ZFIN ID: ZDB-PUB-091221-29
Mys protein regulates protein kinase A activity by interacting with regulatory type I{alpha} subunit during vertebrate development
Kotani, T., Iemura, S.I., Natsume, T., Kawakami, K., and Yamashita, M.
Date: 2010
Source: The Journal of biological chemistry   285(7): 5106-5116 (Journal)
Registered Authors: Kawakami, Koichi, Kotani, Tomoya
Keywords: DEVELOPMENT DIFFERENTIATION, GENETICS, PROTEIN/Protein-protein interactions, SIGNAL TRANSDUCTION, Cell differentiation, Protein kinase A (PKA), Hedgehog signaling, Vertebrate development, Zebrafish
MeSH Terms:
  • Animals
  • Cell Line
  • Colforsin/pharmacology
  • Cyclic AMP-Dependent Protein Kinases/genetics
  • Cyclic AMP-Dependent Protein Kinases/metabolism*
  • Embryo, Nonmammalian/metabolism
  • Humans
  • Immunoblotting
  • Immunoprecipitation
  • In Situ Hybridization
  • Mass Spectrometry
  • Oligonucleotides, Antisense/pharmacology
  • Protein Binding/drug effects
  • Veratrum Alkaloids/pharmacology
  • Zebrafish
  • Zebrafish Proteins/antagonists & inhibitors
  • Zebrafish Proteins/genetics
  • Zebrafish Proteins/metabolism*
PubMed: 20018846 Full text @ J. Biol. Chem.
During embryonic development, protein kinase A (PKA) plays a key role in cell fate specification by antagonizing Hedgehog (Hh) signaling pathway. However, the mechanism by which PKA activity is regulated remains unknown. Here we show that Misty somites (Mys) protein regulates the level of PKA activity during embryonic development in zebrafish. We isolate PKA regulatory type Ialpha subunit (Prkar1a) as a protein interacting with Mys by pull-down assay in HEK293 cells followed by mass-spectrometry analysis. We show an interaction between endogenous Mys and Prkar1a in the zebrafish embryo. Mys binds to Prkar1a in its C-terminus region, termed PRB domain, and activates PKA in vitro. Conversely, knockdown of Mys in zebrafish embryos results in reduction in PKA activity. We also show that knockdown of Mys induces ectopic activation of Hh target genes in the eyes, neural tube and somites downstream of Smoothened, a protein essential for transduction of Hh-signaling activity. The altered patterning of gene expression is rescued by activation of PKA. Together, our results reveal a molecular mechanism of regulation of PKA activity that is dependent on a protein-protein interaction and demonstrate that PKA activity regulated by Mys is indispensable for negative regulation of Hh signaling pathway in Hh-responsive cells.