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C. elegans asymmetric cell division

Temperature control of asymmetric cell division in C. elegans

An example using CherryTemp™ for fast and accurate temperature switch during microscopic observation.

Assymetric cell division in C.elegans

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With its perfectly documented cell-lineage development, C. elegans is a model of choice to study asymmetric cell division. Asymmetric division occurs when one cell gives rise to two daughter cells with distinct cellular fate, function and, sometimes, different size. Asymmetric division is found in many organisms: from yeast, bacteria, fly to higher vertebrates. This process has been extensively studied, as it is fundamental to generate cell lineage diversity in the embryo.

 

Asymmetric cell division in C. elegans embryo

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A succession of asymmetric divisions in C. elegans ensures the establishment of the organism anterior/posterior, dorso/ventral and left/right axes. In a very nice review for the wormbook, Gönczy and Rose detailed the cellular events leading to the first embryo asymmetric division. At fertilization, the sperm centrosome is pushed towards one side of the embryo, this will define the position of the first mitotic spindle and the division site. At the end of the first division, anterior/posterior axis of the embryo is organized. The symmetric embryo P0 is now partitioned into an anterior larger blastomere AB and a posterior smaller one P1. During the second division EMS and Abp define the ventral and dorsal side of the embryo, respectively. Finally, at the end of the third division, the left-right asymmetry is set up with ABal and ABpl.

The positioning of the sperm centrosome before the first division requires an interaction with the cell cortical acto-myosin network. This network is also critical for the positioning of PAR proteins. PAR proteins play crucial role in the determination of cell polarity and C. elegans PAR mutants showed loss of anterior-posterior polarity along with defective cell fate destiny. Before the embryo first division, PAR proteins are equally distributed at the cell cortex. Hyman’s group showed that the repositioning of PAR proteins in the embryo is facilitated by flows of the cortical acto-myosin network (Goehring et al., 2011).  For an extensive review on cell polarity and asymmetric division see (Cowan and Hyman, 2004).

 

How does asymmetric cell division take place?

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In his review (Li, 2013), R.Li explains that there are three requirements for asymmetric division to take place: segregation of cell fate determinant (RNA transcripts, cell polarity proteins), chromosome repartition in the two daughter cells, and cell division itself. Before division, cell determinants factors are unequally localized to the cell poles of the mother cell establishing cell polarization. Of the polarized poles will depend spindle orientation and the orientation of the cell plane division (see figure). When cytokinesis occurs, these cell determinants are inherited by only one of the two daughter cells, this one will then adopt a distinct cellular fate from its mother. For an extensive review on asymmetric cell division see Rose and Gönczy, 2013.

 

Experimental temperature control of asymmetric division in C. elegans development

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C. elegans embryos are immotile, have large cells and the existence of numerous temperature-sensitive mutant make them particularly suitable for combining protein activation/inactivation study and live-cell imaging. JG White’s group used a genetic screen to identify new temperature-sensitive cell-division mutant. These mutants were defective for spindle-orientation, cytokinesis, or had abnormal microtubules organization, all of these cellular processes affect asymmetric cell division (O’connell et al., 1998). More specifically, temperature-sensitive gpa-16 mutants have been shown to be defective in Galfa protein, important for spindle orientation in EMS cells and for left-right axes implementation.

 

Experimental temperature control of asymmetric division in C. elegans development

temperature-control-asymmetric-c-elegans

The use of temperature-sensitive mutants requires shifting C. elegans from a permissive to a restrictive temperature. This is usually done by incubating worms at the desire temperature. While as a genetic tool it is very powerful, in practice it is not possible to do rapid temperature-shift and follow the downstream effects with live-cell imaging. CherryTemp is a temperature controller, which allows you to shift worms temperature (5-45C) in seconds while acquiring microscope images. If you are using cell polarity or cell division temperature-sensitive mutants and you want to know what happens at the subcellular level right after you switch from a permissive to a restrictive temperature, then CherryTemp is what you need. By tightly increasing or decreasing your worm temperature you can generate hypomorhic-controlled activity of your temperature-sensitive protein. CherryTemp is robust, easy to use and it fits within any microscope settings

Reference

R. Li ,The Art of Choreographing Asymmetric Cell Division, Dev cell, 2013

LS. Rose  P. Gönczy Asymmetric cell division and axis formation in the embryo , wormbook 2013

NW Goehring, PK Trong, JS Bois,  D Chowdhury, EM Nicola , AA Hyman, Grill SW.Polarization of PAR proteins by advective triggering of a pattern-forming system, Science, 2011

CR.Cowan, AA. Hyman. Asymmetric cell division in C. elegans: cortical polarity and spindle positioning. Annu Rev Cell Dev Biol. 2004.

KF. O’Connell, CM. Leys and JG. White A Genetic Screen for Temperature-Sensitive Cell-Division Mutants of Caenorhabditis elegans genetics 1998

Tsou MF, Ku W, Hayashi A, Rose LS. PAR-dependent and geometry-dependent mechanisms of spindle positioning. J Cell Biol. 2003

S. Ben-Aroya, X. Pan, JD. Boeke, and P. Hieter, Making temperature-sensitive mutants, Methods Enzymol. 2010


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