A commentary on the interpretation of in vitro biochemical measures of brown adipose tissue thermogenesis

1989 ◽  
Vol 67 (8) ◽  
pp. 811-819 ◽  
Author(s):  
Paul Trayhurn ◽  
Rachel E. Milner
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Binding Sites ◽  
Protein Concentration ◽  
Uncoupling Protein ◽  
Proton Conductance ◽  
Binding Studies ◽  
Brown Adipose ◽  
Brown Adipose Tissue Thermogenesis

In this article we comment on the various in vitro biochemical measurements employed to assess the thermogenic activity and capacity of brown adipose tissue. The meaning and significance of changes in tissue weight, protein content, cell number, and mitochondrial mass are each summarized. In addition, various indices of the proton conductance pathway – mitochondrial swelling, proton conductance, uncoupling protein concentration, and GDP binding studies – are discussed. The issue of unmasking and masking of GDP binding sites is reviewed; recent reports have clearly demonstrated unmasking and masking, and it is concluded that GDP binding studies are an index of the activity of uncoupling protein, rather than a measure of its concentration. It is suggested that tissue mass, mitochondrial content, mitochondrial GDP binding, and uncoupling protein concentration represent core measurements for the biochemical assessment of the thermogenic activity and capacity of brown adipose tissue. Auxiliary measurements include Scatchard analysis of GDP binding data to distinguish changes in the number of binding sites from potential changes in Kd, and mitochondrial swelling studies, as an additional index of proton permeability. The distinction between thermogenic activity (GDP binding, proton permeability) and capacity (uncoupling protein content), both on a per unit of mitochondrial protein and per tissue basis, is emphasized.Key words: brown adipose tissue, thermogenesis, uncoupling protein, mitochondria.

scholarly journals Acute effects of a β-adrenoceptor agonist (BRL 26830A) on rat brown-adipose-tissue mitochondria. Increased GDP binding and GDP-sensitive proton conductance without changes in the concentration of uncoupling protein

1988 ◽  
Vol 249 (3) ◽  
pp. 759-763 ◽  
Author(s):  
R E Milner ◽  
S Wilson ◽  
J R Arch ◽  
P Trayhurn
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Binding Sites ◽  
Uncoupling Protein ◽  
Beta Agonist ◽  
Proton Conductance ◽  
Binding Data ◽  
Adrenoceptor Agonist ◽  
Brown Adipose ◽  
Acute Increase

GDP binding, proton conductance and the specific concentration of uncoupling protein were measured in brown-adipose-tissue mitochondria of rats treated acutely with the novel beta-agonist, BRL 26830A. At 1 h after dosing with BRL 26830A, mitochondrial GDP binding was increased more than 2-fold. The increase in binding resulted from an increase in the number of binding sites. An iterative analysis of Scatchard binding data suggested that there is only one high-affinity GDP-binding site (Kd 0.3 microM) in brown-adipose-tissue mitochondria. The acute increase in GDP binding produced by treatment with BRL 26830A occurred without any alteration in the specific mitochondrial concentration of uncoupling protein, as determined by radioimmunoassay. Treatment with the beta-agonist did, however, lead to a small increase in the GDP-sensitive component of mitochondrial proton conductance. These results indicate that GDP-binding sites on uncoupling protein can be rapidly unmasked after treatment with a brown-fat-specific beta-agonist, and that the increase in binding reflects an increase in the activity of the mitochondrial proton-conductance pathway.

Brown fat thermogenesis during hibernation and arousal in Richardson's ground squirrel

1989 ◽  
Vol 256 (1) ◽  
pp. R42-R48 ◽  
Author(s):  
R. E. Milner ◽  
L. C. Wang ◽  
P. Trayhurn
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Cold Acclimation ◽  
Binding Sites ◽  
Ground Squirrel ◽  
Protein Concentration ◽  
Uncoupling Protein ◽  
Ground Squirrels ◽  
Mitochondrial Content ◽  
Brown Adipose

