MassIVE MSV000094855

Partial Public PXD052535

Using 14mer affinity pulldown to identify sRNA associating proteins in the bacterium Deinococcus radiodurans

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Description

We employed a 14-mer pulldown protocol that was originally developed for eukaryotic miRNA interacting RBPs. In this assay, the sRNA (Dsr2) and a scrambled RNA with the same length were in vitro transcribed with a 5 located, 14 nt long tag and subsequently incubated with D. radiodurans lysates. The resulting co-purified proteins were subjected to LC-MS/MS. Using this alternative approach (hereinafter referred to as 14 mer pull-down), we successfully enriched a total of 66 proteins in the 14 mer Dsr2 samples compared to the control sample. Except for several metabolic and hypothetical proteins, many of these proteins have potential nucleic acid binding functions were enriched, including three ribosomal proteins, two translation initiation factors, two TetR transcriptional regulators, five tRNA modification or processing proteins, DNA polymerase I, and five DNA repair proteins (e.g., RuvB, SbcD, and RecR). DR_2281 is one of the most enriched RBPs which contains a double stranded RNA binding motif (DSBM) but its function is unknown. In contrast, five other proteins (PNPase, RhlB, Ffh, DR_2281, KhpA and KhpB) were known as RBPs previously or contain known RNA binding domains. PNPase, RhlB, and Ffh have been revealed to play crucial roles in oxidative stress response in D. radiodurans by interacting with either oxidized RNA or Signal Recognition Particle RNA (Qpr6). PNPase also plays an indispensable role in paradoxically stabilizing sRNAs bound to other RBPs and promoting their function in gene regulation in E. coli (indicating a similar role of PNPase in D. radiodurans). [doi:10.25345/C5XS5JT8Z] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: small RNA, RNA-protein interaction, RNA binding protein, RNA regulation Deinococcus radiodurans

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Principal Investigators:
(in alphabetical order) 		Lydia M. Contreras, the University of Texas at Austin, United States
Submitting User: Runhuahan
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