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Human Regulation of Circadian Rhythm

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Western Blotting

Background: β-transducin repeat-containing protein (β-TrCP or FBW1A) is an F-box family protein characterized by the presence of the protein-protein mediating F-box domain first described in cyclin F. F-box proteins act as substrate adaptors that target proteins containing a specific phosphorylated sequence element, referred to as a phosphodegron, to the SCF E3 ubiquitin ligase complex for ubiquitination (1,2). β-TrCP targets many important proteins with diverse functions, such as p53, H-Ras, Smad4, IκBα, β-catenin, and the cell cycle checkpoint protein claspin, for ubiquitin-mediated degradation (3-5). Research studies have shown that inhibition of β-TrCP expression has a demonstrated benefit in the treatment of prostate cancer (6).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Chromatin IP, Immunoprecipitation, Western Blotting

Background: Reverse orientation c-erbA gene α (Rev-erbα, EAR-1, or NR1D1) is a widely expressed member of the orphan nuclear receptor family of proteins (1). Rev-erbα is highly expressed in adipose tissue, skeletal muscle, brain and liver, and regulates cellular proliferation and differentiation. Expression increases during differentiation in adipocytes and ectopic expression of Rev-erbα potentiates the adipocyte differentiation of 3T3-L1 cells (2). In addition, expression oscillates with circadian rhythm in liver cells and Rev-erbα regulates expression of BMAL1, ApoA-I and ApoC-III, all key regulators of circadian rhythm (3-7). Phosphorylation of Rev-erbα Ser55 and Ser59 by GSK-3β appears to stabilize Rev-erbα protein levels and is important for synchronizing and maintaining the circadian clock (8). Rev-erbα also regulates inflammation by targeting the NF-κB responsive genes IL-6 and COX-2 (9). Rev-erbα lacks the activation function 2 domain required for ligand-dependent activation of transcription by other members of the nuclear receptor family; thus it behaves as a constitutive repressor protein, recruiting the nuclear receptor co-repressor (N-CoR)/HDAC3 complex to target genes to repress transcription (10).

$303
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Western Blotting

Background: Reverse orientation c-erbA gene α (Rev-erbα, EAR-1, or NR1D1) is a widely expressed member of the orphan nuclear receptor family of proteins (1). Rev-erbα is highly expressed in adipose tissue, skeletal muscle, brain and liver, and regulates cellular proliferation and differentiation. Expression increases during differentiation in adipocytes and ectopic expression of Rev-erbα potentiates the adipocyte differentiation of 3T3-L1 cells (2). In addition, expression oscillates with circadian rhythm in liver cells and Rev-erbα regulates expression of BMAL1, ApoA-I and ApoC-III, all key regulators of circadian rhythm (3-7). Phosphorylation of Rev-erbα Ser55 and Ser59 by GSK-3β appears to stabilize Rev-erbα protein levels and is important for synchronizing and maintaining the circadian clock (8). Rev-erbα also regulates inflammation by targeting the NF-κB responsive genes IL-6 and COX-2 (9). Rev-erbα lacks the activation function 2 domain required for ligand-dependent activation of transcription by other members of the nuclear receptor family; thus it behaves as a constitutive repressor protein, recruiting the nuclear receptor co-repressor (N-CoR)/HDAC3 complex to target genes to repress transcription (10).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: β-transducin repeat-containing protein (β-TrCP or FBW1A) is an F-box family protein characterized by the presence of the protein-protein mediating F-box domain first described in cyclin F. F-box proteins act as substrate adaptors that target proteins containing a specific phosphorylated sequence element, referred to as a phosphodegron, to the SCF E3 ubiquitin ligase complex for ubiquitination (1,2). β-TrCP targets many important proteins with diverse functions, such as p53, H-Ras, Smad4, IκBα, β-catenin, and the cell cycle checkpoint protein claspin, for ubiquitin-mediated degradation (3-5). Research studies have shown that inhibition of β-TrCP expression has a demonstrated benefit in the treatment of prostate cancer (6).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: Casein Kinase I (CK1 or CKI) is the name given to a family of kinases consisting of multiple isoforms (α, α', β, γ1-3, δ, and ε) with a conserved N-terminal kinase domain and a variable C-terminal sequence that determines subcellular localization and regulates enzyme activity (1-3). Indeed, multiple inhibitory autophosphorylation sites have been identified near the C terminus of CK1ε (3). This ubiquitously expressed family of protein kinases has been implicated in multiple processes including DNA repair, cell morphology, and Wnt signaling (4). Perhaps the best understood role of CK1 is to provide the priming phosphorylation of β-catenin at Ser45 to produce the consensus GSK-3 substrate motif (S/T-X-X-X-pS) (4).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Western Blotting

