Dataset Information

Characterization of the liver kinase B1-mouse protein-25 -Ste-20-related adaptor protein complex in adult mouse skeletal muscle.

ABSTRACT: In liver, the AMP-activated protein kinase kinase (AMPKK) complex was identified as the association of liver kinase B1 (LKB1), mouse protein 25 (MO25?/?), and Ste-20-related adaptor protein (STRAD?/?); however, this complex has yet to be characterized in skeletal muscle. We demonstrate the expression of the LKB1-MO25-STRAD complex in skeletal muscle, confirm the absence of mRNA splice variants, and report the relative mRNA expression levels of these proteins in control and muscle-specific LKB1 knockout (LKB1(-/-)) mouse muscle. LKB1 detection in untreated control and LKB1(-/-) muscle lysates revealed two protein bands (50 and 60 kDa), although only the heavier band was diminished in LKB1(-/-) samples [55 ± 2.5 and 13 ± 1.5 arbitrary units (AU) in control and LKB1(-/-), respectively, P < 0.01], suggesting that LKB1 is not represented at 50 kDa, as previously cited. The 60-kDa LKB1 band was further confirmed following purification using polyethylene glycol (43 ± 5 and 8.4 ± 4 AU in control and LKB1(-/-), respectively, P < 0.01) and ion-exchange fast protein liquid chromatography. Mass spectrometry confirmed LKB1 protein detection in the 60-kDa protein band, while none was detected in the 50-kDa band. Coimmunoprecipitation assays demonstrated LKB1-MO25-STRAD complex formation. Quantitative PCR revealed significantly reduced LKB1, MO25?, and STRAD? mRNA in LKB1(-/-) muscle. These findings demonstrate that the LKB1-MO25-STRAD complex is the principal AMPKK in skeletal muscle.

SUBMITTER: Smith CD

PROVIDER: S-EPMC3233888 | biostudies-other | 2011 Dec

REPOSITORIES: biostudies-other

ACCESS DATA

Publications

Characterization of the liver kinase B1-mouse protein-25 -Ste-20-related adaptor protein complex in adult mouse skeletal muscle.

Smith Cody D CD Compton Richard A RA Bowler Joshua S JS Kemp Jonathan T JT Sudweeks Sterling N SN Thomson David M DM Winder William W WW

Journal of applied physiology (Bethesda, Md. : 1985) 20110908 6

In liver, the AMP-activated protein kinase kinase (AMPKK) complex was identified as the association of liver kinase B1 (LKB1), mouse protein 25 (MO25α/β), and Ste-20-related adaptor protein (STRADα/β); however, this complex has yet to be characterized in skeletal muscle. We demonstrate the expression of the LKB1-MO25-STRAD complex in skeletal muscle, confirm the absence of mRNA splice variants, and report the relative mRNA expression levels of these proteins in control and muscle-specific LKB1 k ...[more]

PMID: 21903876

Similar Datasets

Project description:Protein S-glutathionylation is an important reversible post-translational modification implicated in redox signaling. Oxidative modifications to protein thiols can alter the activity of metabolic enzymes, transcription factors, kinases, phosphatases, and the function of contractile proteins. However, the extent to which muscle contraction induces oxidative modifications in redox sensitive thiols is not known. The purpose of this study was to determine the targets of S-glutathionylation redox signaling following fatiguing contractions. Anesthetized adult male CB6F1 (BALB/cBy × C57BL/6) mice were subjected to acute fatiguing contractions for 15?min using in vivo stimulations. The right (stimulated) and left (unstimulated) gastrocnemius muscleswere collected 60?min after the last stimulation and processed for redox proteomics assay of S-glutathionylation. Using selective reduction with a glutaredoxin enzyme cocktail and resin-assisted enrichment technique, we quantified the levels of site-specific protein S-glutathionylation at rest and following fatiguing contractions. Redox proteomics revealed over 2200 sites of S-glutathionylation modifications, of which 1290 were significantly increased after fatiguing contractions. Muscle contraction leads to the greatest increase in S-glutathionylation in the mitochondria (1.03%) and the smallest increase in the nucleus (0.47%). Regulatory cysteines were significantly S-glutathionylated on mitochondrial complex I and II, GAPDH, MDH1, ACO2, and mitochondrial complex V among others. Similarly, S-glutathionylation of RYR1, SERCA1, titin, and troponin I2 are known to regulate muscle contractility and were significantly S-glutathionylated after just 15?min of fatiguing contractions. The largest fold changes (> 1.6) in the S-glutathionylated proteome after fatigue occurred on signaling proteins such as 14-3-3 protein gamma and MAP2K4, as well as proteins like SERCA1, and NDUV2 of mitochondrial complex I, at previously unknown glutathionylation sites. These findings highlight the important role of redox control over muscle physiology, metabolism, and the exercise adaptive response. This study lays the groundwork for future investigation into the altered exercise adaptation associated with chronic conditions, such as sarcopenia.

Dataset Information

Characterization of the liver kinase B1-mouse protein-25 -Ste-20-related adaptor protein complex in adult mouse skeletal muscle.

Publications

Characterization of the liver kinase B1-mouse protein-25 -Ste-20-related adaptor protein complex in adult mouse skeletal muscle.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure