The GUL-1 protein binds multiple RNAs and regulates cell wall remodeling in concert with the MAK-1 pathway in Neurospora crassa
The Neurospora crassa GUL-1 is part of the COT-1 pathway, which plays key roles in regulating polar hyphal growth and cell wall remodeling. We show that GUL-1 is a bona fide mRNA binding protein (RBP) that can associate with 828 “core” mRNA species. When cell wall integrity (CWI) is challenged, the repertoire of associated RNA species increases up to 2628 mRNAs (including its own transcript). GUL-1 can bind mRNAs of genes related to translation, cell wall remodeling, circadian clock, endoplasmatic reticulum (ER), as well as CWI and osmoregulatory response MAPK pathway components. GUL-1 interacts with over 100 different proteins, including stress granule proteins, components of the translational and cell wall remodeling machinery, as well as ER components and those of the MAPK, COT-1 and STRIPAK complexes. Several additional RBPs were also shown to physically interact with GUL-1. We demonstrate that under stress conditions GUL-1 can localize to the ER and that GUL-1 affects the CWI pathway as is evident via altered phosphorylation levels of MAK-1, interaction with mak-1 transcript and involvement in the expression level of the transcription factor adv-1. Taken together, we suggest that the RBP GUL-1 functions in multiple cellular processes, including the regulation of cell wall remodeling. The latter is mechanistically concerted with the MAK-1 pathway in a stress-related manner. Methods: In this study we show that GUL-1 is a bona fide mRNA binding protein (RBP) that associates with different mRNAs, including its own transcript, as determined by RNA antisense purification and RNA immunoprecipitation experiments. Results: We demonstrate that GUL-1 may physically interact with multiple proteins included translation processes, cell wall remodeling and cell wall integrity (CWI). We show that GUL-1 affects the CWI pathway as is evident via altered phosphorylation levels of MAK-1, interaction with mak-1 transcript and involvement in the expression level of the transcription factor adv-1. Conclusions:We suggest that the RBP GUL-1 functions in the regulation of cell wall remodeling in concert with the MAK-1 pathway in a stress-related manner. Overall design: RNA immunoprecipitation analysis was performed on the basis of the protocol described by Niranjanakumari et al., 2002, with minor modifications.