The Goodwin model: Simulating the effect of cycloheximide and heat shock on the sporulation rhythm of Neurospora crassa

The Goodwin model is a negative feedback oscillator which describes rather closely the putative molecular mechanism of the circadian clock of Neurospo ra and Drosophila. An essential feature is that one or two clock proteins a re synthesized and degraded in a rhythmic fashion. When protein synthesis i n N. crassa (wild-type frq(+) and long-period mutant frg(7)) was inhibited by continuous incubation with increasing concentrations of cycloheximide (C HX) the period of the circadian sporulation rhythmicity is only slightly in creased. The explanation of this effect may be seen in the inhibition of pr otein synthesis and protein degradation. In the model, increasing inhibitio n of both processes led to very similar results with respect to period leng th. That protein degradation is, in fact, inhibited by CHX is shown by dete rmining protein degradation in N. crassa by means of pulse chase experiment s. Phase response curves (PRCs) of the N. crassa sporulation rhythm toward CHX which were reported in the literature and investigated in this paper re vealed significant differences between frq(+) and the long period mutants f rq(7) and csp-1 frq(7). These PRCs were also convincingly simulated by the model, if a transient inhibition of protein degradation by CHX is assumed a s well as a lower constitutive degradation rate of FRQ-protein in the frq(7 )/csp-1 frq(7) mutants. The lower sensitivities of frq(7) and csp-1 frq(7) towards CHX may thus be explained by a lower degradation rate of clock prot ein FRQ(7). The phase shifting by moderate temperature pulses (from 25 to 3 0 degrees C) can also be simulated by the Goodwin model and shows large pha se advances at about CT 16-20 as observed in experiments. In case of higher temperature pulses (from 35 to 42 or 45 degrees C = heat shock) the phase position and form of the PRC changes as protein synthesis is increasingly i nhibited. It is known from earlier experiments that heat shock not only inh ibits the synthesis of many proteins but also inhibits protein degradation. Taking this into account, the Goodwin model also simulates the PRCs of hig h temperature (heat shock) pulses. (C) 1999 Academic Press.