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In Vitro Selection with a Site-Specifically Modified RNA Library Reveals the Binding Preferences of N6-Methyladenosine Reader Proteins.

Abstract：

:Epitranscriptomic RNA modifications can serve as recognition elements for the recruitment of effector proteins (i.e., "readers") to modified transcripts. While these interactions play an important role in mRNA regulation, there is a major gap in our understanding of the sequence determinants critical for the binding of readers to modified sequence motifs. Here, we develop a high-throughput platform, relying upon in vitro selection with a site-specifically modified random sequence RNA library and next-generation sequencing, to profile the binding specificity of RNA modification reader proteins. We apply our approach to interrogate the effect of sequence context on the interactions of YTH-domain proteins with N6-methyladenosine (m6A)-modified RNA. We find that while the in vitro binding preferences of YTHDC1 strongly overlap with the well-characterized DR(m6A)CH motif, the related YTH-domain proteins YTHDF1 and YTHDF2 can bind tightly to noncanonical m6A-containing sequences. Our results reveal the principles underlying substrate selection by m6A reader proteins and provide a powerful approach for investigating protein-modified RNA interactions in an unbiased manner.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Arguello AE,Leach RW,Kleiner RE

doi

10.1021/acs.biochem.9b00485

subject

Has Abstract

pub_date

2019-08-06 00:00:00

pages

3386-3395

issue

31

eissn

0006-2960

issn

1520-4995

journal_volume

58

pub_type

杂志文章

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