Submitted on December 22, 2004
Accepted on March 7, 2005
A B56 Regulatory Subunit of Protein Phosphatase 2A Localizes to Nuclear Speckles in Cardiac Myocytes
Marisa S Gigena1, Akihito Ito2, Hiroshi Nojima3, and Terry B Rogers1*
1 Biochemistry and Molecualr Biology, University of Maryland School of Medicine, Baltimore, MD, USA
2 Pathology, Oskaka University Medical School, Osaka, Japan
3 Department for Molecular Genetics, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan
* To whom correspondence should be addressed. E-mail: trogers{at}som.umaryland.edu.
Protein phosphatase 2A (PP2A) is widely distributed in heart tissues yet its precise cellular functions are poorly understood. This study is based on the notion that PP2A action is governed by interactions of the core enzyme with B targeting/regulatory subunits. The subcellular localizations of two B subunits, B56
and B56
1 were assessed by adenoviral-driven expression of epitope-tagged (hemaglutinin, HA) in cultured neonatal and adult rat ventricular myocytes. Confocal imaging revealed that HA-B56 was excluded from the nucleus and decorated striated structures, while HA-B561 was principally found in the nucleus. Precise immunolabeling studies showed that B561 was concentrated in intranuclear structures known as nuclear speckles, macromolecular structures that accumulate transcription and splicing factors. Western blot analyses revealed that overexpression of either B subunit had no effect on the levels of other PP2A subunits in cultured neonatal cardiac cells. However, overexpression of only B561 increased whole-cell PP2A activity by 40% when measured in cell extracts. Finally, B561 did not alter global gene expression or expression of hypertrophic gene markers such as -skeletal actin. However, morphimetric analyses of confocal images revealed that B561 alters the dynamic assembly/disassembly process of nuclear speckles in heart cells. These studies provide new insight into mechanisms of PP2A targeting in the subnuclear architecture in cardiac myocytes and into the role of this phosphatase in nuclear signaling.
