1. Multiple Wnt Signaling Pathways Converge to Orient the Mitotic Spindle in Early C. elegans Embryos Walston T. et al. Developmental Cell, Vol. 7, 831–841, December, 2004

2. ABpl The fate of the EMS daughters is controlled by a Wnt/b-cat pathway. The orientation of the EMS division is controlled by a different Wnt pathway involving Wnt(MOM-2), Porcupine(Porc;MOM-1), Fz(MOM-5), GSK-3(GSK-3b) and CK1  (KIN-19). A pathway involving MES-1, a receptor tyrosine kinase, and SRC-1, a Src family tyrosine kinase, acts redundantly with Wnt signaling with respect to the fate of EMS daughters and the orientation of the EMS spindle. mom-1 (Porc), mom-2 (Wnt), mom-5 (Fz), and mom-3 (uncloned), cause spindle alignment defects in the ABar blastomere of the 8-cell embryo. Background

4. Although many Wnt signaling components have been identified that participate in spindle orientation, the role of the Dsh family has not been clearly characterized. (dsh-1, dsh-2, mig-5) Loss of function of the CKI homolog, kin-19, causes defects in the fate of EMS daughter cells. Although the role of CKI in spindle alignment has not been examined, CKI localizes to centrosomes and mitotic spindles in vertebrate systems. The nontranscriptional Wnt spindle alignment pathway requires contact from the C blastomere to align the spindle of ABar. Wnt/b-catenin pathway regulates the timing of spindle rotation in ABar, presumably by specifying the fate of neighboring blastomeres. We demonstarate...

5. dsh-2(or302) dsh-1(RNAi); dsh-2(or302); mig-5(RNAi) Defects in Alignment of the EMS and ABar Spindles.

6. Defects in EMS

7. KIN-19/CK Ⅰ localizes to centrosomes and DSH-2 accumulates between P2 and EMS. microtubule condensed chromosome ~4cell stage4~6cell stage6~32cell stage cytoplasmboundary between P2 and EMS cortex of all cells Consistent with P2 signaling to EMS to specify endoderm fate and EMS spindle orientation. KIN-19 RNAi → does not affect Dsh-2 localization.

8. Fz APC Axin b-cat NEMO TCF RNA pol. Positive : Dsh, CK Ⅰ, GSK-3, Src (Dsh background) Negative : JNK, APC, Axin, b-cat, NEMO, TCF, RNA pol. Spindle defects in ABar.

9. Contact with the C blastomere aligns the spindle in ABar. Caudal homolog laser killed Mom-2(Wnt) Mom-5(Fz) canonicalnon-canonical C EMS ABar

10. ABar spindle defects visualized by  -tubulin::GFP dsh-2(RNAi); mig-5(RNAi)wrm-1(RNAi) TBB-2/  -tubulin::GFP

11. Three Wnt signaling pathways operate in the Early C. elegans embryos.

12. G  signaling in spindle orientation ?? GSK-3 in spindle orientation ?? Discussion

13. Heterotrimeric G-protein in spindle orientation G  : GPB-1, GPB-2 G  : GPC-1, GPC-2 Gotta M et al, Nat Cell Biol, 2001. A P D V ABa ABp P2 EMS

14. Heterotrimeric G-protein in spindle orientation C. elegans has 20G  genes. → GOA-1, GPA-16 Conclusion : G  signaling, not G , participates spindle orientation in C. elegans. Gotta M et al, Nat Cell Biol, 2001.

15. Cdc42 regulates GSK-3  and APC to control cell polarity. There is a larger complex containing GSK-3 , Par6, PKC . Scratch-induced cell migration assay : Cdc42, p-GSK-3  -cat and APC localize at the leading edge of migrating cell. Etienne-Manneville S et al. Nature, 2003. Astrocyte MTOC : microtubule organizing centre

16. Activation of G  signaling downstream of Wnt-11/Xfz7 regulates Cdc42 activity. Penzo-Mendez A et al. Dev Biol, 2003. During xenopus gastrulation

17. Cdc42, p-GSK-3   -cat, APC Leading edge Dvl-1 ? Dvl-2 ? Dvl-3 ? Src Canonical Wnt Fz Non-canonical Wnt Fz G  Dvl GSK-3  Axin APC  -cat CK Ⅰ  -cat Tcf/Lef Dvl RhoA MEKK, SEK JNK PKC Cdc42 Par-6 PKC  Polarity Cell fate, spindle rotation Mes-1