title: F-box and leucine-rich repeat protein 22 is a cardiac-enriched F-box protein that regulates sarcomeric protein turnover and is essential for maintenance of contractile function in vivo.
abstract: BACKGROUND
  The emerging role of the ubiquitin-proteasome system in cardiomyocyte function and homeostasis implies the necessity of tight regulation of protein degradation. However, little is known about cardiac components of this machinery.
  
  
  OBJECTIVE
  We sought to determine whether molecules exist that control turnover of cardiac-specific proteins.
  
  
  RESULTS
  Using a bioinformatic approach to identify novel cardiac-enriched sarcomere proteins, we identified F-box and leucine-rich repeat protein 22 (Fbxl22). Tissue-specific expression was confirmed by multiple tissue Northern and Western Blot analyses as well as quantitative reverse-transcriptase polymerase chain reaction on a human cDNA library. Immunocolocalization experiments in neonatal and adult rat ventricular cardiomyocytes as well as murine heart tissue located Fbxl22 to the sarcomeric z-disc. To detect cardiac protein interaction partners, we performed a yeast 2-hybrid screen using Fbxl22 as bait. Coimmunoprecipitation confirmed the identified interactions of Fbxl22 with S-phase kinase-associated protein 1 and Cullin1, 2 critical components of SCF (Skp1/Cul1/F-box) E3- ligases. Moreover, we identified several potential substrates, including the z-disc proteins α-actinin and filamin C. Consistently, in vitro overexpression of Fbxl22-mediated degradation of both substrates in a dose-dependent fashion, whereas proteasome inhibition with MG-132 markedly attenuated degradation of both α-actinin and filamin C. Finally, targeted knockdown of Fbxl22 in rat cardiomyocytes as well as zebrafish embryos results in the accumulation of α-actinin associated with severely impaired contractile function and cardiomyopathy in vivo.
  
  
  CONCLUSIONS
  These findings reveal the previously uncharacterized cardiac-specific F-box protein Fbxl22 as a component of a novel cardiac E3 ligase. Fbxl22 promotes the proteasome-dependent degradation of key sarcomeric proteins, such as α-actinin and filamin C, and is essential for maintenance of normal contractile function in vivo.
authors.affiliation: Department of Internal Medicine III, University of Heidelberg, Heidelberg, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Spaich
authors.forename: Rainer D
authors.initials: RD
authors.lastname: Will
authors.forename: Steffen
authors.initials: S
authors.lastname: Just
authors.forename: Saskia
authors.initials: S
authors.lastname: Spaich
authors.forename: Christian
authors.initials: C
authors.lastname: Kuhn
authors.forename: Derk
authors.initials: D
authors.lastname: Frank
authors.forename: Ina M
authors.initials: IM
authors.lastname: Berger
authors.forename: Stefan
authors.initials: S
authors.lastname: Wiemann
authors.forename: Bernhard
authors.initials: B
authors.lastname: Korn
authors.forename: Manfred
authors.initials: M
authors.lastname: Koegl
authors.forename: Johannes
authors.initials: J
authors.lastname: Backs
authors.forename: Hugo A
authors.initials: HA
authors.lastname: Katus
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Rottbauer
authors.forename: Norbert
authors.initials: N
authors.lastname: Frey
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000831:Animals, Newborn; D002478:Cells, Cultured; D044783:F-Box Proteins / Q000502:physiology*; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000378:metabolism* / Q000502:physiology; D021381:Protein Transport / Q000502:physiology; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000502:physiology*; D012518:Sarcomeres / Q000378:metabolism* / Q000502:physiology
chemical_list: D044783:F-Box Proteins; C561886:FBXO22 protein, human; C578029:Fbox22 protein, mouse; D018160:Receptors, Cytoplasmic and Nuclear