Dataset Information

Knockdown of the C. elegans kinome identifies kinases required for normal protein homeostasis, mitochondrial network structure, and sarcomere structure in muscle.

ABSTRACT:

Background

Kinases are important signalling molecules for modulating cellular processes and major targets of drug discovery programs. However, functional information for roughly half the human kinome is lacking. We conducted three kinome wide, >90%, RNAi screens and epistasis testing of some identified kinases against known intramuscular signalling systems to increase the functional annotation of the C. elegans kinome and expand our understanding of kinome influence upon muscle protein degradation.

Results

96 kinases were identified as required for normal protein homeostasis, 74 for normal mitochondrial networks and 50 for normal sarcomere structure. Knockdown of kinases required only for normal protein homeostasis and/or mitochondrial structure was significantly less likely to produce a developmental or behavioural phenotype than knockdown of kinases required for normal sarcomere structure and/or other sub-cellular processes. Lastly, assessment of kinases for which knockdown produced muscle protein degradation against the known regulatory pathways in C. elegans muscle revealed that close to half of kinase knockdowns activated autophagy in a MAPK dependent fashion.

Conclusions

Roughly 40% of kinases studied, 159 of 397, are important in establishing or maintaining muscle cell health, with most required for both. For kinases where decreased expression triggers protein degradation, autophagy is most commonly activated. These results increase the annotation of the C. elegans kinome to roughly 75% and enable future kinome research. As 33% of kinases identified have orthologues expressed in human muscle, our results also enable testing of whether identified kinases function similarly in maintaining human muscle homeostasis.

SUBMITTER: Lehmann S

PROVIDER: S-EPMC3849176 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

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Publications

Knockdown of the C. elegans kinome identifies kinases required for normal protein homeostasis, mitochondrial network structure, and sarcomere structure in muscle.

Lehmann Susann S Bass Joseph J JJ Szewczyk Nathaniel J NJ

Cell communication and signaling : CCS 20130923

<h4>Background</h4>Kinases are important signalling molecules for modulating cellular processes and major targets of drug discovery programs. However, functional information for roughly half the human kinome is lacking. We conducted three kinome wide, >90%, RNAi screens and epistasis testing of some identified kinases against known intramuscular signalling systems to increase the functional annotation of the C. elegans kinome and expand our understanding of kinome influence upon muscle protein d ...[more]

PMID: 24060339

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Project description:BackgroundSkeletal muscle is central to locomotion and metabolic homeostasis. The laboratory worm Caenorhabditis elegans has been developed into a genomic model for assessing the genes and signals that regulate muscle development and protein degradation. Past work has identified a receptor tyrosine kinase signalling network that combinatorially controls autophagy, nerve signal to muscle to oppose proteasome-based degradation, and extracellular matrix-based signals that control calpain and caspase activation. The last two discoveries were enabled by following up results from a functional genomic screen of known regulators of muscle. Recently, a screen of the kinome requirement for muscle homeostasis identified roughly 40% of kinases as required for C. elegans muscle health; 80 have identified human orthologues and 53 are known to be expressed in skeletal muscle. To complement this kinome screen, here, we screen most of the phosphatases in C. elegans.MethodsRNA interference was used to knockdown phosphatase-encoding genes. Knockdown was first conducted during development with positive results also knocked down only in fully developed adult muscle. Protein homeostasis, mitochondrial structure, and sarcomere structure were assessed using transgenic reporter proteins. Genes identified as being required to prevent protein degradation were also knocked down in conditions that blocked proteasome or autophagic degradation. Genes identified as being required to prevent autophagic degradation were also assessed for autophagic vesicle accumulation using another transgenic reporter. Lastly, bioinformatics were used to look for overlap between kinases and phosphatases required for muscle homeostasis, and the prediction that one phosphatase was required to prevent mitogen-activated protein kinase activation was assessed by western blot.ResultsA little over half of all phosphatases are each required to prevent abnormal development or maintenance of muscle. Eighty-six of these phosphatases have known human orthologues, 57 of which are known to be expressed in human skeletal muscle. Of the phosphatases required to prevent abnormal muscle protein degradation, roughly half are required to prevent increased autophagy.ConclusionsA significant portion of both the kinome and phosphatome are required for establishing and maintaining C. elegans muscle health. Autophagy appears to be the most commonly triggered form of protein degradation in response to disruption of phosphorylation-based signalling. The results from these screens provide measurable phenotypes for analysing the combined contribution of kinases and phosphatases in a multi-cellular organism and suggest new potential regulators of human skeletal muscle for further analysis.

Dataset Information

Knockdown of the C. elegans kinome identifies kinases required for normal protein homeostasis, mitochondrial network structure, and sarcomere structure in muscle.

Background

Results

Conclusions

Publications

Knockdown of the C. elegans kinome identifies kinases required for normal protein homeostasis, mitochondrial network structure, and sarcomere structure in muscle.

Similar Datasets

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