AMPK Methods and Protocols

made by fusing a series of organelle-targeting sequences (OTS) to

ABKAR (osABKARs) targeting it to various subcellular localiza-

tions. A collection of osABKARs allowed for monitoring the

AMPK dynamics at various subcellular compartments such as the

plasma membrane, Golgi apparatus, endoplasmic reticulum (ER),

mitochondria, and lysosomes as well as previously reported cyto-

solic and nuclear compartments (Fig.2).

In addition to these unimolecular AMPK biosensors, bimolec-

ular kinase activity reporter for AMPK (BimABKAR) was also

developed [14]. In this system, the donor fluorophore

Table 1
(continued)

Protein name Gene name Main localization

Peroxisome-proliferator-activated receptor gamma

coactivator 1-alpha

PGC1A Nucleoplasm

Tumor protein p73 TP73 Nucleoplasm

Cyclin-dependent kinase inhibitor 1B CDKN1B Nucleus

Forkhead box protein O3 FOXO3a Nucleus

Mdm4 MDM4 Nucleus

6-Phosphofructo-2-kinase/fructose-2,6-

bisphosphatase 2

PFKFB2 Nucleus

Hepatocyte nuclear factor 4 HNF4 Nucleus

Carbohydrate-responsive element-binding protein MLXIPL Nucleus

Phosphatase 1 regulatory subunit 12C PPP1R12C Nucleus

Serine/threonine-protein kinase PAK 2

Astrocytic phosphoprotein PEA-15 PEA15 Nucleus, cytosol

Clock component cryptochrome 1 CRY1 Nucleus, nuclear membrane,

microtubules

Serine/threonine-protein kinase PAK 2 PAK2 Nucleus, vesicles

Microtubule-associated protein tau MAPT Plasma membrane

Phospholipase D1 PLD1 Plasma membrane

Thioredoxin-interacting protein TXNIP Plasma membrane

Vasodilator-stimulated phosphoprotein VASP Plasma membrane, cell junctions,

focal adhesion sites

Brain-specific angiogenesis inhibitor 1-associated

protein 2

BAIAP2 Plasma membrane, cytosol

Cingulin CGN Tight junctions

Serine/threonine-protein kinase B-raf BRAF Vesicle

258 Takafumi Miyamoto et al.