title: Protein phosphorylation is a key event of flagellar disassembly revealed by analysis of flagellar phosphoproteins during flagellar shortening in Chlamydomonas.
abstract: Cilia are disassembled prior to cell division, which is proposed to regulate proper cell cycle progression. The signaling pathways that regulate cilia disassembly are not well-understood. Recent biochemical and genetic data demonstrate that protein phosphorylation plays important roles in cilia disassembly. Here, we analyzed the phosphoproteins in the membrane/matrix fraction of flagella undergoing shortening as well as flagella from steady state cells of Chlamydomonas. The phosphopeptides were enriched by a combination of IMAC and titanium dioxide chromatography with a strategy of sequential elution from IMAC (SIMAC) and analyzed by tandem mass spectrometry. A total of 224 phosphoproteins derived from 1296 spectral counts of phosphopeptides were identified. Among the identified phosphoproteins are flagellar motility proteins such as outer dynein arm, intraflagellar transport proteins as well as signaling molecules including protein kinases, phosphatases, G proteins, and ion channels. Eighty-nine of these phosphoproteins were only detected in shortening flagella, whereas 29 were solely in flagella of steady growing cells, indicating dramatic changes of protein phosphorylation during flagellar shortening. Our data indicates that protein phosphorylation is a key event in flagellar disassembly, and paves the way for further study of flagellar assembly and disassembly controlled by protein phosphorylation.
authors.affiliation: Protein Science Laboratory of Ministry of Education, School of Life Sciences, Tsinghua University, Beijing 100084, China. panjunmin@tsinghua.edu.cn
authors.forename: Junmin
authors.initials: J
authors.lastname: Pan
authors.forename: Bianca
authors.initials: B
authors.lastname: Naumann-Busch
authors.forename: Liang
authors.initials: L
authors.lastname: Wang
authors.forename: Michael
authors.initials: M
authors.lastname: Specht
authors.forename: Martin
authors.initials: M
authors.lastname: Scholz
authors.forename: Kerstin
authors.initials: K
authors.lastname: Trompelt
authors.forename: Michael
authors.initials: M
authors.lastname: Hippler
mesh_terms: D000595:Amino Acid Sequence; D015153:Blotting, Western; D016825:Chlamydomonas reinhardtii / Q000378:metabolism*; D002845:Chromatography / Q000379:methods; D005407:Flagella / Q000378:metabolism*; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D015398:Signal Transduction
chemical_list: D010750:Phosphoproteins