The thermogenic activity [mitochondrial guanosine 5'-diphosphate (GDP) binding] and capacity (uncoupling protein concentration, cytochrome oxidase activity) of brown adipose tissue have been investigated at different phases of the seasonally linked hibernation cycle in Richardson's ground squirrel. The amount of axillary brown adipose tissue and the total mitochondrial content of the tissue were substantially greater in hibernating squirrels than in squirrels caught posthibernation in April or May; cold acclimation induced qualitatively similar differences. The specific mitochondrial concentration of uncoupling protein was high under all conditions (compared with other species), differing little between hibernating, posthibernating, and cold-acclimated squirrels. The thermogenic capacity of brown adipose tissue in Richardson's ground squirrels is therefore modulated almost exclusively by changes in the mitochondrial content of the tissue. Mitochondrial GDP binding was increased on cold acclimation, but similar binding levels were observed in hibernating and posthibernation (May) animals. GDP binding and the GDP-sensitive component of acetate-induced mitochondrial swelling were increased during the early stages of arousal from hibernation. These changes, which indicate an activation of the thermogenic proton conductance pathway in arousal, occurred without an alteration in the specific mitochondrial concentration of uncoupling protein. Increased GDP binding during arousal is clearly due to the unmasking of binding sites, reflecting an acute activation of preexisting uncoupling protein.

scholarly journals TAF7L modulates brown adipose tissue formation

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Haiying Zhou ◽  
Bo Wan ◽  
Ivan Grubisic ◽  
Tommy Kaplan ◽  
Robert Tjian
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Core Promoter ◽  
Uncoupling Protein ◽  
Dna Looping ◽  
Chromatin Interactions ◽  
Brown Adipose ◽  
Important Regulator

Brown adipose tissue (BAT) plays an essential role in metabolic homeostasis by dissipating energy via thermogenesis through uncoupling protein 1 (UCP1). Previously, we reported that the TATA-binding protein associated factor 7L (TAF7L) is an important regulator of white adipose tissue (WAT) differentiation. In this study, we show that TAF7L also serves as a molecular switch between brown fat and muscle lineages in vivo and in vitro. In adipose tissue, TAF7L-containing TFIID complexes associate with PPARγ to mediate DNA looping between distal enhancers and core promoter elements. Our findings suggest that the presence of the tissue-specific TAF7L subunit in TFIID functions to promote long-range chromatin interactions during BAT lineage specification.

Rapid changes in number of GDP binding sites on brown adipose tissue mitochondria

1986 ◽  
Vol 251 (2) ◽  
pp. E192-E195
Author(s):  
A. G. Swick ◽  
R. W. Swick
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Binding Sites ◽  
Uncoupling Protein ◽  
Membrane Integrity ◽  
Binding Activity ◽  
Bat Mitochondria ◽  
Brown Adipose ◽  
Rapid Changes ◽  
Binding To Mitochondria

GDP binding to brown adipose tissue (BAT) mitochondria increased more than twofold in 20 min when rats were moved from 27 to 4 degrees C. When animals housed at 4 degrees C for 2 h were returned to 27 degrees C, GDP binding decreased sharply in 20 min and returned to control levels in 2 h. These results are consistent with a rapid unmasking and remasking of GDP binding sites. GDP binding to mitochondria from warm and acutely cold treated rats was not modified by prior swelling, by freeze-thawing, nor by sonication of the mitochondria before assay. GDP-inhibitable proton conductance, as measured by passive swelling, was unaffected by this brief exposure to cold but more than doubled in rats kept at 4 degrees C for 10 days. We hypothesize that the rate of GDP-inhibitable swelling may be a reflection of uncoupling protein concentration in the BAT mitochondria, whereas physiological thermogenic activity is more appropriately indicated by GDP binding. The alterations in binding activity appear not to be due to changes in the mitochondrial membrane integrity.

scholarly journals Regulation of GDP binding and uncoupling-protein concentration in brown-adipose-tissue mitochondria. The effects of cold-acclimation, warm-reacclimation and noradrenaline

1988 ◽  
Vol 249 (2) ◽  
pp. 451-457 ◽  
Author(s):  
T Peachey ◽  
R R French ◽  
D A York
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Protein Concentration ◽  
Uncoupling Protein ◽  
Potassium Acetate ◽  
Zucker Rats ◽  
Mitochondrial Uncoupling ◽  
Mitochondrial Uncoupling Protein ◽  
Acute Response ◽  
Brown Adipose

