Comprehensive evaluation of RNAlater effect on proteome and phosphoproteome

ABSTRACT: Tumors such as pancreatic cancer contain a variety of enzymes including DNases, RNases and proteases which may lead to autolysis of DNA, RNA and proteins during sample processing. In general, RNAlater can be used to keep nucleic acids intact since it contains high concentrations of quaternary ammonium sulfates which denature those enzymes. Although a few studies were carried out to find effect of RNAlater mainly on DNA and RNA, it is largely unknown whether RNAlater affect both proteome and phosphoproteome. Thus, we carried out a systematic and comprehensive analysis of the RNAlater effect on the proteome and phosphoproteome using high-resolution mass spectrometry. Pancreatic tumor tissues from three patients were pulverized using Covaris CP02 Cryoprep device and incubated in RNAlater at 4 °C for 24 hours. We used the 6-plex TMT to measure quantitative information of proteome and phosphoproteome. Peptides fractionated by mid pH reverse phase liquid chromatography were divided into two parts: a portion (~8%) of each mRP fraction was used to profile the global proteome and the rest (~92%) was to survey phosphorylation enriched by IMAC–based approach. As a result, the global profiling identified 186,941 unmodified peptides of 9,152 protein groups, 18,705 phosphopeptides (15,842 phosphosites). When analyzing the unbiased proteomics and phosphoproteomics data, we observed no significant quantitative changes in both proteins and phosphorylation, if any, induced by RNAlater. Therefore, this result confirms that stored tissues in RNAlater do not significantly affect the proteome and phosphoproteome of pancreatic cancer tumors.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Pancreas

DISEASE(S): Pancreatic Cancer

SUBMITTER: Min-Sik Kim  

LAB HEAD: Min-Sik Kim

PROVIDER: PXD010710 | Pride | 2019-05-06

REPOSITORIES: Pride