PARN Knockdown in Cell Lines Results in Differential and Cell-Specific Alterations in the Expression of Cancer-Associated mRNAs.

Nishith Babu,Srividya Arjuna,Anirban Chakraborty,Sandra Nazareth,Dechamma Nanjappa

doi:10.31557/apjcp.2022.23.1.207

Abstract

Ribonucleases (RNases) is the collective term used for the group of enzymes that are involved in mRNA degradation. The shortening of the poly (A) tail through deadenylation is the preferred mechanism of degradation of most eukaryotic mRNAs and poly (A)-specific ribonuclease (PARN) is the most important player in deadenylation. Besides its primarily role in mRNA stability, PARN is also involved in several non-conventional functions. It is conceivable that a decreased RNase activity can alter the stability of cancer-associated mRNAs and this alteration may be differential in cells of different origin. Methods:The effects of siRNA-mediated knockdown of PARN on the post-transcriptional expression of 16 oncogenes and 18 tumor suppressor genes in cells derived from different lineages (NCI-H460 and NCI-H522; lung cancer) and (HEK-293; kidney) were investigated. Further, the effects of PARN depletion on proliferation and death of the lung cancer cells were investigated. Results: Quantitative real time PCR analysis revealed an cell-specific alteration in the expression of the target onco and tumor suppressor genes upon PARN depletion, differently, for cells derived from different lineages. The tumor suppressor genes showed a consistent pattern of down regulation upon PARN depletion in all the three cell types tested. In contrast, the expression of oncogenes was not consistent; while some oncogenes showed overexpression in HEK 293 cells, the majority of them were downregulated in the lung cancer cells. Further, PARN depletion did not alter the proliferation of lung cancer cells, which was in contrast to previous reports. Conclusion:The results of this study reveal that PARN deficiency leads to an altered stability of cancer-associated mRNA, distinctly, in cells of different lineages. Despite previous reports suggesting a potential therapeutic role of PARN in cancer, our results suggest that PARN may not be an important biomarker, particularly in lung cancer.

Highlights

Almost all mRNAs contain a poly (A) tail at its 3’end, a stretch of approximately 25-200 adenosine residues added post-transcriptionally, which becomes crucial for its processing, export into the cytoplasm and translation (Dehlin et al, 2000; Garneau et al, 2007; Lee et al, 2012)
In HEK-293, down regulation of poly (A)-specific ribonuclease (PARN) mRNA was seen for both siRNAs (Figure 1A)
The down-regulation was significant for PARN siRNA2 compared to PARN siRNA1 and siRNA2 was used for further experiments

Summary

Introduction

Almost all mRNAs contain a poly (A) tail at its 3’end, a stretch of approximately 25-200 adenosine residues added post-transcriptionally, which becomes crucial for its processing, export into the cytoplasm and translation (Dehlin et al, 2000; Garneau et al, 2007; Lee et al, 2012). Among the many RNases, the deadenylases are most crucial for deadenylation by virtue of their high affinity to the poly (A) tail of the mRNA (Dhanraj et al, 2015). Poly(A) specific ribonuclease (PARN) is believed to be the key deadenylase in regulating mRNA turnover rates in mammalian cells (Lee et al, 2012). Org), PARN mRNA displays low tissue specificity and is expressed in all tissues. PARN protein shows cytoplasmic and nuclear expression in most tissues

Methods

Results

Conclusion