Interaction of alpha-actinin-4 with class I PxxP motif-containing OK/SW-CL.16 protein.
BACKGROUND
The aim of this study was to identify novel binding partners for alpha-actinin-4 (actinin-4), an essential component of the glomerular filtration barrier.


METHODS
We performed bacterial two-hybrid screenings of a human kidney cDNA library using as a bait human actinin-4 containing the spectrin-like repeat R1. The identified interactions were further verified by in vitro affinity assays. To investigate the expression of the identified molecules in the kidney and other tissues, a human tissue cDNA panel screening and in situ hybridization were performed.


RESULTS
One isolated cDNA from the library encoded OK/SW-CL.16 protein with a segment similar to known actinin-interacting regions. OK/SW-CL.16 protein also contained the class I PxxP motif, which can participate in binding to Src homology 3 (SH3) domain-containing signaling proteins. In vitro affinity assays showed interactions of recombinant actinin-4 R1 and full-length renal actinin-4 with recombinant OK/SW-CL.16 protein. A tissue cDNA panel screening revealed a ubiquitous expression of OK/SW-CL.16 mRNA. In situ hybridization showed glomerular expression of OK/SW-CL.16 mRNA, mainly in podocytes and mesangial cells.


CONCLUSIONS
Our results suggest that OK/SW-CL.16 protein is a novel binding partner for actinin-4. OK/SW-CL16 protein might act as a linker between actinin-4 and some SH3 domain-containing signaling proteins in podocytes.
Togashi
Masaru
M
Third Department of Internal Medicine, Akita University School of Medicine, Akita, Japan.
Okuyama
Shin
S
Wakui
Hideki
H
Komatsuda
Atsushi
A
Suzuki
Daisuke
D
Toyoda
Masao
M
Sawada
Ken-ichi
K
D000185:Actinin / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D015723:Gene Library; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D050199:Podocytes / Q000737:chemistry* / Q000378:metabolism*; D011392:Proline / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000235:genetics
C111453:ACTN4 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008840:Microfilament Proteins; C524808:OK-SK-CL.16 protein, human; D000185:Actinin; D011392:Proline