Dataset Information

Variation in the Mitochondrial Proteome: Quantifying and Localizing Proteins Across Five Tissues in Mice. Mouse BXD, 8 Strains by 5 Metabolic Tissues (BAT, Liver, Heart, Quadriceps, Brain) by 2 fractions (Mitochondria and Total Cell)

ABSTRACT: We used 8 strains the BXD mouse genetic reference population, including parents C57BL/6J and DBA/2J, to study the proteome of the heart, whole brain, liver, quadriceps, and brown adipose tissue (BAT) across 8 different genotypes. For each sample, we examined the proteome of the mitochondria, as well as the whole cell fraction (including mitochondria). Later, we supplemented this study with an additional 19 strains of BXD mouse, with one strain overlapping, in the BAT for both whole cell and mitochondrial extract to examine tissue-specific hypotheses generated from the original set of 8 strains. All together, these data allowed us to (1) examine proteome expression of the mitochondria, (2) examine differences in proteome expression across tissue, (3) examine differences in proteome expression across strain, and (4) examine all three of the preceding factors simultaneously against one-another. This allowed us to uncover several proteins which are expressed in the mitochondria but are not reported as such in the literature, of which we validated a few by traditional staining techniques. Furthermore, we compare our proteome data against transcriptome data gathered in the same individual mice. We observe (1) that differences in expression of genes involved in oxidative phosphorylation are far more tissue-dependent at the protein level than the transcript level, and (2) and we observe that the phenotypic relationship between BAT and thermogenesis is far more evident at the protein expression data than it is from the transcript expression. Together, these data provide two conclusions: (1) next-gen mass spectrometry proteomics techniques can identify organelle localizations that were not evident from other techniques (perhaps particularly those proteins which are expressed both inside and outside the mitochondria); and (2) proteome and transcriptome data are complimentary, and proteome data may help us find and test new hypotheses even for physiological pathways which have been well studied at the genetic and transcriptomic levels.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Quadriceps, Heart, Brain, Brown Adipose Tissue, Liver

SUBMITTER: Evan Williams

LAB HEAD: Ruedi Aebersold

PROVIDER: PXD005044 | Pride | 2018-06-29

REPOSITORIES: Pride

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Publications

Quantifying and Localizing the Mitochondrial Proteome Across Five Tissues in A Mouse Population.

Williams Evan G EG Wu Yibo Y Wolski Witold W Kim Jun Yong JY Lan Jiayi J Hasan Moaraj M Halter Christian C Jha Pooja P Ryu Dongryeol D Auwerx Johan J Aebersold Ruedi R

Molecular & cellular proteomics : MCP 20180626 9

We have used SWATH mass spectrometry to quantify 3648 proteins across 76 proteomes collected from genetically diverse BXD mouse strains in two fractions (mitochondria and total cell) from five tissues: liver, quadriceps, heart, brain, and brown adipose (BAT). Across tissues, expression covariation between genes' proteins and transcripts-measured in the same individuals-broadly aligned. Covariation was however far stronger in certain subsets than others: only 8% of transcripts in the lowest expre ...[more]