Background: PPARγ coactivator-1α (PGC-1α) was originally identified as a transcriptional coactivator whose expression closely correlated with adaptive thermogenesis following exposure to cold temperatures (1). Named for its association with the nuclear receptor peroxisome-proliferator activated receptor (PPARγ), PGC-1α interacts with a diverse array of transcription factors to regulate numerous aspects of cell physiology (2). PGC-1α helps to regulate cell processes important in adaptive thermogenesis and energy metabolism, including the related functions of glucose uptake, gluconeogenesis, insulin secretion, and mitochondrial biogenesis (3). Long thought to be a potential therapeutic target for the treatment of type II diabetes, obesity, cardiomyopathy, or other metabolic disorders (reviewed in 4), a recent functional survey found no obvious differences in PPARγ activity associated with recognized PGC-1α variants (5).

$303
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: Circadian rhythms govern many key physiological processes that fluctuate with a period of approximately 24 hours. These processes include the sleep-wake cycle, glucose, lipid and drug metabolism, heart rate, hormone secretion, renal blood flow, and body temperature, as well as basic cellular processes such as DNA repair and the timing of the cell division cycle (1,2). The mammalian circadian system consists of many individual tissue-specific clocks (peripheral clocks) that are controlled by a master circadian pacemaker residing in the suprachiasmatic nuclei (SCN) of the brain (1,2). The periodic circadian rhythm is prominently manifested by the light-dark cycle, which is sensed by the visual system and processed by the SCN. The SCN processes the light-dark information and synchronizes peripheral clocks through neural and humoral output signals (1,2).The cellular circadian clockwork consists of interwoven positive and negative regulatory loops, or limbs (1,2). The positive limb includes the CLOCK and BMAL1 proteins, two basic helix-loop-helix-PAS containing transcription factors that bind E box enhancer elements and activate transcription of their target genes. CLOCK is a histone acetyltransferase (HAT) protein, which acetylates both histone H3 and H4 (3). BMAL1 binds to CLOCK and enhances its HAT activity (3). The CLOCK/BMAL1 dimer exhibits a periodic oscillation in both nuclear/cytoplasmic localization and protein levels, both of which are regulated by phosphorylation (4,5). CLOCK/BMAL1 target genes include the Cry and Per genes, whose proteins form the negative limb of the circadian clockwork system (1,2). CRY and PER proteins (CRY1, CRY2, PER1, PER2 and PER3) form oligomers that also periodically shuttle between the nucleus and cytoplasm. When in the nucleus, CRY/PER proteins inhibit CLOCK/BMAL1-mediated transcriptional activation, thus completing the circadian transcriptional loop (1,2). In tissues, roughly six to eight percent of all genes exhibit a circadian expression pattern (1,2). This 24-hour periodicity in gene expression results from coordination of the positive and negative regulatory limbs of the cellular clockwork system, and is fine-tuned by outside signals received from the SCN.

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Chromatin IP, Immunoprecipitation, Western Blotting

Background: Circadian rhythms govern many key physiological processes that fluctuate with a period of approximately 24 hours. These processes include the sleep-wake cycle, glucose, lipid and drug metabolism, heart rate, hormone secretion, renal blood flow, and body temperature, as well as basic cellular processes such as DNA repair and the timing of the cell division cycle (1,2). The mammalian circadian system consists of many individual tissue-specific clocks (peripheral clocks) that are controlled by a master circadian pacemaker residing in the suprachiasmatic nuclei (SCN) of the brain (1,2). The periodic circadian rhythm is prominently manifested by the light-dark cycle, which is sensed by the visual system and processed by the SCN. The SCN processes the light-dark information and synchronizes peripheral clocks through neural and humoral output signals (1,2).The cellular circadian clockwork consists of interwoven positive and negative regulatory loops, or limbs (1,2). The positive limb includes the CLOCK and BMAL1 proteins, two basic helix-loop-helix-PAS containing transcription factors that bind E box enhancer elements and activate transcription of their target genes. CLOCK is a histone acetyltransferase (HAT) protein, which acetylates both histone H3 and H4 (3). BMAL1 binds to CLOCK and enhances its HAT activity (3). The CLOCK/BMAL1 dimer exhibits a periodic oscillation in both nuclear/cytoplasmic localization and protein levels, both of which are regulated by phosphorylation (4,5). CLOCK/BMAL1 target genes include the Cry and Per genes, whose proteins form the negative limb of the circadian clockwork system (1,2). CRY and PER proteins (CRY1, CRY2, PER1, PER2 and PER3) form oligomers that also periodically shuttle between the nucleus and cytoplasm. When in the nucleus, CRY/PER proteins inhibit CLOCK/BMAL1-mediated transcriptional activation, thus completing the circadian transcriptional loop (1,2). In tissues, roughly six to eight percent of all genes exhibit a circadian expression pattern (1,2). This 24-hour periodicity in gene expression results from coordination of the positive and negative regulatory limbs of the cellular clockwork system, and is fine-tuned by outside signals received from the SCN.

