Changes in mRNA expression profile underlie phenotypic adaptations in creatine kinase‐deficient muscles

Abstract

We have studied the mechanisms that regulate the remodeling of the glycolytic, mitochondrial and structural network of muscles of creatine kinase M (M-CK)/sarcomeric mitochondrial creatine kinase (ScCKmit) knockout mice by comparison of wild-type and mutant mRNA profiles on cDNA arrays. The magnitudes of changes in mRNA levels were most prominent in M-CK/ScCKmit (CK 3a3 ) double mutants but did never exceed those of previously observed changes in protein level for any protein examined. In gastrocnemius of CK 3a3 mice we measured a 2.5-fold increase in mRNA level for mitochondrial encoded cytochrome c oxidase (COX)-III which corresponds to the increase in protein content. The level of the nuclear encoded mRNAs for COX-IV, H + -ATP synthase-C, adenine nucleotide translocator-1 and insulin-regulatable glucose transporter-4 showed a 1.5-fold increase, also in agreement with protein data. In contrast, no concomitant up-regulation in mRNA and protein content was detected for the mitochondrial inorganic phosphatecarrier, voltage-dependent anion channel and certain glycolytic enzymes. Our results reveal that regulation of transcript level plays an important role, but it is not the only principle involved in the remodeling of mitochondrial and cytosolic design of CK 3a3 muscles. ß

Adenylate Kinase 1 Deficiency Induces Molecular and Structural Adaptations to Support Muscle Energy Metabolism

“…Intriguingly, other mitochondrial import proteins like the PiC and VDAC were not increased. In this regard muscles of M-CK knockout mice have a similar pattern in molecular and cytoarchitectural adaptations (24,25). In these mutants, ScCKmit and ANT1 mRNA levels were maintained whereas ScCKmit and ANT protein levels were dramatically increased.…”

“…Macroarrays were prepared by spotting individual plasmids with cDNA insert (134 different mouse sequences for glycolytic enzymes, mitochondrial enzymes and transporters, fatty acid metabolism enzymes, muscle regulatory factors, proteins active in Ca 2ϩ signaling, glucose transport, tissue oxygenation as well as cytoskeletal components and transcription factors (see www.ncmls.kun. nl/celbio/data.htm for detailed information) onto Hybond N ϩ membranes using a gridding robot (24). 32 P-labeled single-stranded cDNA was used as probe to hybridize membranes with gridded cDNA arrays as previously described in detail elsewhere (24).…”

“…nl/celbio/data.htm for detailed information) onto Hybond N ϩ membranes using a gridding robot (24). 32 P-labeled single-stranded cDNA was used as probe to hybridize membranes with gridded cDNA arrays as previously described in detail elsewhere (24). Four AK1 knockout and control mice were analyzed and statistically compared.…”

Impaired Intracellular Energetic Communication in Muscles from Creatine Kinase and Adenylate Kinase (M-CK/AK1) Double Knock-out Mice

“…As ultra-fast local and global ATP regeneration is required to sustain contractile activity in fast-twitch muscle (24,52), this would underscore the particular importance of CK-and AK-catalyzed phosphotransfer in these muscle types. Moreover, mice carrying a single mutation in the AK1 or M-CK gene show more pronounced adaptations at the molecular and architectural level in their fast-rather than slow-twitch fibers (24,(37)(38)(39). Thus, tight coordination between cellular sites of ATP consumption and ATP generation by the complete set of high energy phosphoryl transfer pathways would not only be essential for safeguarding cellular energetic economy (6,33,54,65) but also for preserving the differential physiological contractile characteristics of slow-and fast-twitch muscles.…”

“…Likewise, in skeletal muscles carrying a null mutation in either the M-CK or AK1 gene, leading to complete lack of corresponding protein expression and activity, an adaptive rewiring of flux through the remaining intact phosphotransfer circuit occurs (6,18,19). In addition, M-CK and AK1 mutant muscles respond with similar but not identical ultrastructural and molecular adaptations, suggesting an inherent plasticity of the bioenergetic network (6,24,(37)(38)(39).…”