The calcitonin/calcitonin gene-related peptide (CGRP) pre-mRNA is alternatively processed in a tissue-specific manner leading to the production of calcitonin mRNA in thyroid C cells and CGRP mRNA in neurons. A candidate calcitonin/CGRP splice regulator (CSR) isolated from rat brain was shown to inhibit calcitonin-specific splicing in vitro. CSR specifically binds to two regions in the calcitonin-specific exon 4 RNA previously demonstrated to function as a bipartate exonic splice enhancer (ESE). The two regions, A and B element, are necessary for inclusion of exon 4 into calcitonin mRNA. A novel RNA footprinting method based on the UV cross-linking assay was used to define the site of interaction between CSR and B element RNA. Base changes at the CSR binding site prevented CSR binding to B element RNA and CSR was unable to inhibit in vitro splicing of pre-mRNAs containing the mutated CSR binding site. When expressed in cells that normally produce predominantly CGRP mRNA, a calcitonin/CGRP gene containing the mutated CSR binding site expressed predominantly calcitonin mRNA. These observations demonstrate that CSR binding to the calcitonin-specific ESE regulates calcitonin/CGRP pre-mRNA splicing.
"LocusID 24241 maps to probe sets 1369116_a_at, 1369117_at and 1370775_a_at (Table 4), and corresponds to the rat calcitonin alpha/calcitonin gene-related peptide (CGRP) gene. The mammalian Calca/CGRP gene has six exons, and is considered a model gene for the study of alternative splicing . Splicing together the first four exons produces the mRNA for calcitonin alpha, whereas splicing together exons one to three, five and six produces the mRNA for CGRP (Figure 6). "
[Show abstract][Hide abstract] ABSTRACT: Affymetrix GeneChip technology enables the parallel observations of tens of thousands of genes. It is important that the probe set annotations are reliable so that biological inferences can be made about genes which undergo differential expression. Probe sets representing the same gene might be expected to show similar fold changes/z-scores, however this is in fact not the case.
We have made a case study of the mouse Surf4, chosen because it is a gene that was reported to be represented by the same eight probe sets on the MOE430A array by both Affymetrix and Bioconductor in early 2004. Only five of the probe sets actually detect Surf4 transcripts. Two of the probe sets detect splice variants of Surf2. We have also studied the expression changes of the eight probe sets in a public-domain microarray experiment. The transcripts for Surf4 are correlated in time, and similarly the transcripts for Surf2 are also correlated in time. However, the transcripts for Surf4 and Surf2 are not correlated. This proof of principle shows that observations of expression can be used to confirm, or otherwise, annotation discrepancies. We have also investigated groups of probe sets on the RAE230A array that are assigned to the same LocusID, but which show large variances in differential expression in any one of three different experiments on rat. The probe set groups with high variances are found to represent cases of alternative splicing, use of alternative poly(A) signals, or incorrect annotations.
Our results indicate that some probe sets should not be considered as unique measures of transcription, because the individual probes map to more than one transcript dependent upon the biological condition. Our results highlight the need for care when assessing whether groups of probe sets all measure the same transcript.
"The DNA to mRNA transcription with alternative RNA splicing was found in 1982 for Calcitonin/CGRP gene expression . We draw focus to model Calcitonin/CGRP gene expression while combining other recent biological knowledge  . As in Figure 2, the Calcitonin gene expression consists of four introns and five exons, and the transcription process progresses with step1, step2, step3 and step4. "
[Show abstract][Hide abstract] ABSTRACT: The research on modeling and simulation of complex biological systems is getting more important in Systems Biology. In this respect, we have developed Hybrid Function Petri net (HFPN) that was newly developed from existing Petri net because of their intuitive graphical representation and their capabilities for mathematical analyses. However, in the process of modeling metabolic, gene regulatory or signal transduction pathways with the architecture, we have realized three extensions of HFPN, (i) an entity should be extended to contain more than one value, (ii) an entity should be extended to handle other primitive types, e.g. boolean, string, (iii) an entity should be extended to handle more advanced type called object that consists of variables and methods, are necessary for modeling biological systems with Petri net based architecture. To deal with it, we define a new enhanced Petri net called hybrid functional Petri net with extension (HFPNe). To demonstrate the effectiveness of the enhancements, we model and simulate with HFPNe four biological processes that are diffcult to represent with the previous architecture HFPN.
Genome informatics. International Conference on Genome Informatics 02/2004; 15(1):180-97.
[Show abstract][Hide abstract] ABSTRACT: The calcitonin/calcitonin gene-related peptide (CGRP) pre-mRNA is alternatively processed in a tissue-specific manner leading to the production of calcitonin mRNA in thyroid C cells and CGRP mRNA in neurons. Sequences in the human calcitonin-specific fourth exon function as an exonic splice enhancer (ESE) which is required for incorporation of exon 4 into calcitonin mRNA. Deletion of these sequences from the rat calcitonin/CGRP gene was reported to have no effect on calcitonin splicing. We demonstrate that sequences in the rat calcitonin/CGRP fourth exon act as an ESE. In addition, we observed that three proteins in HeLa nuclear extract, of apparent molecular weights of 40, 55 and 85 kDa, specifically interact with the exon 4 ESE. The 40-kDa protein is human transformer 2β (hTra2β), a homolog of the Drosophila splice regulator transformer 2. hTra2β is required for calcitonin splicing in vitro, one of the first biological functions identified for hTra2β. The 55-kDa protein is SRp55, a member of the SR family of phosphoproteins. Binding of SRp55 to an ESE required for calcitonin mRNA splicing suggests that the different levels of SRp55 present in different cell types may regulate calcitonin/CGRP alternative splicing.
Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression 01/2003; 1625(2):141-152. DOI:10.1016/S0167-4781(02)00600-0 · 1.70 Impact Factor
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