Mucolipin 1 channel activity is regulated by protein kinase A-mediated phosphorylation.
Mucolipins constitute a family of cation channels with homology with the transient receptor potential family. Mutations in MCOLN1 (mucolipin 1) have been linked to mucolipidosis type IV, a recessive lysosomal storage disease characterized by severe neurological and ophthalmologic abnormalities. At present, little is known about the mechanisms that regulate MCOLN1 activity. In the present paper, we addressed whether MCOLN1 activity is regulated by phosphorylation. We identified two PKA (protein kinase A) consensus motifs in the C-terminal tail of MCOLN1, containing Ser(557) and Ser(559). Ser(557) was the principal phosphorylation site, as mutation of this residue to alanine caused a greater than 75% reduction in the total levels of phosphorylated MCOLN1 C-terminal tail. Activation of PKA with forskolin promoted MCOLN1 phosphorylation, both in vitro and in vivo. In contrast, addition of the PKA inhibitor H89 abolished MCOLN1 phosphorylation. We also found that PKA-mediated phosphorylation regulates MCOLN1 channel activity. Forskolin treatment decreased MCOLN1 channel activity, whereas treatment with H89 increased MCOLN1 channel activity. The stimulatory effect of H89 on MCOLN1 function was not observed when Ser(557) and Ser(559) were mutated to alanine residues, indicating that these two residues are essential for PKA-mediated negative regulation of MCOLN1. This paper presents the first example of regulation of a member of the mucolipin family by phosphorylation.
Vergarajauregui
Silvia
S
Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Oberdick
Ross
R
Kiselyov
Kirill
K
Puertollano
Rosa
R
D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004594:Electrophysiology; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000196:embryology / Q000502:physiology; D007700:Kinetics; D008247:Lysosomes; D009081:Mucolipidoses / Q000235:genetics; D009154:Mutation; D010766:Phosphorylation; D010957:Plasmids; D011994:Recombinant Proteins / Q000378:metabolism; D050053:TRPM Cation Channels / Q000235:genetics / Q000378:metabolism*; D050051:Transient Receptor Potential Channels
C416674:MCOLN1 protein, human; D011994:Recombinant Proteins; D050053:TRPM Cation Channels; D050051:Transient Receptor Potential Channels; D005982:Glutathione Transferase; D017868:Cyclic AMP-Dependent Protein Kinases