test for incomplete lineage sorting: mito‐nuclear discordance in the reef sponge Callyspongia vaginalis

Conflicting patterns of population differentiation between the mitochondrial and nuclear genomes (mito-nuclear discordance) have become increasingly evident as multilocus data sets have become easier to generate. Incomplete lineage sorting (ILS) of nucDNA is often implicated as the cause of such discordance, stemming from the large effective population size of nucDNA relative to mtDNA. However, selection, sex-biased dispersal and historical demography can also lead to mito-nuclear discordance. Here, we compare patterns of genetic diversity and subdivision for six nuclear protein-coding gene regions to those for mtDNA in a common Caribbean coral reef sponge, Callyspongia vaginalis, along the Florida reef tract. We also evaluated a suite of summary statistics to determine which are effective metrics for comparing empirical and simulated data when testing drivers of mito-nuclear discordance in a statistical framework. While earlier work revealed three divergent and geographically subdivided mtDNACOI haplotypes separated by 2.4% sequence divergence, nuclear alleles were admixed with respect to mitochondrial clade and geography. Bayesian analysis showed that substitution rates for the nuclear loci were up to 7 times faster than for mitochondrial COI. Coalescent simulations and neutrality tests suggested that mito-nuclear discordance in C. vaginalis is not the result of ILS in the nucDNA or selection on the mtDNA but is more likely caused by changes in population size. Sperm-mediated gene flow may also influence patterns of population subdivision in the nucDNA.

TERT Gene Promoter Mutations In Myelodysplastic Syndromes (MDS)

Mutations at the protein-coding region of TERT gene (chromosome 5p15.33) have been well characterized in patients affected by a constitutional telomere syndrome, including diskeratosis, aplastic anemia and pulmonary fibrosis (Calado RT, Hematology, ASH 2009:338). In rare cases somatic mutations may occur in de novo MDS/AML (Calado RT, Young NS. Blood 2008;111:4446). Mutations involving the regulatory region of TERT gene have been recently identified in a consistent proportion of familial and sporadic melanoma and in a subset of tumors originating from tissues with low rate of self-renewal (glioblastoma, liposarcoma, oligodendroglioma bladder cancer, upper urinary tract cancer). These mutations create new binding motifs for Ets/TCF transcription factors, thus increasing TERT gene transcription (Killela PJ et al, PNAS 2013;110:6021; Horn S et al, Science 2013;339:959). As far as we know TERT promoter mutations were never described in MDS. Material and MethodsIndex Case. A 52 years old woman was selected because of a complex phenotype including abnormal skin pigmentation, familial pulmonary fibrosis, osteoarthritis, and a refractory cytopenia with multilineage dysplasia (RCMD, WHO 2008) with a 47,XX,+8 karyotype.Screening Cases. Mutational analysis was extended to a cohort of 115 patients (72 males; 44 females, median age 69) referred to the Laboratory of Cytogenetics, Hematology Department,University of Perugia, Italy.Cytogenetics was normal in 49 cases (43%). Abnormalities included: isolated del(5q) (seven cases, 6%); del(5q) plus one additional change (three cases, 3%); isolated del(20q) (fourteen cases, 12%); trisomy 8 (five cases, 4%); monosomy 7 (two cases, 2%); -Y (three cases, 3%); del(11q) (two cases, 2%); complex karyotype (twenty-four cases, 21%); other aberrations (six cases, 5%).Genomic DNA was extracted from bone marrow (BM) of all cases and from peripheral blood T lymphocytes of index case using Salting out method. In all cases TERT promoter was screened using PCR based DHPLC assay (Wave system; MD Transgenomic Inc. Omaha, NE). The 274bp amplicon was amplified with forward primer 5'GTCCTGCCCCTTCACCTTC3' and reverse primer 5'AGCACCTCGCGGTAGTGG3' using Robust Start Taq KAPA2G (Biosystems, Woburn, MA). DNA from abnormal elution profiles were re-amplified and sequenced by Sanger method (ABI 3500 Genetic Analyzer, Applied Biosystems). Variations were detected using Finch TV v. 1.4.0. In the index case 23 amplicons encompassing all 16 exons of TERT gene were also screened (NC_000005.9). Mutations cloning was carried out after RNA extraction (Trizol, Invitrogen, Life Technologies, Paisley, UK), reverse transcription (ThermoscriptTM RT-PCR System, Invitrogen) and amplification (TERT_2CF (5'-CAGCGCTACTGGCAAATGCG-3' Ta-61,4°C; and TERT_2543R (5'-GGCACTGGACGTAGGACTTG-3' Ta-61,4°C). PCR products were sub-cloned into pGEM-T easy vector (Promega, Madison, WI, USA) and sequenced. ResultsIndex Case. Patient was a compound heterozygous for two germline variations: a nonsense mutation c.1209C>A p.C403* (exon 2) and a missense mutation c.2455C>T p.R819C (exon 8). A putative somatic A>C transition 57 bp before the ATG start codon was detected only in BM cells (Table).Screening Cases. DHPLC analysis showed three patients (2.6%) with a two-peak elution profile. Sequencing revealed a 10 bp duplication (c.1-110_1-101) in case 2; a c.1-124 C>T point mutation in case 3; a point mutation c.1-78 C>T in case 4 (Table). CommentWe showed that TERT gene promoter variations are recurrent events in 2.6% of MDS patients. Only low/int1 risk MDS (IPSS) were affected in this series. The c.1-57A>C, previously reported as a germline activating variation in familial melanoma (Horn S et al, Science 2013;339:959), was likely a somatic mutation in our index case, thus supporting its role in clonal MDS proliferation. We first found the melanoma activating c.1-124 C>T point mutation (Huang FW et al, Science 2013;339:957; Killela PJ et al, PNAS 2013;110:6021) in a MDS with isolated del(5q). The new variation of our case n.4 does not appear to introduce a new transcription factor binding site (http://www.cbrc.jp/research/db/TFSEARCH.html), whereas the 10bp duplication of case 2 indicates an hypothetical binding site for Ikaros. Further results from ongoing functional studies in these cases will be presented. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Primers for fourteen protein-coding genes and the deep phylogeny of the true yeasts

