Determination of left-right patterning of the mouse

Determination of left-right patterning of the mouse -...

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“Randomization of left-right asymmetry  due to loss of nodal cilia generating  leftward flow of extraembryonic fluid in  mice lacking  KIF3B  motor protein” (Nonaka, S.  et al.  1998)  And   “Determination of left-right patterning  of the mouse embryo by artificial nodal  flow” (Nonaka, S.  et al.  2002) By Mark Keresztes & 
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Left-Right Asymmetry Review The consistent left-right asymmetry in  vertebrates is marked by the expression of Nodal,  lefty  and Pitx2 proteins on the left side of the  embryo. The asymmetry is generated by a variety of  mechanisms depending on the organism. Proton potassium pump in  Xenopus  and chicks Notch pathway activation and Nodal signaling The movement of cilia on the surface of the node in  humans and mice 
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Left-Right Asymmetry Review Important step in the development of many  organisms  Left-right asymmetry first becomes anatomically  apparent during heart looping Left-right asymmetry first becomes genetically  apparent during somitogenesis stage. Asymmetrical expression of  lefty1, lefty2, nodal, Pitx2
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Nonaka et al., 1998
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Hypothesis Nodal cilia play an important roll in L-R  determination To test their hypothesis, they mutated a gene in mice  that is important for nodal ciliogenesis
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Introduction Kinesin superfamily proteins (KIFs) are a molecular  motor superfamily  involved in the transportation  of proteins and lipids KIF3A, KIF3B and KAP3 form a  heterotrimer (KIF3 complex) in  mice KIF3 complex transports proteins essential for production of  monocilia (Hirokawa  et al.  2006)
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Introduction The node is important for L-R determination Nodal cells have monocilia on their ventral surface Ciliary beating was speculated to be important for  establishing L-R asymmetry To determine the importance of KIF3 in biological  processes, Nonaka  et al.  generated KIF3B knockout  mice. Result was a randomization of Left-Right (L-R) asymmetry
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Methods Used a targeting vector to disrupt KIF3B Electroporated into cells from a line of  embryonic stem cells Disruption of KIF3B was confirmed by  immunoblotting against KIF3B    kif3B-/-  embryos at 9.5 dpc gave no staining Mutant strains maintained by backcrossing Subjected to a variety of analysis Immunoblotting, electron/light/video  microscopy, whole-mount  in situ  hybridization
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Results kif3B-/-  embryos exhibited randomized left-right  asymmetry 16 of 46  kif3B-/-   embryos showed reversed heart looping at  the 9.5 dpc stage kif3B-/-  embryos turned their tails to either left or right  while wild-type turned their tails uniformly to the right wild-type   kif3B-/- 
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Determination of left-right patterning of the mouse -...

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