Universität zu Köln

Erkelenz, Steffen, Theiss, Stephan, Kaisers, Wolfgang, Ptok, Johannes, Walotka, Lara, Mueller, Lisa, Hillebrand, Frank, Brillen, Anna-Lena, Sladek, Michael and Schaal, Heiner (2018). Ranking noncanonical 5 ' splice site usage by genome-wide RNA-seq analysis and splicing reporter assays. Genome Res., 28 (12). S. 1826 - 1841. COLD SPRING HARBOR: COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT. ISSN 1549-5469

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Abstract

Most human pathogenic mutations in 5' splice sites affect the canonical GT in positions +1 and +2, leading to noncanonical dinucleotides. On the other hand, noncanonical dinucleotides are observed under physiological conditions in similar to 1% of all human 5'ss. It is therefore a challenging task to understand the pathogenic mutation mechanisms underlying the conditions under which noncanonical 5'ss are used. In this work, we systematically examined noncanonical 5' splice site selection, both experimentally using splicing competition reporters and by analyzing a large RNA-seq data set of 54 fibroblast samples from 27 subjects containing a total of 2.4 billion gapped reads covering 269,375 exon junctions. From both approaches, we consistently derived a noncanonical 5'ss usage ranking GC > TT > AT > GA > GG > CT. In our competition splicing reporter assay, noncanonical splicing was strictly dependent on the presence of upstream or downstream splicing regulatory elements (SREs), and changes in SREs could be compensated by variation of Ul snRNA complementarity in the competing 5'ss. In particular, we could confirm splicing at different positions (i.e., -1, +1, +5) of a splice site for all noncanonical dinucleotides weaker than GC. In our comprehensive RNA-seq data set analysis, noncanonical 5'ss were preferentially detected in weakly used exon junctions of highly expressed genes. Among high-confidence splice sites, they were 10-fold overrepresented in clusters with a neighboring, more frequently used 5'ss. Conversely, these more frequently used neighbors contained only the dinucleotides GT, GC, and TT, in accordance with the above ranking.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Erkelenz, Steffen UNSPECIFIED UNSPECIFIED UNSPECIFIED Theiss, Stephan UNSPECIFIED UNSPECIFIED UNSPECIFIED Kaisers, Wolfgang UNSPECIFIED UNSPECIFIED UNSPECIFIED Ptok, Johannes UNSPECIFIED UNSPECIFIED UNSPECIFIED Walotka, Lara UNSPECIFIED UNSPECIFIED UNSPECIFIED Mueller, Lisa UNSPECIFIED UNSPECIFIED UNSPECIFIED Hillebrand, Frank UNSPECIFIED UNSPECIFIED UNSPECIFIED Brillen, Anna-Lena UNSPECIFIED UNSPECIFIED UNSPECIFIED Sladek, Michael UNSPECIFIED UNSPECIFIED UNSPECIFIED Schaal, Heiner UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-163745
DOI:	10.1101/gr.235861.118
Journal or Publication Title:	Genome Res.
Volume:	28
Number:	12
Page Range:	S. 1826 - 1841
Date:	2018
Publisher:	COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
Place of Publication:	COLD SPRING HARBOR
ISSN:	1549-5469
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language U1 SNRNA; INTRONS REQUIRES; SR-PROTEINS; COMPLEX E; REVEALS; RECOGNITION; BINDING; SEQUENCES; MECHANISM; COMPLEMENTARITY Multiple languages Biochemistry & Molecular Biology; Biotechnology & Applied Microbiology; Genetics & Heredity Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/16374