Trans-splicing in Higher Eukaryotes: Implications for Cancer Development?

Trans-splicing, the possibility of exons from distinct pre-mRNAs to join together, is still a concept in gene expression that is generally regarded of limited significance. However, recent work has provided evidence that in human tumors trans-splicing events may precede chromosomal rearrangements. In fact, it has been suggested that the trans-spliced molecules could act as “guides” that facilitate the genomic translocation. This perspective highlights the development of the ideas of trans-splicing in higher eukaryotes during the last 25 years, from a bizarre phenomenon to a biological event that is attaining stronger recognition

Genetic variations regulate alternative splicing in the 5' untranslated regions of the mouse glioma-associated oncogene 1, Gli1

<p>Abstract</p> <p>Background</p> <p>Alternative splicing is one of the key mechanisms that generate biological diversity. Even though alternative splicing also occurs in the 5' and 3' untranslated regions (UTRs) of mRNAs, the understanding of the significance and the regulation of these variations is rather limited.</p> <p>Results</p> <p>We investigated 5' UTR mRNA variants of the mouse Gli1 oncogene, which is the terminal transcriptional effector of the Hedgehog (HH) signaling pathway. In addition to identifying novel transcription start sites, we demonstrated that the expression ratio of the Gli1 splice variants in the 5' UTR is regulated by the genotype of the mouse strain analyzed. The GT allele, which contains the consensus intronic dinucleotides at the 5' splice site of intron 1B, favors exon 1B inclusion, while the GC allele, having a weaker 5' splice site sequence, promotes exon 1B skipping. Moreover, the alternative Gli1 5' UTRs had an impact on translational capacity, with the shorter and the exon 1B-skipped mRNA variants being most effective.</p> <p>Conclusions</p> <p>Our findings implicate novel, genome-based mechanisms as regulators of the terminal events in the mouse HH signaling cascade.</p

Isolation and characterization of a novel cytochrome P-450-like pseudogene

SummaryA rabbit liver P-450-like pseudogene has been isolated from a [lambda] phage genomic library. Sequence analysis revealed structural homology with respect to the rat P-450b and P-450e genes as well as a similar intron-exon organization. A 5'-proximal TATA box-like sequence and two 3'-distal putative polyadenylation signals were identified, and all putative intronexon boundaries except at the 3'-splice site of intron 2 were found to follow the GT/AG rule. With allowance for apparent deletions and insertions, the structural homology of the amino acid sequence deduced from the pseudogene with respect to rabbit P-450 isozyme 2 is lower for exons 1 through 4 (18-28%) than for exons 5 through 9 (42-65%). S1 nuclease mapping showed that mRNAs complementary to the DNA sequence of exon 9 are expressed. However, due to the alterations in the pseudogene, it appears that functional P-450 would not be produced from such mRNAs.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/26288/1/0000373.pd

Tamoxifen treatment of breast cancer cells : Impact on Hedgehog/GLI1 signaling

The selective estrogen receptor (ER) modulator tamoxifen (TAM) has become the standard therapy for the treatment of ER+ breast cancer patients. Despite the obvious benefits of TAM, a proportion of patients acquire resistance to treatment, and this is a significant clinical problem. Consequently, the identification of possible mechanisms involved in TAM-resistance should help the development of new therapeutic targets. In this study, we present in vitro data using a panel of different breast cancer cell lines and demonstrate the modulatory effect of TAM on cellular proliferation and expression of Hedgehog signaling components, including the terminal effector of the pathway, the transcription factor GLI1. A variable pattern of expression following TAM administration was observed, reflecting the distinctive properties of the ER+ and ER´ cell lines analyzed. Remarkably, the TAM-induced increase in the proliferation of the ER+ ZR-75-1 and BT474 cells parallels a sustained upregulation of GLI1 expression and its translocation to the nucleus. These findings, implicating a TAM-GLI1 signaling cross-talk, could ultimately be exploited not only as a means for novel prognostication markers but also in efforts to effectively target breast cancer subtypes. © 2016 by the authors; licensee MDPI, Basel, Switzerland