PMID: 29945935

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Project description:Thoracic perivascular adipose tissue (PVAT) is a unique adipose depot that likely influences vascular function and susceptibility to pathogenesis in obesity and metabolic syndrome. Surprisingly, PVAT has been reported to share characteristics of both brown and white adipose, but a detailed direct comparison to interscapular brown adipose tissue (BAT) has not been performed. Here we show by full genome DNA microarray analysis that global gene expression profiles of PVAT are virtually identical to BAT, with equally high expression of Ucp-1, Cidea and other genes known to be uniquely or very highly expressed in BAT. PVAT and BAT also displayed nearly identical phenotypes upon immunohistochemical analysis, and electron microscopy confirmed that PVAT contained multilocular lipid droplets and abundant mitochondria. Compared to white adipose tissue (WAT), PVAT and BAT from C57BL/6 mice fed a high fat diet for 13 weeks had markedly lower expression of immune cell-enriched mRNAs, suggesting resistance to obesity-induced inflammation. Indeed, staining of BAT and PVAT for macrophage markers (F4/80, CD68) in obese mice showed virtually no macrophage infiltration, and FACS analysis of BAT confirmed the presence of very few CD11b+/CD11c+ macrophages in BAT (1.0%) in comparison to WAT (31%). In summary, murine PVAT from the thoracic aorta is virtually identical to interscapular BAT, is resistant to diet-induced macrophage infiltration, and thus may play an important role in protecting the vascular bed from thermal and inflammatory stress. 8-week-old male C57BL6/J mice were fed a normal (ND) or high fat diet (HFD) (Research Diets 12451, 45 kcal% fat) for 13 weeks. Mice were then euthanized and four different adipose depots were harvested for RNA analysis: perivascular fat from the lesser curvature of the aortic arch (PVAT), interscapular brown adipose (BAT), inguinal adipose tissue (SAT), and epididymal adipose tissue (VAT). 250 ng total RNA pooled from two mice was used for cDNA synthesis; 3 biological replicates per tissue and diet were performed for a total of 24 hybridizations.

Project description:Rodents are commonly housed below thermoneutrality (~20°C) 1. Under these conditions there is a substantial effect on rodent physiology including the hyperactivation of brown (BAT) and beige adipose tissue 2. Here, we raised animals from weaning, on an obesogenic diet at thermoneutrality (28°C) to closer mimic human physiology and determine the impact of a) moderate cold exposure (i.e. 20°C, a temperature reduction of ~8°C) or b) treatment with YM-178, a highly-selective, clinically used β3-adrenoreceptor agonist on classical BAT or subcutaneous inguinal (IWAT) beige depots. Under these conditions, uncoupling protein 1 mRNA was undetectable in IWAT in all groups. Maintenance at 20°C drove weight gain and a 125% increase in subcutaneous fat, an effect not seen with YM-178 administration thus suggesting a direct effect of ambient temperature in promoting weight gain and adiposity in obese rats. Using exploratory adipose tissue proteomics we reveal novel processes and pathways associated with cold-induced weight gain in BAT (i.e. histone deacetylation and glycosphingolipid biosynthesis) and IWAT (i.e. NAD+ binding and retinol metabolism). Conversely, YM-178 had minimal metabolic-related effects on BAT and drove a pro-inflammatory phenotype in IWAT. Exercise training elicits diverse effects on brown (BAT) and white adipose tissue (WAT) physiology in rodents housed below their thermoneutral zone (i.e. 28-32°C). In these conditions, BAT is chronically hyperactive and, unlike human residence, closer to thermoneutrality. Therefore, we set out to determine the effects of exercise training in obese animals at 28°C (i.e. thermoneutrality) on BAT and WAT in its basal (i.e. inactive) state. Sprague-Dawley rats (n=12) were housed at thermoneutrality from 3 weeks of age and fed a high-fat diet. At 12 weeks of age half these animals were randomised to 4-weeks of swim-training (1 hour/day, 5 days per week). Following a metabolic assessment interscapular and perivascular BAT and inguinal (I)WAT were taken for analysis of thermogenic genes and the proteome. Exercise attenuated weight gain but did not affect total fat mass or thermogenic gene expression. Proteomics revealed an impact of exercise training on2-oxoglutarate metabolic process, mitochondrial respiratory chain complex IV, carbon metabolism and oxidative phosphorylation. This was accompanied by an upregulation of multiple proteins involved in skeletal muscle physiology suggesting an adipocyte to myocyte switch in BAT. UCP1 mRNA was undetectable in IWAT with proteomics highlighting changes to DNA binding, the positive regulation of apoptosis, HIF-1 signalling and cytokine-cytokine receptor interaction.

Dataset Information

Variation in the Mitochondrial Proteome: Quantifying and Localizing Proteins Across Five Tissues in Mice. Mouse BXD, 8 Strains by 5 Metabolic Tissues (BAT, Liver, Heart, Quadriceps, Brain) by 2 fractions (Mitochondria and Total Cell)

Publications

Quantifying and Localizing the Mitochondrial Proteome Across Five Tissues in A Mouse Population.

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