کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8644375 | 1569758 | 2018 | 38 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Molecular cloning of novel transcripts of the adaptor-related protein complex 2 alpha 1 subunit (AP2A1) gene, using Next-Generation Sequencing
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کلمات کلیدی
NMDPremature translation termination codonNonsense-mediated mRNA decay (NMD)ISPPTCIGVNGSAdaptin - آدپتینSemiconductor sequencing - ترتیب نیمههادیNext-generation sequencing - تعیین توالی نسل بعدیAlternative splicing - جابجایی جایگزینnonsense-mediated mRNA decay - فرسودن mRNA ناشی از بی معنی استIntegrative Genomics Viewer - مجتمع ژنومیک بینندهUTR یا untranslated regions - منطقه ترجمه نشدهuntranslated region - منطقه غیر ترجمهsplice variant - نوع اتصالاتpolymerase chain reaction - واکنش زنجیره ای پلیمرازPCR - واکنش زنجیرهٔ پلیمراز
موضوعات مرتبط
علوم زیستی و بیوفناوری
بیوشیمی، ژنتیک و زیست شناسی مولکولی
ژنتیک
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
The adaptor-related protein (AP) complexes play important roles in cargo selection and vesicle formation, and hence in intracellular membrane trafficking. Five different AP complexes are currently known, each consisting of four subunits, known as adaptins. AP-2, the most thoroughly characterized of the five AP complexes, facilitates clathrin-mediated endocytosis. In this study, we describe the discovery and molecular cloning of seventy-seven novel alternatively spliced transcripts of the human AP2A1 gene, which encodes the αA adaptin of the AP-2 complex. For this purpose, we have used Next-Generation Sequencing (NGS), a powerful tool for studying alternative splicing. In brief, we subcultured fifty-five established human cell lines, originating from several distinct cancerous and normal tissues, extracted total RNA, and synthesized first-strand cDNA. Next, we used nested touchdown PCR to amplify the whole coding region of the AP2A1 transcripts of each cell line, mixed all PCR products, and proceeded to NGS library construction, template preparation, and semiconductor sequencing. Extensive bioinformatic analysis revealed thirteen novel splice junctions of previously annotated exons, as also verified via nested PCR with primers targeting these splice junctions. Moreover, consecutive nested PCRs led to the determination of the primary structure of seventy-seven novel AP2A1 transcripts, all of which were shown to comprise at least one premature translation termination codon, thus representing nonsense-mediated mRNA decay (NMD) candidates. NMD is a mechanism that cells use to control gene expression. Consequently, alterations in the levels of these potentially non-coding AP2A1 transcripts could lead to a decrease in the number of AP2A1 mRNA molecules, when needed. Undoubtedly, the exact role of these new APA1 splice variants merits elucidation.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Gene - Volume 678, 15 December 2018, Pages 55-64
Journal: Gene - Volume 678, 15 December 2018, Pages 55-64
نویسندگان
Panagiotis G. Adamopoulos, Christos K. Kontos, Marios A. Diamantopoulos, Andreas Scorilas,