The Saccharomycetales or ‘true yeasts’ consist of more than 800 described species, including many of scientific, medical and commercial importance. Considerable progress has been made in determining the phylogenetic relationships of these species, largely based on rDNA sequences, but many nodes for early-diverging lineages cannot be resolved with rDNA alone. rDNA is also not ideal for delineating recently diverged species. From published full-genome sequence data, we have identified 14 regions of protein-coding genes that can be PCR-amplified in a large proportion of a diverse collection of 25 yeast species using degenerate primers. Phylogenetic analysis of the sequences thus obtained reveals a well-resolved phylogeny of the Saccharomycetales with many branches having high bootstrap support. Analysis of published sequences from the Saccharomyces paradoxus species complex shows that these protein-coding gene fragments are also informative about genealogical relationships amongst closely related strains. Our set of protein-coding gene fragments is therefore suitable for analysing both ancient and recent evolutionary relationships amongst yeasts.

Phylogenomics supports Panpulmonata: Opisthobranch paraphyly and key evolutionary steps in a major radiation of gastropod molluscs

Pulmonates, with over 30,000 described species, represent the largest radiation of non-marine animals outside of Arthropoda. The pulmonate lung was a key evolutionary innovation enabling diversification of terrestrial and freshwater snails and slugs. However, recent studies drew conflicting conclusions about pulmonate monophyly, and support for a sister group is lacking, hindering our understanding of this major animal radiation. Analyses of mitochondrial protein-coding genes recovered a paraphyletic Pulmonata grading into a monophyletic Opisthobranchia, a traditional group of sea slugs long considered sister to pulmonates. Conversely, analyses of datsets dominated by nuclear rDNA indicated Opisthobranchia is paraphyletic with respect to Pulmonata. No study resolved the placement of two key taxa: Sacoglossa, an opisthobranch group including photosynthetic sea slugs, and Siphonarioidea, intertidal limpet-like snails traditionally in Pulmonata. To examine evolutionary relationships at the base of the pulmonate radiation, we performed a phylogenomic analysis of 102 nuclear protein-coding gene regions for 19 gastropods. Opisthobranchia was recovered as paraphyletic with respect to Panpulmonata, a clade in which Sacoglossa was sister to Pulmonata, with Siphonarioidea as the basal pulmonate lineage. Siphonarioideans share a similar gill structure with shelled sacoglossans but lack the contractile pneumostome of pulmonates, suggesting descent from an evolutionary intermediate that facilitated the pulmonate radiation into non-marine habitats.

