A specific interaction between muskelin and the cyclin-dependent kinase 5 activator p39 promotes peripheral localization of muskelin.
Previous studies implicate cyclin-dependent kinase 5 in cell adhesion and migration of epithelial cells of the cornea and lens. To explore molecular interactions underlying these functions, we performed yeast two-hybrid screening of an embryonic rat lens library for proteins that interact with cyclin-dependent kinase 5 and its regulators, p35 and p39. This screen identified a specific interaction between p39 and muskelin, an intracellular protein known to affect cytoskeletal organization in adherent cells. Immunohistochemistry detected muskelin in the developing lens and in other tissues, including brain and muscle. Glutathione S-transferase pull-down experiments and co-immunoprecipitations confirmed the specificity of the p39-muskelin interaction. Deletion analysis of p39 showed that muskelin binds to the p39 C terminus, which contains a short insertion (amino acids 329-366) absent from p35. Similar analysis of muskelin mapped the interaction with p39 to the fifth kelch repeat. Co-expression of p39 and muskelin in COS1 cells or lens epithelial cells altered the intracellular localization of muskelin, recruiting it to the cell periphery. These findings demonstrate a novel interaction between muskelin and the cyclin-dependent kinase 5 activator p39 and suggest that p39 may regulate the subcellular localization of muskelin.
Ledee
Dolena R
DR
NEI, National Institutes of Health, Bethesda, Maryland 20892-0704, USA.
Gao
Chun Y
CY
Seth
Ranjana
R
Fariss
Robert N
RN
Tripathi
Brajendra K
BK
Zelenka
Peggy S
PS
D000199:Actins / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D019556:COS Cells; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules; D002460:Cell Line; D002465:Cell Movement; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D017353:Gene Deletion; D015723:Gene Library; D005982:Glutathione Transferase / Q000378:metabolism; D015151:Immunoblotting; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins; D007908:Lens, Crystalline / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009132:Muscles / Q000378:metabolism; D009841:Oligonucleotides / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D010770:Phosphotransferases / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism*; D012313:RNA / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014162:Transfection; D020798:Two-Hybrid System Techniques
D000199:Actins; C492236:Cdk5r1 protein, mouse; D015815:Cell Adhesion Molecules; D018076:DNA, Complementary; D047908:Intracellular Signaling Peptides and Proteins; C503020:Mkln1 protein, mouse; C503021:Mkln1 protein, rat; D009841:Oligonucleotides; D010455:Peptides; D011506:Proteins; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D012313:RNA; D005982:Glutathione Transferase; D010770:Phosphotransferases; D051360:Cyclin-Dependent Kinase 5; C495907:Cdk5 protein, rat; D018844:Cyclin-Dependent Kinases