SummaryDiurnal oscillations of gene expression controlled by the circadian clock and its connected feeding rhythm enable organisms to coordinate their physiologies with daily environmental cycles. While available techniques yielded crucial insights into regulation at the transcriptional level, much less is known about temporally controlled functions within the nucleus and their regulation at the protein level. Here, we quantified the temporal nuclear accumulation of proteins and phosphoproteins from mouse liver by SILAC proteomics. We identified around 5,000 nuclear proteins, over 500 of which showed a diurnal accumulation. Parallel analysis of the nuclear phosphoproteome enabled the inference of the temporal activity of kinases accounting for rhythmic phosphorylation. Many identified rhythmic proteins were parts of nuclear complexes involved in transcriptional regulation, ribosome biogenesis, DNA repair, and the cell cycle and its potentially associated diurnal rhythm of hepatocyte polyploidy. Taken together, these findings provide unprecedented insights into the diurnal regulatory landscape of the mouse liver nucleus.

This study encompasses a collection of experiences with regard to numerous matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) sample preparation techniques in terms of their suitability for di †erent peptide and protein analytes. Variants of both established and new sample preparation techniques for the MALDI-MS analysis of peptides and proteins are described. The importance of matrix selection, matrix and analyte concentration, pH adjustment, crystallization conditions and the use of additives is evaluated. The tolerance of the di †erent sample preparations towards salts, bu †ers, synthetic polymers, detergents, denaturants and other contaminants, and also the inÑuence of the preparation methods on undesired amino acid side-chain oxidation, are investigated. Moreover, the performance of on-target tryptic digestion and on-target disulÐde reduction is shown and a microscale puriÐcation procedure is described.

This study encompasses a collection of experiences with regard to numerous matrix‐assisted laser desorption/ionization mass spectrometry (MALDI‐MS) sample preparation techniques in terms of their suitability for different peptide and protein analytes. Variants of both established and new sample preparation techniques for the MALDI‐MS analysis of peptides and proteins are described. The importance of matrix selection, matrix and analyte concentration, pH adjustment, crystallization conditions and the use of additives is evaluated. The tolerance of the different sample preparations towards salts, buffers, synthetic polymers, detergents, denaturants and other contaminants, and also the influence of the preparation methods on undesired amino acid side‐chain oxidation, are investigated. Moreover, the performance of on‐target tryptic digestion and on‐target disulfide reduction is shown and a microscale purification procedure is described. According to this study, there is no universally applicable sample preparation for a broad variety of analytes. Rather, it is necessary to specifically adapt the sample preparation to the analyte properties. © 1997 John Wiley & Sons, Ltd.

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