A regulatory light chain of ciliary outer arm dynein in Tetrahymena thermophila

TY - JOUR

T1 - A regulatory light chain of ciliary outer arm dynein in Tetrahymena thermophila

AU - Christensen, Søren Tvorup

AU - Guerra, C

AU - Wada, Y

AU - Valentin, T

AU - Angeletti, R H

AU - Satir, P

AU - Hamasaki, T

N1 - Keywords: Animals; Calcium; Cilia; Cyclic AMP; Dynein ATPase; Electrophoresis, Polyacrylamide Gel; Phosphorylation; Tetrahymena thermophila

PY - 2001

Y1 - 2001

N2 - 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.

AB - 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.

U2 - 10.1074/jbc.M008412200

DO - 10.1074/jbc.M008412200

M3 - Journal article

C2 - 11274140

SN - 0021-9258

VL - 276

SP - 20048

EP - 20054

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 23

ER -