Cloning and functional characterization of an uncoupling protein homolog in hummingbirds.
The cDNA of an uncoupling protein (UCP) homolog has been cloned from the swallow-tailed hummingbird, Eupetomena macroura. The hummingbird uncoupling protein (HmUCP) cDNA was amplified from pectoral muscle (flight muscle) using RT-PCR and primers for conserved domains of various known UCP homologs. The rapid amplification of cDNA ends (RACE) method was used to complete the cloning of the 5' and 3' ends of the open reading frame. The HmUCP coding region contains 915 nucleotides, and the deduced protein sequence consists of 304 amino acids, being approximately 72, 70, and 55% identical to human UCP3, UCP2, and UCP1, respectively. The uncoupling activity of this novel protein was characterized in yeast. In this expression system, the 12CA5-tagged HmUCP fusion protein was detected by Western blot in the enriched mitochondrial fraction. Similarly to rat UCP1, HmUCP decreased the mitochondrial membrane potential as measured in whole yeast by uptake of the fluorescent potential-sensitive dye 3',3-dihexyloxacarbocyanine iodide. The HmUCP mRNA is primarily expressed in skeletal muscle, but high levels can also be detected in heart and liver, as assessed by Northern blot analysis. Lowering the room's temperature to 12-14 degrees C triggered the cycle torpor/rewarming, typical of hummingbirds. Both in the pectoral muscle and heart, HmUCP mRNA levels were 1.5- to 3.4-fold higher during torpor. In conclusion, this is the first report of an UCP homolog in birds. The data indicate that HmUCP has the potential to function as an UCP and could play a thermogenic role during rewarming.
Vianna
C R
CR
Department of Physiology, Biosciences Institute, University of Sao Paulo, São Paulo 05508, Brazil.
Hagen
T
T
Zhang
C Y
CY
Bachman
E
E
Boss
O
O
Gereben
B
B
Moriscot
A S
AS
Lowell
B B
BB
Bicudo
J E
JE
Bianco
A C
AC
D000595:Amino Acid Sequence; D000818:Animals; D001717:Birds / Q000235:genetics* / Q000502:physiology; D002352:Carrier Proteins / Q000235:genetics*; D003001:Cloning, Molecular; D007473:Ion Channels; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000235:genetics*; D026901:Membrane Transport Proteins; D008928:Mitochondria / Q000502:physiology; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D010802:Phylogeny; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis; D012441:Saccharomyces cerevisiae / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D022722:Thermogenesis; D014018:Tissue Distribution; D000071256:Uncoupling Protein 1; D000071246:Uncoupling Protein 2; D000071247:Uncoupling Protein 3
D002352:Carrier Proteins; D007473:Ion Channels; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D024101:Mitochondrial Proteins; D011506:Proteins; D012333:RNA, Messenger; C000604371:UCP1 protein, human; C000604389:UCP2 protein, human; C000604392:UCP3 protein, human; C000604373:Ucp1 protein, rat; C000604391:Ucp2 protein, rat; C000604394:Ucp3 protein, rat; D000071256:Uncoupling Protein 1; D000071246:Uncoupling Protein 2; D000071247:Uncoupling Protein 3