AMP-activated protein kinase in skeletal muscle: From structure and localization to its role as a master regulator of cellular metabolism

The AMP-activated protein kinase (AMPK) is a metabolite sensing serine/threonine kinase that has been termed the master regulator of cellular energy metabolism due to its numerous roles in the regulation of glucose, lipid, and protein metabolism. In this review, we first summarize the current literature on a number of important aspects of AMPK in skeletal muscle. These include the following: (1) the structural components of the three AMPK subunits (i.e. AMPKα, β, and γ), and their differential localization in response to stimulation in muscle; (2) the biochemical regulation of AMPK by AMP, protein phosphatases, and its three known upstream kinases, LKB1, Ca²⁺/calmodulin-dependent protein kinase kinase (CaMKK), and transforming growth factor-β-activated kinase 1 (TAK1); (3) the pharmacological agents that are currently available for the activation and inhibition of AMPK; (4) the physiological stimuli that activate AMPK in muscle; and (5) the metabolic processes that AMPK regulates in skeletal muscle. Received 04 May 2008; received after revision 14 June 2008; accepted 14 July 2008     

Let-7b is a novel regulator of hepatitis C virus replication

The non-coding microRNA (miRNA) is involved in the regulation of hepatitis C virus (HCV) infection and offers an alternative target for developing anti-HCV agent. In this study, we aim to identify novel cellular miRNAs that directly target the HCV genome with anti-HCV therapeutic potential. Bioinformatic analyses were performed to unveil liver-abundant miRNAs with predicted target sequences on HCV genome. Various cell-based systems confirmed that let-7b plays a negative role in HCV expression. In particular, let-7b suppressed HCV replicon activity and down-regulated HCV accumulation leading to reduced infectivity of HCVcc. Mutational analysis identified let-7b binding sites at the coding sequences of NS5B and 5'-UTR of HCV genome that were conserved among various HCV genotypes. We further demonstrated that the underlying mechanism for let-7b-mediated suppression of HCV RNA accumulation was not dependent on inhibition of HCV translation. Let-7b and IFNα-2a also elicited a synergistic inhibitory effect on HCV infection. Together, let-7b represents a novel cellular miRNA that targets the HCV genome and elicits anti-HCV activity. This study thereby sheds new insight into understanding the role of host miRNAs in HCV pathogenesis and to developing a potential anti-HCV therapeutic strategy.     

Increased plasma fibrinogen and the release of a fibrinogen enhancing factor in tumour-bearing rats

Zusammenfassung Die Freisetzung eines die Fibrinogensynthese stimulierenden Faktors aus nekrotischem Gewebe in die Blutbahn ist für den Anstieg des Plasmafibrinogens bei experimentellen Rattentumoren verantwortlich.     

Pyridine nucleotide oxidized to reduced ratio as a regulator of muscular performance

Zusammenfassung Die Pyridinnukleotid-Gehalte von trainierten und untrainierten Muskeln werden nach zehnminütiger Arbeit miteinander verglichen. Das Verhältnis der oxidierten zu reduzierten Pyridinnukleotiden ist beim trainierten Muskel stärker oxidiert als beim nicht trainierten Muskel.     

MicroRNA miR-196a is a central regulator of HOX-B7 and BMP4 expression in malignant melanoma

Since bone morphogenetic proteins (BMPs) play an important role in melanoma progression, we aimed to determine the molecular mechanisms leading to overexpression of BMP4 in melanoma cells compared to normal melanocytes. With our experimental approach we revealed that loss of expression of a microRNA represents the starting point for a signaling cascade finally resulting in overexpression of BMP4 in melanoma cells. In detail, strongly reduced expression of the microRNA miR-196a in melanoma cells compared to healthy melanocytes leads to enhanced HOX-B7 mRNA and protein levels, which subsequently raise Ets-1 activity by inducing basic fibroblast growth factor (bFGF). Ets-1 finally accounts for induction of BMP4 expression. We were furthermore able to demonstrate that bFGF-mediated induction of migration is achieved via activation of BMP4, thus determining BMP4 as major modulator of migration in melanoma. In summary, our study provides insights into the early steps of melanoma progression and might thereby harbor therapeutic relevance.     

miR-124-3p is a chronic regulator of gene expression after brain injury

Traumatic brain injury (TBI) initiates molecular and cellular pathologies that underlie post-injury morbidities, including hippocampus-related memory decline and epileptogenesis. Non-coding small RNAs are master regulators of gene expression with the potential to affect multiple molecular pathways. To evaluate whether hippocampal gene expression networks are chronically regulated by microRNAs after TBI, we sampled the dentate gyrus of rats with severe TBI induced by lateral fluid-percussion injury 3 months earlier. Ingenuity pathway analysis revealed 30 upregulated miR-124-3p targets, suggesting that miR-124-3p is downregulated post-TBI (z-score?=?? 5.146, p?<?0.05). Droplet digital polymerase chain reaction (ddPCR) and in situ hybridization confirmed the chronic downregulation of miR-124-3p (p?<?0.05). Quantitative PCR analysis of two targets, Plp2 and Stat3, indicated that their upregulation correlated with the miR-124-3p downregulation (r?=?? 0.647, p?<?0.05; r?=?? 0.629, p?<?0.05, respectively). Immunohistochemical staining of STAT3 confirmed the increased protein expression. STRING analysis showed that 9 of the 30 miR-124-3p targets belonged to a STAT3 network. Reactome analysis and data mining connected the targets especially to inflammation and signal transduction. L1000CDS2 software revealed drugs (e.g., importazole, trichostatin A, and IKK-16) that could reverse the observed molecular changes. The translational value of our data was emphasized by in situ hybridization showing chronic post-traumatic downregulation of miR-124-3p in the dentate gyrus of TBI patients. Analysis of another brain injury model, status epilepticus, highlighted the fact that chronic downregulation of miR-124 is a common phenomenon after brain injury. Together, our findings indicate that miR-124-3p is a chronic modulator of molecular networks relevant to post-injury hippocampal pathologies in experimental models and in humans.     

