Spindle orientation

We have previously shown that a 209 amino acid region (aa288-497, asymmetric localization domain) of Insc is necessary and sufficient for apical cortical localization and for mitotic spindle orientation along the apical-basal axis (Tio et al 1999). In a yeast two-hybrid screen we identified Partner of Inscuteable (Pins), a novel 658aa protein with multiple repeats of the Tetratricopeptide (TPR) motif. Affinity purification experiments using embryonic extracts demonstrate that Pins complexes with Insc in vivo. In vitro protein interaction assays demonstrates that Pins interacts with the Insc asymmetric localization domain (see Yu et al 2000). 

Downstream events protein localization spindle orientation cell fate resolution... 

In Pins - embryos the initiation steps of apical complex formation occur normally. However, this complex cannot be maintained in mitotic neuroblasts. Hence, the importance of the maintenance of this complex for asymmetric cell division can be ascertained by assessing how Pins- neural progenitors divide. Pins- embryos exhibit all of the defects seen in insc mutants. Mitotic spindle orientation is defective. In the cells of mitotic domain 9 the 90° reorientation, which normally occurs in wild-type resulting in the orientation of the spindle along the apical—basal axis (Fig. 3A), fails to occur in the mutant (Fig. 3B). Mitotic spindle orientation of neuroblasts in the segmented CNS, deduced from DNA staining, also often fails to... 

Chia A certain proportion of the time Insc failed to localize and subsequently downstream events such as spindle orientation and Pros and Numb asymmetric localization were abnormal. 

Nurse Didn t someone suggest that cyclin A/Cdc2 activity which comes up earlier in the cell cycle might have a role in spindle orientation ... 

Chia We see a defect in spindle orientation when there is a failure of localization of the apical complex. This is also what is seen in the cdc2 alleles. 

Kuchinke U, Grawe F, Knust E 1998 Control of spindle orientation in Drosophila by the Par-3-related PDZ-domain protein Bazooka. Curr Biol 8 1357—1365... 

Cheng NN, Kirby CM, Kemphues KJ 1995 Control of cleavage spindle orientation in Caenorhabditis elegans-. the role of the genes par-2 and par-3. Genetics 139 549-559 Edgar LG, Wolf N, Wood WB 1994 Early transcription in Caenorhabditis elegans embryos. Development 120 443—451... 

Gonzalez C (2007) Spindle orientation, asymmetric division and tumour suppression in Drosophila stem cells. Nat Rev Genet 8 462-472... 

Lgl binds to myosin II, which functions in cytokinesis (see Figure 19-20). Lgl Itself is uniformly localized around the cortex. Lgl is phosphorylated by the apical complex and may be inactivated on the apical side to allow basal Miranda accumulation. Two yeast proteins related to Lgl also bind to myosin II they have been implicated in exocytosis and secretion, specifically in the docking of post-Golgi vesicles with the plasma membrane (Chapter 17). Mutations in the two yeast proteins suppress mutations in the myosin II gene. This observation is interpreted to mean that the function of myosin II in spindle orientation is opposite to that of the Lgl-like proteins reduction of one protein s function is ameliorated by reduction of the other, restoring a semblance of the normal balance. In this case, therefore, myosin and Lgl probably act in opposite directions myosin II moving Miranda or other materials to control spindle orientation and Lgl restraining It. 

Further evidence for the Importance of myosin In spindle orientation comes the finding that another myosin relative,... 

Asymmetry factors exert their influence at least in part by controlling the orientation of the mitotic spindle, so that asymmetrically localized proteins and structures are differentially incorporated into the two daughter cells. Myosin proteins bind to proteins that control asymmetry factors of cells to control spindle orientation. 

Petritsch, C., et al. 2003. The Drosophila myosin VI Jaguar is required for basal protein targeting and correct spindle orientation in mitotic neuroblasts. Devel. Ceil 4 273-281. 

Tabler, J.M. Yamanaka, H. Green, J.B. (2010). PAR-1 promotes primary neurogenesis and asymmetric cell divisions via control of spindle orientation. Development, Vol. 137(15), pp. 2501-2505.

McCudden, C.R. Firnberg, N. Barres, B.A. Siderovski, D.P. Knoblich, J.A. (2005). Mammalian inscuteable regulates spindle orientation and cell fate in the developing retina Neuron, Vol. 48(4), pp. 539-545. 

If we are to understand phenomena such as tissue growth, cell differentiation, and organ development, we must first understand how spindle orientation or direction is regulated during nuclear division. Furthermore, we must understand how the cell cycle is started and stopped. The elucidation of these processes can probably be most readily achieved by using initially homogeneous populations of cells grown in axenic culture because such populations are those that can be most readily manipulated. Theoretically, if the details of the biochemical... 

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