AMPK is a heterotrimeric complex composed of a catalytic α- subunit and two regulatory β- and γ- subunits. Each subunit exists in multiple isoforms encoded by separate genes (α1, α2, β1, β2, γ1, γ2, and γ3), and their combination give rise to a variety of AMPK holoenzymes (). Phosphorylation of Threonine 172 in the catalytic domain of the α subunit is required for AMPK activation. The C-terminal domain is required for binding with the β and γ subunits. The β subunit has scaffold/docking properties and contains two domains, glycogen binding domain and the C-terminal domain for binding with the α and γ subunits [6
]. The β subunit also contains several sites for post-translational modifications such as myristoylation and phosphorylation which may regulate the subcellular localization of the heterotrimer [7
]. The γ subunit contains four cystathionine-β-synthase (CBS) motifs in the C-terminus. Each pairs of the CBS motifs tightly associate in a pseudo dimeric arrangement through their β-sheets forming a so-called Bateman domain. Nucleotide binding assays suggested that each Bateman domain bound one molecule of AMP or ATP in an exclusive manner [8
]. This observation was supported by a recent crystal structure study which in addition revealed a third site containing a non-exchangeable AMP [9
] . The binding of AMP activates while the binding of ATP inactivates the kinase, thus, the two exchangeable sites on the γ subunit are responsible for the sensitivity of AMPK to the AMP/ATP ratio in the environment.
Figure 1 Regulation of the AMPK signaling cascade. The intracellular AMP/ATP ratio is a primary regulator of the system. Increased AMP/ATP activates AMPK via three mechanisms 1) allosteric activation; 2) promotes the Thr 172 phosphorylation of the α- AMPK (more ...)
The AMP/ATP ratio regulates AMPK activity by several mechanisms (). First, the binding of AMP in the γ subunit allosterically activates the kinase. Increased AMP binding also facilitates the phosphorylation of and protects against the dephosphorylation of Thr 172, thus promotes and sustains the kinase activity [10
]. The combined effects of the allosteric activation and Thr 172 phosphorylation increase AMPK activity by 1000-fold [11
]. Since the [ATP] in the cell is several orders of magnitude higher than [AMP], AMPK activity is very low in unstressed conditions. However, because of the large difference in free concentrations of the nucleotides, a small decrease in [ATP] can lead to a large increase of [AMP] (ATP → ADP → AMP) resulting in a great change of AMP/ATP. In this way, AMPK functions as an ultra-sensitive gauge of cellular energetic status [12
Multiple upstream kinases of AMPK (AMPKK) have been reported, including LKB1, Ca2+
/calmodulin-dependent protein kinase kinase β (CaMKKβ), and transforming growth factor-β activated kinase 1 (TAK1) (). LKB1, encoded by the Peutz-Jeghers syndrome tumor suppressor gene, exists in a heterotrimetric complex associated with two accessory proteins, mouse protein 25 (MO25α/β) and Ste20-related adaptor protein (STRADα/β). LKB1 complex functions as a constitutively active kinase that phosphorylates AMPK in the presence of AMP [13
]. LKB1-AMPK pathway exists in majority of cell and tissue types except in Hela cells where LKB1 is absent [14
]. CaMKKβ has considerable sequence and structural homology with LKB1 but its activity is dependent on intracellular Ca2+
levels. Recent studies suggest that CaMKKβ regulates AMPK in a Ca2+
/calmodulin-dependent and AMP-independent manner [11
]. The expression of CaMKKβ in the heart is very low but the CaMKKβ-AMPK cascade likely has an important role in the neurological tissues [15
]. TAK1 was first shown to activate the yeast AMPK homologue Snif1 and purified mammalian AMPK and more recently in human cell culture [17
]. Deletion of TAK1 results in decreased phosphorylation of AMPK in mice [19
Recent studies have also proposed that the AMPKKs may be constitutively active and the regulation of AMPK phoshporylation and activity is, instead, achieved by the dephosphorylation process. Increased AMP has been shown to prevent the dephosphorylation and inactivation of AMPK by protein phosphatase [10
] but the specific phosphatase for the AMPK in vivo
as well as its regulation remains to be defined.
Several endocrine factors/hormones have been reported to regulate AMPK activity, such as adiponectin, leptin, ciliary neurotrophic factor and ghrelin [20
]. The mechanisms of action for these factors/hormones are complex involving both central nervous system and direct effects on the peripheral tissue. It is though very unlikely these factors/hormones interact directly with the AMPK complex.