Abstract
The AMP-activated protein kinase (AMPK) phosphorylates, and thereby inactivates, a number of enzymes involved in the regulation of lipid metabolism. We have studied the expression of the AMPK gene in a variety of rat tissues. The gene is transcribed into a message of approximately 9.5 kb as detected by Northern blotting. Highest expression of the AMPK message was found in skeletal muscle, which contained 20 amol/micrograms total RNA as determined by competitive reverse-transcription/polymerase chain reaction (RT-PCR). In liver, kidney, brain, mammary glands, heart and lung, AMPK mRNA levels ranged over 1-4 amol/micrograms total RNA. Adipose tissue contained less than 1 amol/microgram total RNA. A second AMPK mRNA form was detected by RT-PCR that was 142 bases shorter than the functional transcript. This transcript was apparently generated by alternative splicing of a single exon within the 5'-coding region. The shorter of the two messages, which is not translated into AMPK protein, contributed between 35-60% of AMPK mRNA in most tissues, but only 15-20% in skeletal muscle and heart. As a result, functional AMPK mRNA was sevenfold higher in skeletal muscle than in liver, although AMPK activity was much lower. By Western blotting, relatively large amounts of AMPK protein were detected in skeletal muscle compared to liver. AMPK isolated from skeletal muscle was not activated by treatment with AMPK kinase under conditions where liver AMPK was fully activated. A single 63-kDa polypeptide was immunoprecipitated from rat skeletal muscle using anti-peptide IgG against AMPK. In contrast, two additional polypeptides with apparent molecular masses of 38 kDa and 36 kDa co-precipitated with the 63-kDa AMPK protein from rat liver. These results indicate that the muscle enzyme has a different subunit organization compared to the liver enzyme, which may account for its low catalytic activity. Together, our results indicate a physiological role for AMPK in muscle, in addition to its previously described role in lipid metabolism.
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