$108
250 PCR reactions
500 µl
SimpleChIP® Human NR1D1 Promoter Primers contain a mix of forward and reverse PCR primers that are specific to a region of the human nuclear receptor subfamily 1, group D, member 1 (NR1D1) promoter. These primers can be used to amplify DNA that has been isolated using chromatin immunoprecipitation (ChIP). Primers have been optimized for use in SYBR® Green quantitative real-time PCR and have been tested in conjunction with SimpleChIP® Enzymatic Chromatin IP Kits #9002 and #9003 and ChIP-validated antibodies from Cell Signaling Technology®. The NR1D1 gene encodes for the transcriptional repressor Rev-erbα, an orphan nuclear receptor that regulates cell proliferation, differentiation, and circadian rhythms in multiple tissues and organ systems.
REACTIVITY
Human

Background: The chromatin immunoprecipitation (ChIP) assay is a powerful and versatile technique used for probing protein-DNA interactions within the natural chromatin context of the cell (1,2). This assay can be used to either identify multiple proteins associated with a specific region of the genome or to identify the many regions of the genome bound by a particular protein (3-6). ChIP can be used to determine the specific order of recruitment of various proteins to a gene promoter or to "measure" the relative amount of a particular histone modification across an entire gene locus (3,4). In addition to histone proteins, the ChIP assay can be used to analyze binding of transcription factors and co-factors, DNA replication factors, and DNA repair proteins. When performing the ChIP assay, cells are first fixed with formaldehyde, a reversible protein-DNA cross-linking agent that "preserves" the protein-DNA interactions occurring in the cell (1,2). Cells are lysed and chromatin is harvested and fragmented using either sonication or enzymatic digestion. Fragmented chromatin is then immunoprecipitated with antibodies specific to a particular protein or histone modification. Any DNA sequences that are associated with the protein or histone modification of interest will co-precipitate as part of the cross-linked chromatin complex and the relative amount of that DNA sequence will be enriched by the immunoselection process. After immunoprecipitation, the protein-DNA cross-links are reversed and the DNA is purified. Standard PCR or quantitative real-time PCR are often used to measure the amount of enrichment of a particular DNA sequence by a protein-specific immunoprecipitation (1,2). Alternatively, the ChIP assay can be combined with genomic tiling micro-array (ChIP on chip) techniques, high throughput sequencing (ChIP-Seq), or cloning strategies, all of which allow for genome-wide analysis of protein-DNA interactions and histone modifications (5-8). SimpleChIP® primers have been optimized for amplification of ChIP-isolated DNA using real-time quantitative PCR and provide important positive and negative controls that can be used to confirm a successful ChIP experiment.

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Western Blotting

Background: Casein Kinase I (CK1 or CKI) is the name given to a family of kinases consisting of multiple isoforms (α, α', β, γ1-3, δ, and ε) with a conserved N-terminal kinase domain and a variable C-terminal sequence that determines subcellular localization and regulates enzyme activity (1-3). Indeed, multiple inhibitory autophosphorylation sites have been identified near the C terminus of CK1ε (3). This ubiquitously expressed family of protein kinases has been implicated in multiple processes including DNA repair, cell morphology, and Wnt signaling (4). Perhaps the best understood role of CK1 is to provide the priming phosphorylation of β-catenin at Ser45 to produce the consensus GSK-3 substrate motif (S/T-X-X-X-pS) (4).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse

Application Methods: Immunoprecipitation, Western Blotting

Background: Inhibitor of DNA-binding-2 (Id2) is a member of the Id proteins which belong to the helix-loop-helix (HLH) protein family. The Id protein functions by binding to specific transcription factors and preventing their dimerization and DNA binding (1-3). Id2 interacts with a wide variety of transcription factors including E proteins (5), TCS (4), Pax (6) and the tumor suppressor Rb (1). Id2 has been shown to be important in regulating cellular differentiation, proliferation, development and tumorgenesis (7-9). In tumor cells, increased levels of Id2 functionally inactivate Rb, leading to cellular transformation and cancer (10,11). Id2 is therefore a promising therapeutic target for treatment of cancer (12).

