A regulatory light chain of ciliary outer arm dynein in Tetrahymena thermophila.
Ciliary beat frequency is primarily regulated by outer arm dyneins (22 S dynein). Chilcote and Johnson (Chilcote, T. J., and Johnson, K. A. (1990) J. Biol. Chem. 256, 17257-17266) previously studied isolated Tetrahymena 22 S dynein, identifying a protein p34, which showed cAMP-dependent phosphorylation. Here, we characterize the molecular biochemistry of p34 further, demonstrating that it is the functional ortholog of the 22 S dynein regulatory light chain, p29, in Paramecium. p34, thiophosphorylated in isolated axonemes in the presence of cAMP, co-purified with 22 S dynein and not with inner arm dynein (14 S dynein). Isolated 22 S dynein containing phosphorylated p34 showed approximately 70% increase in in vitro microtubule translocation velocity compared with its unphosphorylated counterpart. Extracted p34 rebound to isolated 22 S dynein from either Tetrahymena or Paramecium but not to 14 S dynein from either ciliate. Binding of radiolabeled p34 to 22 S dynein was competitive with p29. Phosphorylated p34 was not present in axonemes isolated from a mutant lacking outer arms. Two-dimensional gel electrophoresis followed by phosphorimaging revealed at least five phosphorylated p34-related spots, consistent with multiple phosphorylation sites in p34 or perhaps multiple isoforms of p34. These new features suggest that a class of outer arm dynein light chains including p34 regulates microtubule sliding velocity and consequently ciliary beat frequency through phosphorylation.
Christensen
S T
ST
Departments of Anatomy and Structural Biology and Developmental and Molecular Biology, Albert Einstein College of Medicine of Yeshiva University, Bronx, New York 10461, USA.
Guerra
C
C
Wada
Y
Y
Valentin
T
T
Angeletti
R H
RH
Satir
P
P
Hamasaki
T
T
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002923:Cilia / Q000378:metabolism*; D000242:Cyclic AMP / Q000494:pharmacology; D004398:Dyneins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D010766:Phosphorylation; D016808:Tetrahymena thermophila / Q000187:drug effects / Q000378:metabolism*
D000242:Cyclic AMP; D004398:Dyneins; D002118:Calcium