Fig. 4
From: Light regulates the degradation of the regulatory protein VE-1 in the fungus Neurospora crassa
Light regulates the accumulation of VE-1 in aerial hyphae by preventing its degradation. a The degradation of VE-1 in aerial hyphae requires FWD-1. Protein samples from cultures of the Δfwd-1 ve-1FLAG strain kept in the dark or light were obtained from mycelia growing vegetatively in liquid media (M), or as supporting vegetative mycelia after 48 h (M48) of transfer from liquid media to the surface of an agar plate with minimal media to induce conidiation. Conidiating aerial hyphae were collected after 48 h (A48) of the induction of conidiation. Proteins were separated by SDS-PAGE and hybridized with antibodies specific for FLAG. One hundred micrograms of proteins was loaded per lane. As loading control, we used a Coomassie staining of each protein sample. b The degradation of VE-1 in conidia requires FWD-1 and an active CSN. Protein samples were isolated from conidia of the ve-1FLAG, Δcsn-5 ve-1FLAG, and Δfwd-1 ve-1FLAG strains isolated from cultures kept in the dark or light. Proteins were separated by SDS-PAGE and hybridized with antibodies specific for FLAG. Fifty micrograms of proteins was loaded per lane. As loading control, we used a Coomassie staining of each protein sample. c Light promotes the accumulation of VE-1 during conidiation. Protein samples from cultures of the ve-1FLAG strain kept in the dark or light were obtained from mycelia growing vegetatively in liquid media (M), or as supporting vegetative mycelia after 15 h (M15) of transfer from liquid media to the surface of an agar plate with minimal media to induce conidiation. Conidiating aerial hyphae were collected after 15 h (A15) of the induction of conidiation. For the dark to light transfer or light to dark transfer, the M15 or A15 samples were incubated in light or dark, respectively, for the indicated times after 15 h of induction of conidiation in dark or light, respectively, and collected for protein purification. Proteins were separated by SDS-PAGE and hybridized with an antibody specific for FLAG. One hundred micrograms of proteins was loaded per lane. As loading control, we used a Coomassie staining of each protein sample. d A model for the regulation of VE-1 degradation during conidiation and light. VE-1 accumulates in the cytoplasm and nuclei of vegetative hyphae following transcription and translation. VE-1 is degraded by the proteasome in a process that requires interaction with the SCFFWD−1 and ubiquitylation (symbolized by stars) and is regulated by the CSN and light. During the development of aerial hyphae VE-1 is highly degraded but light reduces VE-1 degradation and promotes its translocation to the nucleus for gene regulation