The role of calcium in the regulation of the cell cycle of Xenopus laevis embryos

Thomas James Keating, Purdue University

Abstract

Levels of intracellular free calcium ( (Ca$\sp{2+}\rbrack\sb{i})$ in albino Xenopus laevis embryos were measured using aequorin and a photon-counting system. The recordings showed that there are sinusoidal oscillations (Ca$\sp{2+}\rbrack\sb{i}$ having the same frequency as cleavage, with mitosis beginning after the peak of each (Ca$\sp{2+}\rbrack\sb{i}$ cycle and cleavage occurring when (Ca$\sp{2+}\rbrack\sb{i}$ is lowest. The (Ca$\sp{2+}\rbrack\sb{i}$ oscillations are superimposed on a secondary pattern that increases, peaks between third and fifth cleavages and then declines to a level similar to that measured before first cleavage. The oscillations are due to release of calcium from intracellular stores, since the signal is the same in calcium-free solution as in normal medium. Similar calcium oscillations are present in embryos in which cleavage has been blocked with microtubule-disrupting drugs as well as in artificially activated eggs. In nondividing eggs and embryos the secondary pattern increases until about the third cycle and remains elevated for the duration of the recordings. Measurements of (Ca$\sp{2+}\rbrack\sb{i}$ from different subregions of cleavage-blocked embryos shows that the calcium changes occur uniformly throughout the embryos, implying that the (Ca$\sp{2+}\rbrack\sb{i}$ changes are uniform within individual blastomeres in cleaving eggs. When aequorin is injected into individual blastomeres of 64-cell embryos, the shapes of the signals are substantially different from those seen in recordings from whole embryos, indicating that (Ca$\sp{2+}\rbrack\sb{i}$ increases are not uniform across the multicellular embryo but are localized. To investigate further the role of calcium in cell cycle control, I attempted to prevent calcium changes in cycling extracts prepared from Xenopus eggs by adding an antibody directed against phosphatidylinositol 4,5-bisphosphate (PIP$\sb2),$ the precursor for the calcium-releasing second messenger, inositol 1,4,5-trisphosphate (IP$\sb3).$ While the antibody slows cell cycle progression in these extracts, this is the result of an inhibition of protein synthesis rather than a specific effect on the cell cycle machinery. Subsequent experiments suggested that the inhibition of translation is not the result of the antibody blocking production of IP$\sb3$ or diacylglycerol, the byproducts of PIP$\sb2$ hydrolysis, although the exact mechanism is presently unclear.

Degree

Ph.D.

Advisors

Robinson, Purdue University.

Subject Area

Anatomy & physiology|Animals

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