Immunological detection of cDNA clones encoding the uncoupling protein of brown adipose tissue: Evidence for an antigenic determinant within the C-terminal eleven amino acids

1986 ◽  
Vol 6 (1) ◽  
pp. 87-94 ◽  
Author(s):  
Robert G. Ridley ◽  
Hasmukh V. Patel ◽  
Craig L. J. Parfett ◽  
Kimberly A. Olynyk ◽  
Susanna Reichling ◽  
...  

Poly(A)+ RNA was isolated from brown adipose tissue of cold acclimated rats and a fraction enriched for uncoupling protein mRNA was used to generate a cDNA library in pBR 322, Immunological screening of 1,500 colonies with an affinity-purified antiserum against the uncoupling protein yielded five positive clones, pUCPratl–5. Clone pUCPrat2 encoded the C-terminal 54 amino acids of rat uncoupling protein and exhibited 90% amino acid homology with the hamster protein. Clones pUCPrat3–5 encoded only the C-terminal 11 amino acids suggesting that an antigenic determinant lies within this sequence.

1987 ◽  
Vol 65 (11) ◽  
pp. 955-959 ◽  
Author(s):  
Hasmukh V. Patel ◽  
Karl B. Freeman ◽  
Michel Desautels

The time course of changes in the level of uncoupling protein mRNA when cold-acclimated mice were returned to a thermoneutral environment (33 °C) was examined using a cDNA probe. Upon deacclimation, there was a marked loss of uncoupling protein mRNA within 24 h, which precedes the loss of uncoupling protein from mitochondria. This loss of uncoupling protein mRNA was selective, since there was no change in the relative proportion of cytochrome c oxidase subunit IV mRNA or poly(A)+ RNA in total RNA. The results suggest that the decrease in the mitochondrial content of uncoupling protein during deacclimation is likely the result of turnover of existing protein, with very little replacement due to a lower level of its mRNA.


1994 ◽  
Vol 41 (3) ◽  
pp. 315-318 ◽  
Author(s):  
TSUTOMU KAZUMI ◽  
DANIEL RICQUIER ◽  
TETSUO MAEDA ◽  
TADAYUKI MASUDA ◽  
TOSHIKI HOZUMI ◽  
...  

1989 ◽  
Vol 67 (2-3) ◽  
pp. 147-151 ◽  
Author(s):  
Karl B. Freeman ◽  
Michael Heffernan ◽  
Zenobia Dhalla ◽  
Hasmukh V. Patel

The effect of temperature on the amount of uncoupling protein mRNA in rat brown adipose tissue was examined after 1 and 14 days of exposure to cold. The relative amounts after 1 day, compared with rats kept at a thermoneutral temperature of 28 °C, were 3.2 at 19 °C, 3.3 at 11 °C, and 2.1 at 3 °C. This suggests that in warm-acclimated rats, a maximal response to a cold stimulus in brown adipose tissue is reached by 19 °C. In contrast to these results, the relative amounts of uncoupling protein mRNA after 14 days of cold exposure, compared with rats left at 28 °C, were 1.2 at 19 °C, 1.9 at 11 °C, and 2.1 at 3 °C. Since it is known that the amount of uncoupling protein in cold-acclimated rats increases continuously with decrease in temperature, the amount of protein reflects the mRNA levels during later times but not the initial time of exposure to cold.Key words: brown adipose tissue, uncoupling protein mRNA.


1987 ◽  
Vol 142 (3) ◽  
pp. 696-701 ◽  
Author(s):  
Susanna Reichling ◽  
Robert G. Ridley ◽  
Hasmukh V. Patel ◽  
Calvin B. Harley ◽  
Karl B. Freeman

1988 ◽  
Vol 252 (3) ◽  
pp. 843-849 ◽  
Author(s):  
F J López-Soriano ◽  
J A Fernández-López ◽  
T Mampel ◽  
F Villarroya ◽  
R Iglesias ◽  
...  

The net uptake/release of glucose, lactate and amino acids from the bloodstream by the interscapular brown adipose tissue of control, cold-exposed and cold-acclimated rats was estimated by measurement of arteriovenous differences in their concentrations. In the control animals amino acids contributed little to the overall energetic needs of the tissue; glucose uptake was more than compensated by lactate efflux. Cold-exposure resulted in an enhancement of amino acid utilization and of glucose uptake, with high lactate efflux. There was a net glycine and proline efflux that partly compensated the positive nitrogen balance of the tissue; amino acids accounted for about one-third of the energy supplied by glucose to the tissue. Cold-acclimation resulted in a very high increase in glucose uptake, with a parallel decrease in lactate efflux and amino acid consumption. Branched-chain amino acids, however, were more actively utilized. This was related with a much higher alanine efflux, in addition to that of glycine and proline. It is suggested that most of the glucose used during cold-exposure is returned to the bloodstream as lactate under conditions of active lipid utilization, amino acids contributing their skeletons largely in anaplerotic pathways. On the other hand, cold-acclimation resulted in an important enhancement of glucose utilization, with lowered amino acid oxidation. Amino acids are thus used as metabolic substrates by the brown adipose tissue of rats under conditions of relatively scarce substrate availability, but mainly as anaplerotic substrates, in parallel to glucose. Cold-acclimation results in a shift of the main substrates used in thermogenesis from lipid to glucose, with a much lower need for amino acids.


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