Biotin as a regulator of some haematic parameters and of DNA-content of the liver of old rats

Summary Biotin administration to old rats (28 months) causes in the blood an increase of ATP, glucose, triglycerides, alkaline phosphatase and a decrease of cholesterol and acid phosphatase; in the liver DNA and electrostatic interactions between DNA and histones are increased. Such parameters come within the values shown by adult rats.     

Biotin as a regulator of some haematic parameters and of DNA-content of the liver of old rats

Biotin administration to old rats (28 months) causes in the blood an increase of ATP, glucose, triglycerides, alkaline phosphatase and a decrease of cholesterol and acid phosphatase; in the liver DNA and electrostatic interactions between DNA and histones are increased. Such parameters come within the values shown by adult rats.     

Bridging the molecular and biological functions of the oxysterol-binding protein family

Oxysterol-binding protein (OSBP) and OSBP-related proteins (ORPs) constitute a large eukaryotic gene family that transports and regulates the metabolism of sterols and phospholipids. The original classification of the family based on oxysterol-binding activity belies the complex dual lipid-binding specificity of the conserved OSBP homology domain (OHD). Additional protein- and membrane-interacting modules mediate the targeting of select OSBP/ORPs to membrane contact sites between organelles, thus positioning the OHD between opposing membranes for lipid transfer and metabolic regulation. This unique subcellular location, coupled with diverse ligand preferences and tissue distribution, has identified OSBP/ORPs as key arbiters of membrane composition and function. Here, we will review how molecular models of OSBP/ORP-mediated intracellular lipid transport and regulation at membrane contact sites relate to their emerging roles in cellular and organismal functions.     

Diversity and specificity of the mitogen-activated protein kinase phosphatase-1 functions

The balance of protein phosphorylation is achieved through the actions of a family of protein serine/threonine kinases called the mitogen-activated protein kinases (MAPKs). The propagation of MAPK signals is attenuated through the actions of the MAPK phosphatases (MKPs). The MKPs specifically inactivate the MAPKs by direct dephosphorylation. The archetypal MKP family member, MKP-1 has garnered much of the attention amongst its ten other MKP family members. Initially viewed to play a redundant role in the control of MAPK signaling, it is now clear that MKP-1 exerts profound regulatory functions on the immune, metabolic, musculoskeletal and nervous systems. This review focuses on the physiological functions of MKP-1 that have been revealed using mouse genetic approaches. The implications from studies using MKP-1-deficient mice to uncover the role of MKP-1 in disease will be discussed.     

The nucleotide receptor P2Y13 is a key regulator of hepatic High-Density Lipoprotein (HDL) endocytosis

Cell surface receptors for high-density lipoprotein (HDL) on hepatocytes are major partners in the regulation of cholesterol homeostasis. We recently identified a cell surface ATP synthase as a high-affinity receptor for HDL apolipoprotein A-I (apoA-I) on human hepatocytes. Stimulation of this ectopic ATP synthase by apoA-I triggered a low-affinity-receptor-dependent HDL endocytosis by a mechanism strictly related to the generation of ADP. This suggests that nucleotide G-protein- coupled receptors of the P2Y family are molecular components in this pathway. Only P2Y₁ and P2Y₁₃ are present on the membrane of hepatocytes. Using both a pharmacological approach and small interference RNA, we identified P2Y₁₃ as the main partner in hepatic HDL endocytosis, in cultured cells as well as in situ in perfused mouse livers. We also found a new important action of the antithrombotic agent AR-C69931MX as a strong activator of P2Y₁₃-mediated HDL endocytosis. Received 9 May 2005; received after revision 24 June 2005; accepted 1 September 2005     

Effect of spironolactone and phenobarbital on the plasma fibrinogen in rats

Résumé Par un dosage de 20 M/100 g poids de spironolactone ou de phénobarbital pendant 3 jours, un niveau de fibrine élevé peut être produit chez des rats. En même temps, l'incorporation des aminoacides C¹⁴ au fibrinogène est aussi intense. Le phénobarbital est dans tous les cas plus efficace que le spironolactone. Dans notre cas, ces médicaments, connus comme inducteurs des enzymes microsomiques, augmentaient la synthèse protéique, enzymatiquement inactive du foie.     

Bacterial signal peptide recognizes HeLa cell mitochondrial import receptors and functions as a mitochondrial leader sequence

Phage display was used to identify new components of the mammalian mitochondrial receptor complex using Tom20 as a binding partner. Two peptides were identified. One had partial identity (SMLTVMA) with a bacterial signal peptide from Toho-1, a periplasmic protein. The other had partial identity with a mitochondrial inner membrane glutamate carrier. The bacterial signal peptide could carry a protein into mitochondria both in vivo and in vitro. The first six residues of the sequence, SMLTVM, were necessary for import but the two adjacent arginine residues in the 30-amino-acid leader were not critical for import. The signal peptides of Escherichia coli β-lactamase and Bacillsus subtilis lipase could not carry proteins into mitochondria. Presumably, the Toho-1 leader can adopt a structure compatible for recognition by the import apparatus.Received 29 April 2005; received after revision 8 June 2005; accepted 17 June 2005