$327
100 µl
This Cell Signaling Technology (CST) antibody is conjugated to biotin under optimal conditions. The unconjugated Phospho-CREB (Ser133) (87G3) Rabbit mAb #9198 reacts with human, mouse and rat phospho-CREB (Ser133) protein. CST expects that Phospho-CREB (Ser133) (87G3) Rabbit mAb (Biotinylated) #4095 will also recognize phospho-CREB (Ser133) in these species.
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Western Blotting

Background: CREB is a bZIP transcription factor that activates target genes through cAMP response elements. CREB is able to mediate signals from numerous physiological stimuli, resulting in regulation of a broad array of cellular responses. While CREB is expressed in numerous tissues, it plays a large regulatory role in the nervous system. CREB is believed to play a key role in promoting neuronal survival, precursor proliferation, neurite outgrowth, and neuronal differentiation in certain neuronal populations (1-3). Additionally, CREB signaling is involved in learning and memory in several organisms (4-6). CREB is able to selectively activate numerous downstream genes through interactions with different dimerization partners. CREB is activated by phosphorylation at Ser133 by various signaling pathways including Erk, Ca2+, and stress signaling. Some of the kinases involved in phosphorylating CREB at Ser133 are p90RSK, MSK, CaMKIV, and MAPKAPK-2 (7-9).

$260
100 µl
APPLICATIONS
REACTIVITY
D. melanogaster, Hamster, Human, Monkey, Mouse, Rat

Application Methods: Chromatin IP, Flow Cytometry, Immunofluorescence (Frozen), Immunofluorescence (Immunocytochemistry), Immunoprecipitation, Western Blotting

Background: CREB is a bZIP transcription factor that activates target genes through cAMP response elements. CREB is able to mediate signals from numerous physiological stimuli, resulting in regulation of a broad array of cellular responses. While CREB is expressed in numerous tissues, it plays a large regulatory role in the nervous system. CREB is believed to play a key role in promoting neuronal survival, precursor proliferation, neurite outgrowth, and neuronal differentiation in certain neuronal populations (1-3). Additionally, CREB signaling is involved in learning and memory in several organisms (4-6). CREB is able to selectively activate numerous downstream genes through interactions with different dimerization partners. CREB is activated by phosphorylation at Ser133 by various signaling pathways including Erk, Ca2+, and stress signaling. Some of the kinases involved in phosphorylating CREB at Ser133 are p90RSK, MSK, CaMKIV, and MAPKAPK-2 (7-9).

$262
3 nmol
300 µl
SignalSilence® CREB siRNA from Cell Signaling Technology (CST) allows the researcher to specifically inhibit CREB expression using RNA interference, a method whereby gene expression can be selectively silenced through the delivery of double stranded RNA molecules into the cell. All SignalSilence® siRNA products are rigorously tested in-house and have been shown to reduce target protein expression by western analysis.
REACTIVITY
Human

Background: CREB is a bZIP transcription factor that activates target genes through cAMP response elements. CREB is able to mediate signals from numerous physiological stimuli, resulting in regulation of a broad array of cellular responses. While CREB is expressed in numerous tissues, it plays a large regulatory role in the nervous system. CREB is believed to play a key role in promoting neuronal survival, precursor proliferation, neurite outgrowth, and neuronal differentiation in certain neuronal populations (1-3). Additionally, CREB signaling is involved in learning and memory in several organisms (4-6). CREB is able to selectively activate numerous downstream genes through interactions with different dimerization partners. CREB is activated by phosphorylation at Ser133 by various signaling pathways including Erk, Ca2+, and stress signaling. Some of the kinases involved in phosphorylating CREB at Ser133 are p90RSK, MSK, CaMKIV, and MAPKAPK-2 (7-9).

