A quantitative spatial proteomics analysis of proteome turnover in human cells.

Measuring the properties of endogenous cell proteins, such as expression level, subcellular localization, and turnover rates, on a whole proteome level remains a major challenge in the postgenome era. Quantitative methods for measuring mRNA expression do not reliably predict corresponding protein levels and provide little or no information on other protein properties. Here we describe a combined pulse-labeling, spatial proteomics and data analysis strategy to characterize the expression, localization, synthesis, degradation, and turnover rates of endogenously expressed, untagged human proteins in different subcellular compartments.

In vivo study in Trypanosoma brucei links mitochondrial transfer RNA import to mitochondrial protein import.

Trypanosoma brucei imports all mitochondrial transfer RNAs (tRNAs) from the cytosol. By using cell lines that allow independent tetracycline-inducible RNA interference and isopropyl-β-D-thiogalactopyranoside-inducible expression of a tagged tRNA, we show that ablation of Tim17 and mitochondrial heat-shock protein 70, components of the inner-membrane protein translocation machinery, strongly inhibits import of newly synthesized tRNAs. These findings, together with previous results in yeast and plants, suggest that the requirement for mitochondrial protein-import factors might be a conserved feature of mitochondrial tRNA import in all systems.

Dual targeting of a tRNAAsp requires two different aspartyl-tRNA synthetases in Trypanosoma brucei.

The mitochondrion of the parasitic protozoon Trypanosoma brucei does not encode any tRNAs. This deficiency is compensated for by partial import of nearly all of its cytosolic tRNAs. Most trypanosomal aminoacyl-tRNA synthetases are encoded by single copy genes, suggesting the use of the same enzyme in the cytosol and in the mitochondrion.

The single mitochondrial porin of Trypanosoma brucei is the main metabolite transporter in the outer mitochondrial membrane.

All mitochondria have integral outer membrane proteins with beta-barrel structures including the conserved metabolite transporter VDAC (voltage dependent anion channel) and the conserved protein import channel Tom40. Bioinformatic searches of the Trypanosoma brucei genome for either VDAC or Tom40 identified a single open reading frame, with sequence analysis suggesting that VDACs and Tom40s are ancestrally related and should be grouped into the same protein family: the mitochondrial porins. The single T.

Simultaneous cell morphometry and refractive index measurement with dual-wavelength digital holographic microscopy and dye-enhanced dispersion of perfusion medium.

Digital holographic microscopy (DHM) allows optical-path-difference (OPD) measurements with nanometric accuracy. OPD induced by transparent cells depends on both the refractive index (RI) of cells and their morphology. This Letter presents a dual-wavelength DHM that allows us to separately measure both the RI and the cellular thickness by exploiting an enhanced dispersion of the perfusion medium achieved by the utilization of an extracellular dye.