Worm Breeder's Gazette 15(5): 36 (February 1, 1999)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
|1||Laboratory of Molecular Biology, Dept of Ecological Engineering,, Toyohashi University of Technology, Toyohashi 441-8580, Japan|
|2||Northwestern University, Dept of BMBCB, Evanston, IL 60208|
|3||University of Illinois, Dept of Pharmacology, Chicago, IL 60612|
In eukaryotes, kinesins are cytoskeletal motor proteins that mediate intracellular transport of a variety of vesicles and cellular cargo on microtubule tracks, by hydrolysing ATP. We have previously characterized cDNA clones corresponding to 20 genes (klp-1 to klp-20) encoding members of the kinesin super family (Khan et al.,JMB,270,627-639,1997). Using maximum parsimony rules, and ProtML program implemented on the MOLPHY package, we have constructed a phylogenetic tree derived from the motor domain regions of different kinesins. Based on the results of this computation, we have been able to assign nemtaode kinesins in eight distinct classes, and one outgroup where atypical kinesins which do not match with any of the above eight classes in structure have been placed. It is remarkable that the nematode genome encodes representatives of all the known kinesin groups found in mammals (e.g. see Hirokawa, 1996, 1998) To determine the temporal and spatial expression pattern of different kinesin like proteins, we have systematically investigated the transcription of these genes by using in situ hybridization on whole mount embryos, using digioxiginine labeled cDNA probes from representative kinesins from all nine groups. Our results suggest that multiple kinesins co-express in early embryogenesis in C.elegans, and some of these show cell lineage specificity. These results provide the first complete view of the temporal and spatial expression pattern of the kinesin superfamily in a metazoan during early development. We have also used the dsRNAi assay, (Tabara,1998, Fire,1998) for different members of the kinesin gene family to see the resulting phenotypes in the treated animals. Our results are consistent with the predicted roles of different kinesins involved in early embryonic development, such as chromosome movement, and spindle pole orientation (Khan et al., 1997). For example, klp-3 encoded C-terminus retrograde kinesin like protein, and the Chromoskinesin ortholog (CeKLP-12) also helps in chrmosomal locomotion. We have also performed the in situ hybridization of kinesin genes during larval and adult development. Our data suggest differential expression for various members of the kinesin gene family, in a variety of tissues such as the nervous system, musculature, gonads, and hypodermis. For a higher resolution of kinesin gene expression, we are in the process of constructing gfp fusion reporter genes (A. Fire et al.), and analyzing the ectopic expression of the reporter genes in different kinesin mutant backgrounds.Finally we have been able to assign some of the previously uncharcaterized cDNA clones to newly released genomic sequences,that brings the total number of kinesin like proteins to about 25. These new kinesin motors soon will be part of the big motor city once their wheels are checked, and properly aligned. We thank Yuji Kohara, and the sequencing consortium for their continued generosity by providing us with necessary parts for the motor family.