Analysis and characterization of novel Chlamydomonas reinhardtii eyespot mutants
Joseph S. Boyd and Carol L. Dieckmann
Dept. of Molecular and Cellular Biology, University of Arizona, 1007 E. Lowell St., Tucson, AZ 85721 USA
The eyespot apparatus of Chlamydomonas reinhardtii is a complex multi-layered organelle that allows the cell to perceive light. The asymmetric positioning of the eyespot, in association with the daughter four-membered microtubule rootlet, is crucial to its function in generating proper phototactic responses to light. The biogenesis of this organelle, which occurs de novo following each cell division, requires the coordinated assembly of a patch of photoreceptor molecules on the plasma membrane and layers of carotenoid-filled pigment granules in close-packed arrays tightly apposed to thylakoid and chloroplast membranes. Four previously identified loci, eye2, eye3, min1, and mlt1, have been shown to affect the assembly of eyespot pigment granules and/or localization of the photoreceptor patch. Here we describe phenotypic characterization and genetic analysis of three novel non-phototactic eyespot mutants: min3, min4, and mlt2. min3 cells fail to assemble a complete eyespot, possessing a single miniature pigment granule spot apposed to a miniature photoreceptor patch, while min4 cells assemble one or occasionally two miniature eyespots anterior to the wild-type equatorial position. mlt2 cells are characterized by misregulation of both eyespot number and positioning, possessing two to four eyespots per cell. The multiple pigment granule spots of mlt2 coincide with plasma-membrane-localized patches of photoreceptor. These results indicate that while the min3, min4, and mlt2 mutations do not affect the co-association of pigment granule spots with photoreceptor patches, the chloroplast component of the eyespot apparatus is important for maintaining the shape and integrity of the plasma membrane photoreceptor patch.
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