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Translational specialization in pluripotency by RBPMS poises future lineage-decisions

ABSTRACT: Labeling active ribosome associated proteins by L-Azidohomoalanine (AHA)- pulldown. Humane embryonic stem cells were labeled via AHA in methionine-free medium for 40 min. AHA labeled nascent proteins together with ribosomes and associated proteins were isolated with beads. Samples were divided into ribosome plus associated protein fraction and nascent protein fraction by 8M UREA wash.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Prerana Wagle

LAB HEAD: Leo Kurian

PROVIDER: PXD032046 | Pride | 2023-10-26

REPOSITORIES: pride

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			Action	DRS
	20161107_AWH_ColID_245_projectID_256_DenizBartsch_Kurian_control1_40S.raw	Raw
	20161107_AWH_ColID_245_projectID_256_DenizBartsch_Kurian_control1_80S.raw	Raw
	20161107_AWH_ColID_245_projectID_256_DenizBartsch_Kurian_control1_Poly1.raw	Raw
	20161107_AWH_ColID_245_projectID_256_DenizBartsch_Kurian_control1_Poly2.raw	Raw
	20161107_AWH_ColID_245_projectID_256_DenizBartsch_Kurian_control2_40S.raw	Raw

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Translational specialization in pluripotency by RBPMS poises future lineage-decisions

Project description:Immunoprecipitation for RBPMS in human embryonic stem cells. Triplicates of sheep anti-rabbit beads coupled with rabbit anti-RBPMS antibody were compared with and triplicates sheep anti-rabbit beads, which served as IgG control. Bands above heavy IgG, between heavy IgG and light IgG, below IgG and both IgG bands combined were excised from a gel and labeled via dimethyl labeling, before LC-MS run to increase detection resolution.

2023-10-26 | PXD032040 | Pride

Translational specialization in pluripotency by RBPMS poises future lineage-decisions

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Project description:AATF is a predominantly nuclear protein that has been shown to be an apoptosis-inhibiting factor essential for both embryonic development and tumor growth. Several studies have shown a role of AATF in the modulation of pivotal cellular signal transduction pathways such as p53-, mTOR- and HIF-signaling. However, the exact molecular functions underlying its essential nature to cell proliferation and survival have remained elusive. Interestingly, several lines of evidence point towards a pivotal role of this protein in ribosome biogenesis and the maturation of ribosomal RNA. The function of AATF in this process is not clear, especially since the identity of the RNA-binding protein that links the AATF-containing protein complex (ANN) to rRNA precursors is unknown. In this study, we identify AATF in a screen for RNA-binding proteins. Importantly, AATF does not only bind to polyA-tailed RNA but CLIP-experiments reveal its association with ribosomal RNA. The RNA-binding domain of AATF is essential to maintain its positive effect on cellular rRNA levels. Furthermore, AATF binds to both mRNAs encoding for ribosome biogenesis factors as well as proteins involved with this process indicating AATF to be a central hub for the coordination of ribosome maturation. Consequently, our data for the first time identify the molecular basis to the role of AATF in ribosome biosynthesis and corroborate the link between tumorigenesis and ribosome biology.

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BrU-labeled nascent RNA sequencing and m6A-IP from BrU-labeled nascent RNA

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