title: S6K1 is involved in polyploidization through its phosphorylation at Thr421/Ser424.
abstract: Studies on polyploidization of megakaryocytes have been hampered by the lack of synchronized polyploid megakaryocytes. In this study, a relatively synchronized polyploid cell model was successfully established by employing Dami cells treated with nocodazole. In nocodazole-induced cells, cyclin B expression oscillated normally as in diploid cells and polyploid megakaryocytes. By using the nocodazole-induced Dami cell model, we found that 4E-BP1 and Thr421/Ser424 of ribosomal S6 kinase 1(S6K1) were phosphorylated mostly at M-phase in cytoplasm and oscillated in nocodazole-induced polyploid Dami cells, concomitant with increased expression of p27 and cyclin D3. However, phosphorylation of 4E-BP1 and S6K1 on Thr421/Ser424 was significantly decreased in differentiated Dami cells induced by phorbol 12-myristate 13-acetate (PMA), concomitant with increased expression of cyclin D1 and p21 and cyclin D3. Overexpression of the kinase dead form of S6K1 containing the mutation Lys 100 --> Gln in PMA-induced Dami cells increased ploidy whereas overexpression of rapamycin-resistant form of S6K1 containing the mutations Thr421 --> Glu and Ser424 --> Asp significantly dephosphorylated 4E-BP1 and reduced expression of cyclin D1, cyclin D3, p21 and p27, and slightly decreased the ploidy of PMA-induced Dami cells, compared with treatment with PMA alone. Moreover, overexpression of rapamycin-resistant form of S6K1 significantly reversed polyploidization of nocodazole-induced Dami cells. Furthermore, MAP (a novel compound synthesized recently) partly blocked the phosphorylation of S6K1 on Thr421/Ser424 and decreased the expression of p27 and polyploidization in nocodazole-induced Dami cells. Taken together, these data suggested that S6K1/4E-BP1 pathway may play an important role in polyploidization of megakaryocytes.
authors.affiliation: Department of Experimental Medicine, Northern Hospital, Shenyang, Liaoning, China. mark_cdm@yahoo.com
authors.forename: Dongchu
authors.initials: D
authors.lastname: Ma
authors.forename: Huiying
authors.initials: H
authors.lastname: Yu
authors.forename: Di
authors.initials: D
authors.lastname: Lin
authors.forename: Yinghui
authors.initials: Y
authors.lastname: Sun
authors.forename: Liping
authors.initials: L
authors.lastname: Liu
authors.forename: Yage
authors.initials: Y
authors.lastname: Liu
authors.forename: Bing
authors.initials: B
authors.lastname: Dai
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Jianping
authors.initials: J
authors.lastname: Cao
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D002453:Cell Cycle / Q000502:physiology; D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor / Q000187:drug effects; D002867:Chromones / Q000494:pharmacology; D019926:Cyclin B / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008533:Megakaryocytes / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D009025:Morpholines / Q000494:pharmacology; D015739:Nocodazole / Q000494:pharmacology; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011123:Polyploidy; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D038744:Ribosomal Protein S6 Kinases, 90-kDa / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013912:Threonine / Q000378:metabolism*; D000077191:Wortmannin
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000730:Androstadienes; D000970:Antineoplastic Agents; D002273:Carcinogens; D018797:Cell Cycle Proteins; D002867:Chromones; D019926:Cyclin B; C087000:EIF4EBP1 protein, human; D004791:Enzyme Inhibitors; D009025:Morpholines; D010750:Phosphoproteins; D013912:Threonine; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D012694:Serine; D011494:Protein Kinases; C546842:MTOR protein, human; D038744:Ribosomal Protein S6 Kinases, 90-kDa; D058570:TOR Serine-Threonine Kinases; D013755:Tetradecanoylphorbol Acetate; D015739:Nocodazole; D020123:Sirolimus; D000077191:Wortmannin