MilliporeSigma
  • Most human proteins made in both nucleus and cytoplasm turn over within minutes.

Most human proteins made in both nucleus and cytoplasm turn over within minutes.

PloS one (2014-06-10)
Sabyasachi Baboo, Bhaskar Bhushan, Haibo Jiang, Chris R M Grovenor, Philippe Pierre, Benjamin G Davis, Peter R Cook
ABSTRACT

In bacteria, protein synthesis can be coupled to transcription, but in eukaryotes it is believed to occur solely in the cytoplasm. Using pulses as short as 5 s, we find that three analogues--L-azidohomoalanine, puromycin (detected after attaching fluors using 'click' chemistry or immuno-labeling), and amino acids tagged with 'heavy' 15N and 13C (detected using secondary ion mass spectrometry)--are incorporated into the nucleus and cytoplasm in a process sensitive to translational inhibitors. The nuclear incorporation represents a significant fraction of the total, and labels in both compartments have half-lives of less than a minute; results are consistent with most newly-made peptides being destroyed soon after they are made. As nascent RNA bearing a premature termination codon (detected by fluorescence in situ hybridization) is also eliminated by a mechanism sensitive to a translational inhibitor, the nuclear turnover of peptides is probably a by-product of proof-reading the RNA for stop codons (a process known as nonsense-mediated decay). We speculate that the apparently-wasteful turnover of this previously-hidden ('dark-matter') world of peptide is involved in regulating protein production.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Hydrofluoric acid, 48 wt. % in H2O, ≥99.99% trace metals basis
Millipore
Cycloheximide solution, 0.1%, suitable for microbiology
Cystine, European Pharmacopoeia (EP) Reference Standard
USP
Cystine, United States Pharmacopeia (USP) Reference Standard
Sigma-Aldrich
Hydrofluoric acid, ACS reagent, 48%
Sigma-Aldrich
Cycloheximide, Biotechnology Performance Certified
Sigma-Aldrich
Cycloheximide, ≥90% (HPLC)
Sigma-Aldrich
Tetracycline, 98.0-102.0% (HPLC)
Sigma-Aldrich
Cycloheximide, from microbial, ≥94% (TLC)
Sigma-Aldrich
Tetracycline, 98.0-102.0% (HPLC)
Sigma-Aldrich
Cycloheximide solution, Ready-Made Solution, microbial, 100 mg/mL in DMSO, Suitable for cell culture
Supelco
Cycloheximide, PESTANAL®, analytical standard
Supelco
4-tert-Octylphenol monoethoxylate solution, 10 μg/mL in acetone, analytical standard
Sigma-Aldrich
DAPI, for nucleic acid staining
Sigma-Aldrich
DL-Cysteine, technical grade
Sigma-Aldrich
Z-Leu-Leu-Leu-al, ≥90% (HPLC)