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Series GSE230431 Query DataSets for GSE230431
Status Public on Jan 30, 2024
Title GPATCH4 regulates rRNA and snRNA 2’-O-methylation in both DHX15-dependent and -independent manners [CRAC]
Organism Homo sapiens
Experiment type Other
Summary RNA helicases are critical regulators of gene expression and therefore strict regulation of their activities is essential to limit promiscuity and ensure target specify. While some RNA helicases appear dedicated for particular substrates others, such as DHX15, are multifunctional, contributing to more than one gene expression process. Protein cofactors play key roles both in modulating RNA helicase activities and also directing them to appropriate substrate RNAs. The largest family of RNA helicase cofactors, with more than 20 members expressed in human cells, are the G-patch proteins, characterised by a glycine-rich domain via which they associate with specific DEAH/RHA helicases. However, the cognate RNA helicases and cellular functions of numerous human G-patch proteins still remain elusive. Here, we reveal that the G-patch protein GPATCH4 is a cofactor of DHX15 that stimulates its ATPase activity. We show that GPATCH4 interacts with DHX15 in the nucleolus via residues in its G-patch domain and co-migrates with pre-ribosomal particles, wherein loss of DHX15 reduces pre-ribosomal co-migration of GPATCH4. Using a combination of high-throughput sequencing approaches (ChIP, CRAC and small RNA-seq), we discovered that GPATCH4 crosslinks to the ribosomal DNA locus and precursor ribosomal RNAs as well as binding to small nucleolar- and small Cajal body-associated- RNAs that guide rRNA and snRNA modifications. In line with this, using RiboMeth-seq analysis to globally analyse rRNA and snRNA 2’-O-methylation revealed that knockout of GPATCH4 impairs methylation at various sites. Utilizing targeted 2’-O-methylation-sensitive RNase H-based cleavage assays, we demonstrated that the regulation of 2’-O-methylation by GPATCH4 is both dependent on, and independent of, its interaction with DHX15. Intriguingly, the ATPase activity of DHX15 is necessary for efficient methylation of DHX15-dependent sites, suggesting a function of DHX15 in regulating snoRNA-guided 2’-O-methylation of rRNA that requires activation by GPATCH4. Overall, our findings not only extend knowledge on RNA helicase regulation by G-patch proteins but also provide important new insights into how the installation of rRNA and snRNA modifications, which are essential for ribosome assembly/function and pre-mRNA splicing, is regulated.
 
Overall design Flp-In T-Rex HEK293 cell lines expressing GPATCH4-His6-2xFLAG (FLAG) or the FLAG tag alone were crosslinked using UV and analysed by CRAC.
 
Contributor(s) Kanwal N, Krogh N, Memet I, Lemus-Diaz N, Thome C, Welp L, Mizi A, Hackert P, Papantonis A, Urlaub H, Nielsen H, Bohnsack K, Bohnsack M
Citation(s) 38113271
Submission date Apr 24, 2023
Last update date Mar 20, 2024
Contact name Katherine Bohnsack
E-mail(s) katherine.bohnsack@med.uni-goettingen.de
Organization name University Medical Centre Göttingen
Street address Humboldtallee 23
City Göttingen
ZIP/Postal code 37073
Country Germany
 
Platforms (1)
GPL20301 Illumina HiSeq 4000 (Homo sapiens)
Samples (4)
GSM7221980 His-Flag_A
GSM7221981 His-Flag_B
GSM7221982 GPATCH4-His-Flag_A
This SubSeries is part of SuperSeries:
GSE230433 GPATCH4 regulates rRNA and snRNA 2'-O-methylation in both DHX15-dependent and -independent manners
Relations
BioProject PRJNA961127

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Supplementary file Size Download File type/resource
GSE230431_RAW.tar 9.3 Mb (http)(custom) TAR (of BEDGRAPH)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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