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Identification of a novel intracellular function of the secreted ribonuclease RNASE1 in inhibiting gene expression

ABSTRACT: Identification of a novel intracellular function of the secreted ribonuclease RNASE1 in inhibiting gene expression

PROVIDER: PRJNA1218632 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR32307587_1.fastq.gz	Fastqsanger.gz
	SRR32307587_2.fastq.gz	Fastqsanger.gz
	SRR32307588_1.fastq.gz	Fastqsanger.gz
	SRR32307588_2.fastq.gz	Fastqsanger.gz
	SRR32307589_1.fastq.gz	Fastqsanger.gz

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Identification of a novel intracellular function of the secreted ribonuclease RNASE1 in inhibiting gene expression

Project description:RNASE1 is a ribonuclease secreted by cells and degrades extracellular RNAs. Here, we unexpectedly found that endogenous RNASE1, in addition to being secreted, is predominantly localized to the nucleus and primarily functions to inhibit gene expression in human colorectal cancer (CRC) cells. In CRC cells, RNASE1 expression is highly cell type-specific and is restricted to well-differentiated cells where its transcription is activated by the pioneer transcription factor FOXA1. Using CRISPR interference (CRISPRi) utilizing three independent sgRNAs targeting the RNASE1 locus followed by RNA-seq, we found that upon depletion of RNASE1, most of the differentially expressed RNAs are modestly but significantly upregulated suggesting that RNASE1 may predominantly function to inhibit gene expression. In CRC patients, RNASE1 is significantly downregulated and high RNASE1 expression is associated with better patient survival indicating a potential tumor suppressive function. Consistent with this, RNASE1 depletion results in increased proliferation and clonogenicity indicating that RNASE1 inhibits the growth of CRC cells. Finally, a promising RNASE1 target amongst the most significantly upregulated mRNAs upon RNASE1 depletion is DKK1 (Dickkopf inhibitor 1) which is upregulated in CRC and negatively regulated by RNASE1. Collectively, these data uncover an intracellular function of RNASE1 in inhibiting gene expression and CRC cell proliferation.

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Ribonuclease Selection for Ribosome Profiling

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Project description:Ribosome profiling has emerged as a powerful method to assess global gene translation, but methodological and analytical challenges often lead to inconsistencies across labs and model organisms. A critical issue in ribosome profiling is nuclease treatment of ribosome-mRNA complexes, as it is important to ensure both stability of ribosomal particles and complete conversion of polysomes to monosomes. We performed comparative ribosome profiling in yeast and mice with various ribonucleases including I, A, S7 and T1, characterized their cutting preferences, trinucleotide periodicity patterns, and coverage similarities across coding sequences, and showed that they yield comparable estimations of gene expression when ribosome integrity is not compromised. However, ribosome coverage patterns of individual transcripts had little in common between the ribonucleases. We further examined their potency at converting polysomes to monosomes across other commonly used model organisms, including bacteria, nematodes and fruit flies. In some cases, ribonuclease treatment completely degraded ribosome populations. Ribonuclease T1 was the only enzyme that preserved ribosomal integrity while thoroughly converting polysomes to monosomes in all examined species. This study provides a guide for ribonuclease selection in ribosome profiling experiments across most common model systems

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