The Phospho-Code Determining Circadian Feedback Loop Closure and Output in Neurospora

Mol Cell. 2019 May 16;74(4):771-784.e3. doi: 10.1016/j.molcel.2019.03.003. Epub 2019 Apr 3.


In the negative feedback loop driving fungal and animal circadian oscillators, negative elements (FREQUENCY [FRQ], PERIODS [PERs], and CRYPTOCHROMES [CRYs]) are understood to inhibit their own expression, in part by promoting the phosphorylation of their heterodimeric transcriptional activators (e.g., White Collar-1 [WC-1]-WC-2 [White Collar complex; WCC] and BMAL1/Circadian Locomotor Output Cycles Kaput [CLOCK]). However, correlations between heterodimer activity and phosphorylation are weak, contradictions exist, and mechanistic details are almost wholly lacking. We report mapping of 80 phosphosites on WC-1 and 15 on WC-2 and elucidation of the time-of-day-specific code, requiring both a group of phosphoevents on WC-1 and two distinct clusters on WC-2, that governs circadian repression, leading to feedback loop closure. Combinatorial control via phosphorylation also governs rhythmic WCC binding to the promoters of clock-controlled genes mediating the essential first step in circadian output, a group encoding both transcription factors and signaling proteins. These data provide a basic mechanistic understanding for fundamental events underlying circadian negative feedback and output, key aspects of circadian biology.

Keywords: C-box; DNA binding; FRQ; WC-1; WC-2; ccgs; clock-controlled genes; feedback loop; frq transcription; phosphorylation.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • ARNTL Transcription Factors / genetics
  • Circadian Rhythm / genetics*
  • DNA-Binding Proteins / genetics*
  • Feedback, Physiological
  • Fungal Proteins / genetics*
  • Gene Expression Regulation, Fungal
  • Neurospora crassa / genetics*
  • Neurospora crassa / physiology
  • Phosphorylation
  • Promoter Regions, Genetic / genetics
  • Signal Transduction / genetics
  • Transcription Factors / genetics*


  • ARNTL Transcription Factors
  • DNA-Binding Proteins
  • Fungal Proteins
  • Transcription Factors
  • wc-1 protein, Neurospora crassa
  • white collar 2 protein, Neurospora