Fbxl22, A Cardiac-Enriched F-Box Protein, Regulates Sarcomeric Protein Turnover and is Essential for Maintenance of Contractile Function In Vivo

Spaich, S., Will, R.D., Just, S., Spaich, S., Kuhn, C., Frank, D., Berger, I., Wiemann, S., Korn, B., Koegl, M., Backs, J., Katus, H.A., Rottbauer, W., and Frey, N.
Circulation research   111(12): 1504-1516 (Journal)
Registered Authors
Berger, Ina, Just, Steffen, Rottbauer, Wolfgang
cardiac myocytes, cardiomyopathy, proteasome, ubiquitin
MeSH Terms
  • Amino Acid Sequence
  • Animals
  • Animals, Newborn
  • Cells, Cultured
  • F-Box Proteins/physiology*
  • HEK293 Cells
  • Humans
  • Molecular Sequence Data
  • Myocardial Contraction/physiology*
  • Myocytes, Cardiac/metabolism*
  • Myocytes, Cardiac/physiology
  • Protein Transport/physiology
  • Rats
  • Receptors, Cytoplasmic and Nuclear/physiology*
  • Sarcomeres/metabolism*
  • Sarcomeres/physiology
22972877 Full text @ Circ. Res.

Rationale: 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.

Methods and 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.

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