Germination of myxospores from the fruiting bodies of Myxococcus xanthus

Germination of myxospores from fruiting bodies of Myxococcus xanthus was examined under a light microscope as well as by analyzing the incorporation of [3H]uracil into the RNA fraction. Efficient germination was observed in 0.2% Casitone containing 8 mM MgSO4 and 1 mM CaCl2 at 30 degrees C. Under this condition, spherical myxospores were converted into rod-shaped vegetative cells within 5 to 6 h. The germination was severely inhibited in the presence of 1 mM phenylmethylsulfonyl fluoride, a protease inhibitor, indicating that a serine protease(s) is required for the myxospore germination. EGTA (1 mM) also completely blocked germination, indicating that Ca2+ plays an important role in myxospore germination. In 1% Casitone without added Mg2+ and Ca2+ or 0.2% Casamino Acids with 8 mM MgSO4 and 1 mM CaCl2, myxospores lost their refractility under a phase microscope, while no RNA synthesis took place within 6 h, as judged by the incorporation of [3H]uracil. A group of proteins were found to be specifically synthesized during an early stage of germination. In addition, a new major spore-associated protein with a size of 41.5 kDa became detectable in the spore shell fraction 3 h after germination. The present results demonstrate that myxospore germination occurs in at least two steps: the loss of myxospore refractility, followed by an outburst of metabolic activities. The first step can occur even in the absence of energy metabolism, while the second step was blocked by rifampin, EGTA, and protease inhibitors.