Real-time observation of Ca2+-induced basal body reorientation in Chlamydomonas.

Abstract

The two basal bodies of Chlamydomonas are connected by a bridge, the distal fiber, that contains a Ca2+-binding protein, centrin. Although various fibrous structures in many organisms containing centrin or similar proteins have been shown to contract at Ca2+ concentrations >10(-7)-10(-6) M, the contractility of the distal fiber in Chlamydomonas has not been demonstrated. To determine whether it undergoes Ca2+-dependent contraction, we isolated the flagella-basal body complex from the paralyzed-flagella mutant pf18 and measured the angle between the two axonemes at different Ca2+ concentrations. Use of a double mutant with the mutant fa1, deficient in the mechanism for Ca2+-dependent flagellar amputation, enabled the measurement at Ca2+ concentrations > or = 10(-4) M. The angle, 80-120 degrees at 10(-9) M Ca(2-), was found to decrease by about 20 degrees when the Ca2+ concentration was raised above 10(-6) M. The angle increased again when the Ca2+ concentration was lowered below 10(-7) M. The flagellar apparatuses isolated from the double mutant between pf18 and the mutant vfl2 deficient in the structural gene of centrin had an angle of 90-130 degrees at 10(-9) M Ca2+, but the angle did not change when the Ca2+ concentration was increased. Thus centrin must be involved in the basal body reorientation. In detergent-extracted cell models of the pf18fa1 mutant, the angle between the two axonemes was found to decrease transiently by about 15 degrees upon iontophoretic application of Ca2+. Hence, the Ca2+-induced basal body reorientation can take place even when the basal body is contained in the cell body covered by the cell wall. It may function as part of the mechanism for phobic responses wherein Chlamydomonas cells swim backward transiently upon reception of strong light or mechanical stimuli.