$327
50 tests
100 µl
This Cell Signaling Technology antibody is conjugated to Alexa Fluor® 488 fluorescent dye and tested in-house for direct flow cytometry and immunofluorescent analysis in human and mouse cells. The antibody is expected to exhibit the same species cross-reactivity as the unconjugated Phospho-CREB (Ser133) (87G3) Rabbit mAb #9198.
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Flow Cytometry

Background: CREB is a bZIP transcription factor that activates target genes through cAMP response elements. CREB is able to mediate signals from numerous physiological stimuli, resulting in regulation of a broad array of cellular responses. While CREB is expressed in numerous tissues, it plays a large regulatory role in the nervous system. CREB is believed to play a key role in promoting neuronal survival, precursor proliferation, neurite outgrowth, and neuronal differentiation in certain neuronal populations (1-3). Additionally, CREB signaling is involved in learning and memory in several organisms (4-6). CREB is able to selectively activate numerous downstream genes through interactions with different dimerization partners. CREB is activated by phosphorylation at Ser133 by various signaling pathways including Erk, Ca2+, and stress signaling. Some of the kinases involved in phosphorylating CREB at Ser133 are p90RSK, MSK, CaMKIV, and MAPKAPK-2 (7-9).

$111
20 µl
$260
100 µl
$630
300 µl
APPLICATIONS
REACTIVITY
D. melanogaster, Human, Monkey, Mouse, Rat

Application Methods: Chromatin IP, Chromatin IP-seq, Flow Cytometry, Immunofluorescence (Frozen), Immunofluorescence (Immunocytochemistry), Immunohistochemistry (Frozen), Immunohistochemistry (Paraffin), Immunoprecipitation, Western Blotting

Background: CREB is a bZIP transcription factor that activates target genes through cAMP response elements. CREB is able to mediate signals from numerous physiological stimuli, resulting in regulation of a broad array of cellular responses. While CREB is expressed in numerous tissues, it plays a large regulatory role in the nervous system. CREB is believed to play a key role in promoting neuronal survival, precursor proliferation, neurite outgrowth, and neuronal differentiation in certain neuronal populations (1-3). Additionally, CREB signaling is involved in learning and memory in several organisms (4-6). CREB is able to selectively activate numerous downstream genes through interactions with different dimerization partners. CREB is activated by phosphorylation at Ser133 by various signaling pathways including Erk, Ca2+, and stress signaling. Some of the kinases involved in phosphorylating CREB at Ser133 are p90RSK, MSK, CaMKIV, and MAPKAPK-2 (7-9).

$269
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunohistochemistry (Paraffin), Western Blotting

Background: Prospero homeobox protein 1 (PROX1) is a transcription factor known for its roles in organ development and lymphangiogenesis. It plays a critical role in the development of the CNS, lens, retina, liver, pancreas, and heart, and is considered to be the master regulator of the lymphatic system (1,2). PROX1 initiates the differentiation of lymphatic vasculature from the cardinal vein, where it is regulated by Sox18 (3,4). The PROX1 suppressor COUP-TFII represses the Notch pathway in venous endothelium, which prevents arterialization (4). HIF-1α and HIF-1β mediated hypoxia induces PROX1, which suggests a means of promoting lymphangiogenesis. Since the tumor microenvironment is typically hypoxic, regulation of PROX1 by hypoxia may also explain the up-regulation of this transcription factor in some cancers (2). PROX1 promotes colon cancer progression by down-regulating E-cadherin via miR-9, which promotes epithelial-mesenchymal transition (EMT) and metastasis (5). The PROX1 protein can act as a tumor suppressor in cases of hepatocellular carcinoma. PROX1 represses transcription of TWIST1, a transcription factor that promotes metastasis by binding the E-cadherin promoter. The function of PROX1 in other cancers is an area of active research (6).

$303
100 µl
$717
300 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Western Blotting

Background: AMP-activated protein kinase (AMPK) is highly conserved from yeast to plants and animals and plays a key role in the regulation of energy homeostasis (1). AMPK is a heterotrimeric complex composed of a catalytic α subunit and regulatory β and γ subunits, each of which is encoded by two or three distinct genes (α1, 2; β1, 2; γ1, 2, 3) (2). The kinase is activated by an elevated AMP/ATP ratio due to cellular and environmental stress, such as heat shock, hypoxia, and ischemia (1). The tumor suppressor LKB1, in association with accessory proteins STRAD and MO25, phosphorylates AMPKα at Thr172 in the activation loop, and this phosphorylation is required for AMPK activation (3-5). AMPKα is also phosphorylated at Thr258 and Ser485 (for α1; Ser491 for α2). The upstream kinase and the biological significance of these phosphorylation events have yet to be elucidated (6). The β1 subunit is post-translationally modified by myristoylation and multi-site phosphorylation including Ser24/25, Ser96, Ser101, Ser108, and Ser182 (6,7). Phosphorylation at Ser108 of the β1 subunit seems to be required for the activation of AMPK enzyme, while phosphorylation at Ser24/25 and Ser182 affects AMPK localization (7). Several mutations in AMPKγ subunits have been identified, most of which are located in the putative AMP/ATP binding sites (CBS or Bateman domains). Mutations at these sites lead to reduction of AMPK activity and cause glycogen accumulation in heart or skeletal muscle (1,2). Accumulating evidence indicates that AMPK not only regulates the metabolism of fatty acids and glycogen, but also modulates protein synthesis and cell growth through EF2 and TSC2/mTOR pathways, as well as blood flow via eNOS/nNOS (1).