Tamoxifen treatment of breast cancer cells: Impact on Hedgehog/GLI1 signaling

The selective estrogen receptor (ER) modulator tamoxifen (TAM) has become the standard therapy for the treatment of ER+ breast cancer patients. Despite the obvious benefits of TAM, a proportion of patients acquire resistance to treatment, and this is a significant clinical problem. Consequently, the identification of possible mechanisms involved in TAM-resistance should help the development of new therapeutic targets. In this study, we present in vitro data using a panel of different breast cancer cell lines and demonstrate the modulatory effect of TAM on cellular proliferation and expression of Hedgehog signaling components, including the terminal effector of the pathway, the transcription factor GLI1. A variable pattern of expression following TAM administration was observed, reflecting the distinctive properties of the ER+ and ER− cell lines analyzed. Remarkably, the TAM-induced increase in the proliferation of the ER+ ZR-75-1 and BT474 cells parallels a sustained upregulation of GLI1 expression and its translocation to the nucleus. These findings, implicating a TAM-GLI1 signaling cross-talk, could ultimately be exploited not only as a means for novel prognostication markers but also in efforts to effectively target breast cancer subtypes

Circular RNAs Are the Predominant Transcript Isoform from Hundreds of Human Genes in Diverse Cell Types

Most human pre-mRNAs are spliced into linear molecules that retain the exon order defined by the genomic sequence. By deep sequencing of RNA from a variety of normal and malignant human cells, we found RNA transcripts from many human genes in which the exons were arranged in a non-canonical order. Statistical estimates and biochemical assays provided strong evidence that a substantial fraction of the spliced transcripts from hundreds of genes are circular RNAs. Our results suggest that a non-canonical mode of RNA splicing, resulting in a circular RNA isoform, is a general feature of the gene expression program in human cells

A promoter-associated RNA downregulates the oncogenic GLI1 transcription factor in rhabdomyosarcoma cells

Recent experimental evidence demonstrates a novel regulatory mechanism on the expression of the GLI1 oncogene, a transcriptional effector of Hedgehog signaling. This is mediated by a non-coding RNA, antisense to the GLI1 promoter, GLI1AS, which elicits negative feedback on GLI1 expression. Knockdown of GLI1AS was shown to enhance rhabdomyosarcoma tumor growth in a xenograft model, in-line with the observed increase of the expression levels of GLI1, a known proliferative/oncogenic factor in this cellular context

Mechanisms of pre-mRNA splicing: classical versus non-classical pathways

Expression of genetic information proceeds through two major biological events, transcription and translation. However, in eukaryotic cells, the primary transcript (pre-mRNA) is not the template that the translational apparatus scans through, in order to produce the corresponding protein. Pre-mRNAs undergo several modifications (cap site addition, poly At tail addition) prior to becoming mature mRNAs, with the most important one being the excision (splicing) of the intronic sequences. Yet, the mechanisms that regulate the splicing process and the generation of alternatively spliced mRNA products are still poorly understood. Moreover recent findings suggest that this process also has the capability to produce an additional set of RNA products that differ from typical mRNA molecules. In these novel RNA transcripts the order of the exons has been changed relative to genomic DNA. Furthermore, the properties of these transcripts suggest that they may represent circular RNA molecules

Isolation And Characterization Of A Novel Cytochrome P-450-like Gene.

Cytochrome P-450, a pigment occurring widely in nature, is involved in the oxygenation and other metabolic reactions of a large variety of drugs, carcinogens, and other xenobiotics, as well as naturally occurring substances such as steroids and prostaglandins. A number of P-450 isozymes have already been purified to electrophoretic homogeneity from the rabbit, rat, and mouse, as well as from other species, and the amino acid sequence is known in several instances. Two major classes of P-450 inducers are known, represented by the drug phenobarbital and the chemical carcinogen methylcholanthrene. A rabbit liver genomic library was screened using a cDNA clone coding for a phenobarbital-induced rat P-450. A positive plaque was isolated and used to characterize a novel P-450-like gene. The new gene resembles known genes of the phenobarbital family in having nine exons and in the location of the intron-exon junctions. The predicted sequence of the corresponding protein was calculated to have an overall structural homology of 40% with respect to isozyme 2, the major phenobarbital-inducible P-450 in the rabbit. The homology is particularly evident in three small regions common to the cytochrome P-450 family, including the cysteine-containing peptide believed to provide the sulfur ligand to the heme iron atom. Several lines of evidence, including S(,1) mapping, indicate that the gene is expressed. However, neither phenobarbital nor isosafrole, a typical inducer of the methylcholanthrene type, appear to enhance the expression of the gene in rabbit liver. Of particular interest, the amino acid sequence deduced from the gene has higher homology at the carboxyl-terminal region than at the amino-terminal region as compared with the corresponding sequence of isozyme 2.Ph.D.Biological SciencesMolecular biologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/127786/2/8521020.pd