A study of light and its possible role in adaptation to temperature changes in Neurospora

Contributed by Robert L. Metzenberg, Stanford University


Neurospora has a strong, light-entrained circadian rhythm that is regulated by several known genes and is driven by a mechanism that is at least partly understood. However, we understand essentially nothing about what good it does for the organism! A wide variety of Just-So stories can be invented to give adaptive significance to this daily cycle, but let me suggest a testable hypothesis. When nature's light goes on at sunrise, the temperature will almost always start to rise as well -- let's say, 10 Celsius degrees in a smallish fraction of a day. Any organism at the mercy of the ambient temperature must maintain optimal activities of hundreds or even thousands of enzymes if it is also to maintain homeostatic, optimal, concentrations of its pathway metabolites. Such an adjustment of enzyme levels is sure to take many minutes, perhaps hours. An organism that start adapting to this temperature change in response to light, rather than reacting tardily to a rise in temperature, could be well-served by such a light-entrained circadian rhythm.

Responding appropriately is more complicated than it might sound. Virtually all enzymes conduct catalysis more rapidly at higher than at lower temperatures. However, some will increase in the forward rate constant of a fixed number of enzyme molecules very strongly over 10 degrees, say, fourfold; others will do so perhaps twofold, and a few, barely at all. Thus, to maintain the same ratio of activity of enzymes in a pathway and hence the same metabolite concentrations, the relative concentration of the highly temperature-responsive enzyme must be allowed to fall, that of the almost unresponsive enzyme must be made to rise, and the enzyme of intermediate responsiveness must be have its concentration adjusted to an slighter degree, if at all.

Let me put this in a more conventional language, that of the Arrhenius equation for enthalpy of activation, delta Ha. A large value for delta Ha will correspond to a highly temperature-responsive enzyme, that is, that is, as the temperature T, expressed in the Kelvin scale, rises, the enzyme rate constant, k, will increase greatly. The integrated form of the Arrhenius equation is:

ln k = -delta Ha/R(1/T) + C

where R is the universal gas constant and C is the integration constant and can be taken to represent the amount or concentration of enzyme. Obviously this is the equation of a straight line, with ln k along the ordinate, 1/T along the abscissa, and with the slope, -delta Ha/R, being a fixed property of the enzyme. Increasing or decreasing concentrations of enzyme will give a family of lines parallel to the original. Different enzymes with different values of -delta Ha will give intersecting lines of different slopes. The very simple working hypothesis is that, faced with a change in actual or projected temperature, the needs of the organism will be served by adjusting the concentration of each enzyme so that their relative ln k values read off the ordinate will remain the same.

The proposed test of the hypothesis, then, is simple in principle. Neurospora crassa would be grown in constant darkness at two temperatures, say 25C (298K) and 35C, and RNA would be extracted for transcriptional profiling. An additional sample would be grown at 25C in the dark, then illuminated, still at 25C, to simulate "sunrise", and RNA would be extracted.

Here are two predictions of the working hypothesis:

(1) For many transcripts, the 25C "sunrise" profile would resemble the 35C continuous darkness profile, not the 25C continuous darkness profile.

(2) Those transcripts whose concentration increased relatively strongly in the 25C "sunrise" and 35C darkness profiles would correspond to a family of enzymes of low delta Ha, i.e., enzymes whose catalytic activity per molecule increased relatively little. Conversely, of course, transcripts whose concentration decreased substantially should be those corresponding to enzymes of high delta Ha. The degree of increase or decrease of concentration might even be related monotonically to the value of the Arrhenius enthalpy, delta Ha. Finally, it should be noted that values for delta Ha are not likely to be in the literature for most enzymes in Neurospora. However, they could be determined very quickly for any easily assayed enzyme by measurement of the relative velocity, in arbitrary units, at two or more temperatures.

Verification of either or both of these predictions would add substantially to our understanding of a complex property of biological systems -- their response to changes in illumination and temperature.