$122
20 µl
$307
100 µl
$719
300 µl
APPLICATIONS
REACTIVITY
D. melanogaster, Hamster, Human, Monkey, Mouse, Rat, S. cerevisiae

Application Methods: Immunohistochemistry (Paraffin), Immunoprecipitation, Western Blotting

Background: AMP-activated protein kinase (AMPK) is highly conserved from yeast to plants and animals and plays a key role in the regulation of energy homeostasis (1). AMPK is a heterotrimeric complex composed of a catalytic α subunit and regulatory β and γ subunits, each of which is encoded by two or three distinct genes (α1, 2; β1, 2; γ1, 2, 3) (2). The kinase is activated by an elevated AMP/ATP ratio due to cellular and environmental stress, such as heat shock, hypoxia, and ischemia (1). The tumor suppressor LKB1, in association with accessory proteins STRAD and MO25, phosphorylates AMPKα at Thr172 in the activation loop, and this phosphorylation is required for AMPK activation (3-5). AMPKα is also phosphorylated at Thr258 and Ser485 (for α1; Ser491 for α2). The upstream kinase and the biological significance of these phosphorylation events have yet to be elucidated (6). The β1 subunit is post-translationally modified by myristoylation and multi-site phosphorylation including Ser24/25, Ser96, Ser101, Ser108, and Ser182 (6,7). Phosphorylation at Ser108 of the β1 subunit seems to be required for the activation of AMPK enzyme, while phosphorylation at Ser24/25 and Ser182 affects AMPK localization (7). Several mutations in AMPKγ subunits have been identified, most of which are located in the putative AMP/ATP binding sites (CBS or Bateman domains). Mutations at these sites lead to reduction of AMPK activity and cause glycogen accumulation in heart or skeletal muscle (1,2). Accumulating evidence indicates that AMPK not only regulates the metabolism of fatty acids and glycogen, but also modulates protein synthesis and cell growth through EF2 and TSC2/mTOR pathways, as well as blood flow via eNOS/nNOS (1).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Western Blotting

Background: AMP-activated protein kinase (AMPK) is highly conserved from yeast to plants and animals and plays a key role in the regulation of energy homeostasis (1). AMPK is a heterotrimeric complex composed of a catalytic α subunit and regulatory β and γ subunits, each of which is encoded by two or three distinct genes (α1, 2; β1, 2; γ1, 2, 3) (2). The kinase is activated by an elevated AMP/ATP ratio due to cellular and environmental stress, such as heat shock, hypoxia, and ischemia (1). The tumor suppressor LKB1, in association with accessory proteins STRAD and MO25, phosphorylates AMPKα at Thr172 in the activation loop, and this phosphorylation is required for AMPK activation (3-5). AMPKα is also phosphorylated at Thr258 and Ser485 (for α1; Ser491 for α2). The upstream kinase and the biological significance of these phosphorylation events have yet to be elucidated (6). The β1 subunit is post-translationally modified by myristoylation and multi-site phosphorylation including Ser24/25, Ser96, Ser101, Ser108, and Ser182 (6,7). Phosphorylation at Ser108 of the β1 subunit seems to be required for the activation of AMPK enzyme, while phosphorylation at Ser24/25 and Ser182 affects AMPK localization (7). Several mutations in AMPKγ subunits have been identified, most of which are located in the putative AMP/ATP binding sites (CBS or Bateman domains). Mutations at these sites lead to reduction of AMPK activity and cause glycogen accumulation in heart or skeletal muscle (1,2). Accumulating evidence indicates that AMPK not only regulates the metabolism of fatty acids and glycogen, but also modulates protein synthesis and cell growth through EF2 and TSC2/mTOR pathways, as well as blood flow via eNOS/nNOS (1).