Isolation and characterisation of chemotactic mutants of Chlamydomonas reinhardtii obtained by insertional mutagenesis

The swimming behaviour of the green flagellated protist Chlamydomonas reinhardtii is influenced by several different external stimuli including light and chemical attractants. Common components are involved in both the photo- and chemo-sensory transduction pathways, although the nature and organisation of these pathways are poorly understood. To learn move about the mechanism of chemotaxis in Chlamydomonas, we have generated nonchemotactic strains by insertional mutagenesis. The arginine-requiring strain arg7-8 was transformed with DNA carrying the wild-type ARG7 gene. Of the 8630 arginine-independent transformants obtained, five are defective in their chemotaxis towards various sugars. Two of the mutants (CTX2 and CTX3) are blocked only in their response to xylose. Mutant CTX1 is blocked in its response to xylose, maltose and mannitol, but displays normal taxis to sucrose. Mutants CTX4 and CTX5 lack chemotactic responses to all sugars tested. CTX1, CTX4 and CTX5 represent novel chemotactic phenotypes not previously obtained using ultra-violet or chemical mutagenesis. Genetic analysis confirms that each mutation maps to a single nuclear locus that is unlinked to the mating-type locus. Further analysis of CTX4 indicates that the mutant allele is tagged by the transforming ARG7 DNA. CTX4 appears to be defective in a component specific for chemotactic signal transduction since it exhibits wild-type photobehavioural responses (phototaxis and photoshock) as well as the wild-type responses of EGTA-induced trans-flagellum inactivation and acid-induced deflagellation. Insertional mutagenesis has thus permitted the generation of novel chemotactic mutants that will be of value in the molecular dissection oil: the signalling machinery.