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Background: Type I lissencephaly is characterized by reduced or absent gyri and disrupted neuronal positioning. Patients are cognitively impaired and develop increasingly severe seizures. Many children die at an early age. Lis1 turns out to be absolutely vital for all embryonic development in mice, while heterozygotic mutations specifically disrupt brain development. It is likely that total loss of Lis1 is also inconsistent with human development. Homologs of Lis1 exist in various organisms, including slime molds, yeasts and insects. In all systems, there is a link between Lis1 and cytoplasmic dynein. Cytoplasmic dynein can translocate along microtubules, carrying various sorts of cargo with it, including membranous organelles and components of the cytoskeleton. Motor activity has been linked to axon transport, membrane trafficking, organelle distribution, chromosome segregation, spindle orientation, and cell migration. Ndel1, which interacts with both Lis1 and dynein is a target of cyclin dependent kinases, including Cdk5, which functions primarily in the brain, and Cdc2, which functions primarily during mitosis. It is clear that Lis1 and Ndel1 function with dynein and may regulate dynein activity in cells. However, the mechanisms involved are just beginning to be elucidated.
Ongoing projects: a) Do Lis1 and/or Ndel1 impact dynein's enzymatic activity? We have evidence that this occurs in vitro, but the extent to which Lis1 can stimulate bovine brain dynein is relatively small (Mesngon et at 2006). We are currently examining how Ndel1 impacts dynein in this assay. Also, dynein complexes are heterogeneous, so we are interested in whether Lis1 interacts with and stimulates a specific subset of dynein complexes substantially, rather than stimulating all complexes modestly.
b) Do Lis1 and/or Ndel1 control dynein processivity? Overexpression or loss-of-function studies indicate that Lis1 and Ndel1 influence dynein's distribution in cultured cells (Smith et al, 2000; Niethammer et al 2000, Hebbar et al 2008). However, it is not yet clear whether this involves a direct effect on dynein motility or whether they target non-processive dynein to specific regions such as the leading edge, the growth cone, the kinetochore, or the centrosome. We are currently exploring a role for Lis1 and Ndel1 in organelle transport to try to answer this question. We have several mutant Lis1 constructs that will allow us to explore the importance of Lis1-dynein interactions, the Lis1-Ndel1 interactions, and Lis1-Lis1 homodimerization. We also have several Ndel1 mutants that allow us to explore the role of phosphorylation by cdks.
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c) Do Lis1 and/or Ndel1 influence all dynein-dependent events, or only some? Lis1 influences all the dynein-dependent events we have tested to date. Most recently we showed a role for Lis1 and Ndel1 in regulating nuclear envelope breakdown (NEBD) in neural stem cells (Hebbar et al, JCB, 2008). Dynein has been linked to NEBD by other labs, so this is one more link between these proteins and dynein. Moreover, the study revealed an interesting cooperation between cdk5 and cdk1 in this process. Our current model is that cdk1 phosphorylation of Ndel1 primes it for phosphorylation by cdk5, which reduces its interaction with cytoplasmic dynein and promotes interaction with Lis1.
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d) Is there crosstalk between Lis1 and/or Ndel1 and other cytoskeletal proteins? Both dynein and Lis1 interact with the c-terminus of the adenomatous polyposis coli (APC) protein (Hebbar et al, Dev. Neurosci, 2008). APC is mutated in the majority of inherited and spontaneous cases of colorectal cancer. The current dogma is that APC controls expression of a transcriptional regulator in response to developmental cues. However, APC also has direct and indirect links to microtubules, and APC mutations disrupt this interaction as well. APC has been shown in other labs to functions in postmitotic neuronal cells. In our study we found that a cancer-causing APC mutation enhanced the brain phenotype of Lis1+/-, mice, suggesting crosstalk between these pathways. Our lab is associated with the Center for Colon Cancer Research at USC/MUSC, which is dedicated to the study of the biology, therapy and prevention of colon cancer. We are therefore currently pursuing the APC-dynein link in intestinal epithelium. |