We have used a specific immunoassay for uncoupling protein and [3H]GDP binding to study the acute and chronic responses of brown-adipose-tissue (BAT) mitochondria of warm-acclimated rats to housing at 4 degrees C and cold-acclimated rats to housing at 27 degrees C. These studies have shown the following. (1) In the cold-exposed rat the increase in mitochondrial uncoupling-protein concentration parallels the increase in GDP binding from 1 day to 5 days, but that acutely (initial 4 h) the increase in GDP binding is not associated with any change in uncoupling-protein concentration. 2. In the cold-acclimated rat rehoused at 27 degrees C, GDP binding fell by over 50% in the first 2 days, without any change in uncoupling-protein concentrations. 3. Noradrenaline acutely (30 min) increased BAT mitochondrial GDP binding of lean and obese Zucker rats, without any change in uncoupling-protein concentrations. 4. The increases in GDP binding in cold-exposed rats were associated with increases in the rate of swelling of mitochondria in the presence of valinomycin and potassium acetate. The evidence supports the hypothesis that the acute response of the rat to changes in environmental temperature are associated with unmasking or remasking of uncoupling protein, whereas chronically changes in uncoupling-protein concentration predominate.

scholarly journals Evidence for masking of brown adipose tissue mitochondrial GDP-binding sites in response to fasting in rats made obese by dietary manipulation. Effects of reversion to standard diet

1991 ◽  
Vol 279 (2) ◽  
pp. 575-579 ◽  
Author(s):  
P Puigserver ◽  
I Lladó ◽  
A Palou ◽  
M Gianotti
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Binding Sites ◽  
Uncoupling Protein ◽  
Cafeteria Diet ◽  
Standard Diet ◽  
Dietary Manipulation ◽  
Bat Mitochondria ◽  
Brown Adipose ◽  
Obese Rats

A specific immunoassay of uncoupling protein (UCP) and measurement of GDP binding were used to study the chronic responses of brown adipose tissue (BAT) mitochondria from rats made obese by dietary means (cafeteria rats) and from obese rats subsequently fed a standard diet (post-cafeteria rats). We studied the response to fasting in order to assess the masking/unmasking responses in these groups. These studies have shown the following. (1) In the obese rats (cafeteria and post-cafeteria) the chronic increase in mitochondrial UCP concentration compared with controls parallels the increase in GDP binding. (2) In 24 h-fasted control rats the decrease in GDP binding is associated with a change in UCP concentration, but in fasting cafeteria and post-cafeteria obese rats the decrease in GDP binding is not associated with any change in UCP concentration. (3) Post-cafeteria obese rats showed increased GDP binding and higher UCP concentrations than the controls, but these values were less than in cafeteria obese rats. (4) Control rats at 8 months old showed greater GDP binding and had a higher UCP concentration than 11-month-old control rats. (5) The responses of GDP binding and UCP concentration to fasting in post-cafeteria obese rats were similar to those in cafeteria obese rats, suggesting that such abbreviations are related to the obese status itself rather than to the composition of the cafeteria diet. The evidence supports the hypothesis that the response of the cafeteria and post-cafeteria obese rats to fasting is associated with a masking of UCP, whereas with chronic manipulation of diet changes in UCP concentration predominate.

Changes in GDP binding to brown adipose tissue mitochondria and the uncoupling protein

1988 ◽  
Vol 255 (6) ◽  
pp. E865-E870
Author(s):  
A. G. Swick ◽  
R. W. Swick
Keyword(s):  
Alkaline Phosphatase ◽  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Divalent Cations ◽  
Uncoupling Protein ◽  
Bat Mitochondria ◽  
Brown Adipose ◽  
Simplified Method

Incubation in vitro of brown adipose tissue (BAT) mitochondria with divalent cations, spermine, or alkaline phosphatase led to a marked increase in the binding of [3H]GDP. The effect of Mg2+ appeared to be the most specific and led to the largest increase in GDP binding. A simplified method was developed for measuring GDP binding to purified uncoupling protein from rat BAT mitochondria. Application of this method indicates that uncoupling protein from cold-acclimated rats binds twice as much GDP as uncoupling protein from cold-acclimated rats that were briefly returned to thermoneutrality, paralleling changes in GDP binding to the mitochondria. Incubation of BAT mitochondria with Mg2+ led to a smaller increase in GDP binding to the subsequently purified uncoupling protein, suggesting that divalent cations may somehow participate in the regulation of the activity of the uncoupling protein.

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