Molecular phylogeny of the genus Taenia (Cestoda: Taeniidae): Proposals for the resurrection of Hydatigera Lamarck, 1816 and the creation of a new genus Versteria

The cestode family Taeniidae generally consists of two valid genera, Taenia and Echinococcus. The genus Echinococcus is monophyletic due to a remarkable similarity in morphology, features of development and genetic makeup. By contrast, Taenia is a highly diverse group formerly made up of different genera. Recent molecular phylogenetic analyses strongly suggest the paraphyly of Taenia. To clarify the genetic relationships among the representative members of Taenia, molecular phylogenies were constructed using nuclear and mitochondrial genes. The nuclear phylogenetic trees of 18S ribosomal DNA and concatenated exon regions of protein-coding genes (phosphoenolpyruvate carboxykinase and DNA polymerase delta) demonstrated that both Taenia mustelae and a clade formed by Taenia parva, Taenia krepkogorski and Taenia taeniaeformis are only distantly related to the other members of Taenia. Similar topologies were recovered in mitochondrial genomic analyses using 12 complete protein-coding genes. A sister relationship between T. mustelae and Echinococcus spp. was supported, especially in protein-coding gene trees inferred from both nuclear and mitochondrial data sets. Based on these results, we propose the resurrection of Hydatigera Lamarck, 1816 for T. parva, T. krepkogorski and T. taeniaeformis and the creation of a new genus, Versteria, for T. mustelae. Due to obvious morphological and ecological similarities, Taenia brachyacantha is also included in Versteria gen. nov., although molecular evidence is not available. Taenia taeniaeformis has been historically regarded as a single species but the present data clearly demonstrate that it consists of two cryptic species.

Highly recurrent TERT promoter mutations in human melanoma.

Systematic sequencing of human cancer genomes has identified many recurrent mutations in the protein-coding regions of genes but rarely in gene regulatory regions. Here, we describe two independent mutations within the core promoter of telomerase reverse transcriptase (TERT), the gene coding for the catalytic subunit of telomerase, which collectively occur in 50 of 70 (71%) melanomas examined. These mutations generate de novo consensus binding motifs for E-twenty-six (ETS) transcription factors, and in reporter assays, the mutations increased transcriptional activity from the TERT promoter by two- to fourfold. Examination of 150 cancer cell lines derived from diverse tumor types revealed the same mutations in 24 cases (16%), with preliminary evidence of elevated frequency in bladder and hepatocellular cancer cells. Thus, somatic mutations in regulatory regions of the genome may represent an important tumorigenic mechanism.

TERT Promoter Mutations in Familial and Sporadic Melanoma

Cutaneous melanoma occurs in both familial and sporadic forms. We investigated a melanoma-prone family through linkage analysis and high-throughput sequencing and identified a disease-segregating germline mutation in the promoter of the telomerase reverse transcriptase (TERT) gene, which encodes the catalytic subunit of telomerase. The mutation creates a new binding motif for Ets transcription factors and ternary complex factors (TCFs) near the transcription start and, in reporter gene assays, caused up to twofold increase in transcription. We then screened the TERT promoter in sporadic melanoma and observed recurrent ultraviolet signature somatic mutations in 125 of 168 (74%) of human cell lines derived from metastatic melanomas, 45 of 53 corresponding metastatic tumor tissues (85%), and 25 of 77 (33%) primary melanomas. The majority of those mutations occurred at two positions in the TERT promoter and also generated binding motifs for Ets/TCF transcription factors.

Insights of potential G-quadruplex sequences in telomeres and proto-oncogenes

Guanine rich sequences have the ability to fold into stable 4 stranded structures called G-quadruplex under physiological concentrations of Na+ or K+. G-quadruplexes are found in telomeres, being stable structures under the control of telomerase binding proteins. They are also identified throughout the genome and are enriched in promoter regions of protein coding genes, upstream and downstream of the transcription initiation sites. A number of these promoter quadruplexes have been investigated for several proto-oncogenes. The formation of these quadruplexes can lead to chemical intervention of gene expression using a G-quadruplex binding ligand. We review location, configuration, and stabilization of these quadruplexes in some of the important promoters with regards to their potential as anticancer target.