Circadian Activation of the Mitogen-Activated Protein Kinase MAK-1 Facilitates Rhythms in Clock-Controlled Genes in Neurospora crassa

Abstract

The circadian clock regulates the expression of many genes involved in a wide range of biological functions through output pathways such as mitogen-activated protein kinase (MAPK) pathways. We demonstrate here that the clock regulates the phosphorylation, and thus activation, of the MAPKs MAK-1 and MAK-2 in the filamentous fungus Neurospora crassa. In this study, we identified genetic targets of the MAK-1 pathway, which is homologous to the cell wall integrity pathway in Saccharomyces cerevisiae and the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway in mammals. When MAK-1 was deleted from Neurospora cells, vegetative growth was reduced and the transcript levels for over 500 genes were affected, with significant enrichment for genes involved in protein synthesis, biogenesis of cellular components, metabolism, energy production, and transcription. Additionally, of the ϳ500 genes affected by the disruption of MAK-1, more than 25% were previously identified as putative clock-controlled genes. We show that MAK-1 is necessary for robust rhythms of two morning-specific genes, i.e., ccg-1 and the mitochondrial phosphate carrier protein gene NCU07465. Additionally, we show clock regulation of a predicted chitin synthase gene, NCU04352, whose rhythmic accumulation is also dependent upon MAK-1. Together, these data establish a role for the MAK-1 pathway as an output pathway of the circadian clock and suggest a link between rhythmic MAK-1 activity and circadian control of cellular growth.

Involvement of MAK-1 and MAK-2 MAP kinases in cell wall integrity in Neurospora crassa

“…30) The MAK-1 MAPK pathway, composed of MIK-1 MAPKKK, MEK-1 MAPKK, and MAK-1 MAPK, is known to participate in secondary metabolism by regulating the expression of a tyrosinase gene, and plays a role as an output pathway of the circadian clock. 31,32) It is genetically assumed that the MAK-1 pathway contributes to CWI. Cell wall stress is sensed at the cell surface by certain stress sensors such as WSC-1.…”

A Novel Cryptochrome-Dependent Oscillator inNeurospora crassa

“…The vvd knockout (KO) strain was obtained from Christian Heintzen . The Dwc-1::hph strain was obtained from Jay Dunlap (Dartmouth Medical School), and Dwc-1::bar was generated in our lab (Bennett et al 2013). The cry KO [Fungal Genetics Stock Center (FGSC) 12981] was generated by the N. crassa KO project (Colot et al 2006) and obtained from the FGSC.…”

The cell survival pathways of the primordial RNA-DNA complex remain conserved in the extant genomes and may function as proto-oncogenes

“…It is the loss-of-function-mutated PP2A phosphatases that allow MAPK overactivity up to its constitutive expression, resulting in incessantly upregulated hyphal cell proliferation and fusions in some strains (Δpp2A; Δcdc-14; Δcsp-6; Δpzl-1, phosphatase; cell division cycle; cell shape phenotype; phosphatase Z-like). This process fails to respond to the inhibitor butyl-hydroxyperoxide t-BuOOH, Sigma [73,75], thus, imitating chemotherapy-resistant neoplastic cells.…”

“…The light receptors Vivid, White Collar, and Frequency (WC activates the release of frq mRNA) activate the circadian rhythm clock proteins ATF and CREB (activating transcription factor, cyclic AMP-responsive element-binding protein) and the MAPK-OS-MAPK rhythmic pathways, thus, synchronizing cell divisions. The mitogen-activated kinases engage the clock-controlled genes (ccg) and some mitochondrial genes of the cells [73]. The protein products of genes frq and catp (clock ATPase) react with histones in the nucleosome.…”

“…It operates a polyADP-ribose polymerase PARP for restoring its damaged DNA. Several MAPK-ERK pathways are operational, including osmolarity-sensitive OS-MAPK [73]. In hyperosmotic stress, its class 2 phosphatases A2 (PP2A) act.…”