Phosphatase inhibitor 2 promotes acetylation of tubulin in the primary cilium of human retinal epithelial cells.
BACKGROUND
Primary cilia are flagella-like projections from the centriole of mammalian cells that have a key role in cell signaling. Human diseases are linked to defects in primary cilia. Microtubules make up the axoneme of cilia and are selectively acetylated and this is thought to contribute to the stability of the structure. However, mechanisms to regulate tubulin acetylation in cilia are poorly understood.


RESULTS
Endogenous phosphatase inhibitor-2 (I-2) was found concentrated in cilia of human epithelial cells, and was localized to cilia early in the process of formation, prior to the full acetylation of microtubules. Knockdown of I-2 by siRNA significantly reduced the acetylation of microtubules in cilia, without a net decrease in whole cell tubulin acetylation. There was a reduction in the percentage of I-2 knockdown cells with a primary cilium, but no apparent alteration in the cilium length, suggesting no change in microtubule-based transport processes. Inhibition of either histone deacetylases with trichostatin A, or protein phosphatase-1 with calyculin A in I-2 knockdown cells partially rescued the acetylation of microtubules in cilia and the percentage of cells with a primary cilium.


CONCLUSIONS
The regulatory protein I-2 localizes to the primary cilium where it affects both Ser/Thr phosphorylation and is required for full tubulin acetylation. Rescue of tubulin acetylation in I-2 knockdown cells by different chemical inhibitors shows that deacetylases and phosphatases are functionally interconnected to regulate microtubules. As a multifunctional protein, I-2 may link cell cycle progression to structure and stability of the primary cilium.
Wang
Weiping
W
Center for Cell Signaling, University of Virginia, School of Medicine, Charlottesville, 22908, USA.
Brautigan
David L
DL
D000107:Acetylation; D002460:Cell Line; D002923:Cilia / Q000378:metabolism* / Q000648:ultrastructure; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D055785:Gene Knockdown Techniques; D056572:Histone Deacetylase Inhibitors; D006655:Histone Deacetylases / Q000378:metabolism; D006801:Humans; D008870:Microtubules / Q000737:chemistry / Q000378:metabolism; D010641:Phenotype; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D012160:Retina / Q000166:cytology*; D014404:Tubulin / Q000737:chemistry / Q000378:metabolism*; D000097847:Protein Phosphatase Inhibitory Proteins
D056572:Histone Deacetylase Inhibitors; D006655:Histone Deacetylases; D011506:Proteins; D034741:RNA, Small Interfering; D014404:Tubulin; C024833:protein phosphatase inhibitor-2; D000097847:Protein Phosphatase Inhibitory Proteins