Frequency (gene)
Forward genetics has generated many alleles of frq resulting in strains whose circadian clocks vary in period length.
Feldman had been a graduate student with Colin Pittendrigh at Princeton and went to Caltech in 1967 to begin genetic screens for circadian clock mutants.
Colin Pittendrigh and his colleagues had confirmed in 1959 that the daily cycle of asexual development, described in Neurospora crassa earlier by Brandt,[3] was in fact due to regulation by a circadian clock.
[4] In work published not long before Feldman arrived at Caltech, Malcolm L. Sargent, Winslow R. Briggs and Dow O. Woodward at Stanford University reported that overt expression of the developmental rhythm in conidiation was enhanced in a strain of Neurospora called Timex.
[7][2] Feldman used nitrosoguanidine as a mutagen and used race tubes to screen individual strains surviving the mutagenesis for their circadian period length.
[9] In 1986, frq was cloned by Jay Dunlap and his colleagues using a strategy that involved a long chromosome walk and successful application of the then-untried strategy of rescuing an arrhythmic behavioral mutant through transformation of exogenous DNA arising from the chromosome walk.
These frq transcripts both have capacity to encode two FRQ proteins, a long form of 989 amino acids (lFRQ) and a short form of 890 amino acids (sFRQ); both lFRQ and sFRQ are required for strong rhythmicity although the clock is able to persist at certain temperatures, albeit with a weaker rhythmicity, with just one of the proteins present.
[13] The choice of which protein is made is the result of temperature-dependent splicing of the primary transcript such that it includes or excludes the ATG start codon for lFRQ.
[7] Because sFRQ favors a longer period than lFRQ, free running rhythms in wild type Neurospora are somewhat decreased with increased temperature.
The myriad time-of-day specific phosphorylation that characterize FRQ are predicted to provide structure to this otherwise disordered protein.
Neurospora crassa has a relatively strong codon usage bias compared to S. cerevisiae, a commonly used organism for codon-optimization analysis.
One is the FFC, the negative element complex composed of two copies of FRQ, FRH, and Casein kinase 1 as well as, probably, other less strongly bound proteins.
[16] The other complex which acts as the positive element in the feedback loop includes WC-1 and WC-2; they are GATA transcription factors that, together, form the heterodimeric WCC via their PAS domains.
Heavily phosphorylated FRQ undergoes a conformational change that is detected by the FWD-1 protein, which is part of the SCF type E3 ligase.
[33] Recently the transcription factor ADV-1 was identified as a necessary transducer of clock outputs, including circadian rhythmicity in genes critical to somatic cell fusion.
[38] In the frq[9] mutant Neurospora crassa, a non-temperature compensated rhythm of conidiospore development was still observed in constant darkness (DD).
One way to rationalize this is to assume that many are "slaves" to the frequency/white collar oscillator; they do not possess all of the characteristics of a circadian clock on their own because this is supplied by the FWO.
[2][44] Nonetheless bona fide FRQ-based circadian cocks have been found in organisms other than Neurospora both within the Sordariacea, for instance, in the salient fungal pathogen Botrytis,[45] and also as far afield as Pyronema[46] within the Pezizomycetes, an early-diverging lineage of filamentous ascomycetes.
The finding of frq and conserved circadian clock mechanism inside non-Dikarya, Arbuscular Mycorrhizal Fungi expanded the evolutionary history of this gene in Fungal kingdom.
[30] In general, the TTFLs found in fungi and animals share a similar regulatory architecture, with a single step negative feedback loop, PAS-PAS heterodimeric activators that are conserved, and negative element proteins that largely lack structure and are much less well conserved.
