HuFASEBJ'13.pdf

Obscurins comprise a family of giant
(870- to 600-kDa) and small (250- to 55-kDa) proteins
that play important roles in myofibrillogenesis,
cytoskeletal organization, and cell adhesion and are
implicated in hypertrophic cardiomyopathy and tumorigenesis.
Giant obscurins are composed of tandem
structural and signaling motifs, including 2 serine/
threonine kinase domains, SK1 and SK2, present at the
COOH terminus of giant obscurin-B. Using biochemical
and cellular approaches, we show for the first time
that both SK1 and SK2 possess enzymatic activities and
undergo autophosphorylation. SK2 can phosphorylate
the cytoplasmic domain of N-cadherin, a major component
of adherens junctions, and SK1 can interact with
the extracellular domain of the 1-subunit of the Na/
K-ATPase, which also resides in adherens junctions.
Immunostaining of nonpermeabilized myofibers and
cardiocytes revealed that some obscurin kinase isoforms
localize extracellularly. Quantification of the
exofacial expression of obscurin kinase proteins indicated
that they occupy 16 and 5% of the sarcolemmal
surface in myofibers and cardiocytes, respectively.
Treatment of heart lysates with peptide-N-glycosidase F
revealed that while giant obscurin-B localizes intracellularly,
possessing dual kinase activity, a small obscurin
kinase isoform that contains SK1 localizes extracellularly,
where it undergoes N-glycosylation. Collectively,
our studies demonstrate that the obscurin kinase domains
are enzymatically active and may be involved in
the regulation of cell adhesion.