In vivo labelling of functional ribosomes reveals spatial regulation during starvation in Podospora anserina

Abstract

Background: To date, in eukaryotes, ribosomal protein expression is known to be regulated at the transcriptional and/or translational levels. But other forms of regulation may be possible.

Results: Here, we report the successful tagging of functional ribosomal particles with a S7-GFP chimaeric protein, making it possible to observe in vivo ribosome dynamics in the filamentous fungus Podospora anserina. Microscopic observations revealed a novel kind of ribosomal protein regulation during the passage between cell growth and stationary phases, with a transient accumulation of ribosomal proteins and/or ribosome subunits in the nucleus, possibly the nucleolus, being observed at the beginning of stationary phase.

Conclusion: Nuclear sequestration can be another level of ribosomal protein regulation in eukaryotic cells. This may contribute to the regulation of cell growth and division.

Figure 1

Functional complementation the su12-1C1 mutation by su12-GFP. Growth after four days on medium containing 750 mg/l paromomycin of two independent su12-1C1 transformants expressing the S7GFP protein, wild type and untransformed su12-1C1 strain.

Figure 2

Microscopic observation of hyphae expressing the S7GFP protein.A-F, are experimental observations with strains carrying the su12-GFP chimaeric gene. A, C and E visualise GFP fluorescence and B, D and F visualise DAPI straining. The hyphae in A and B is from the growing edge; The hyphae in C and D is taken 1 centimetre away from the growing edge corresponding to about 1 day of stationary phase. The hyphae in E and F is taken 5 centimetres away from the growing edge corresponding to about 1 week of stationary phase. The picture in E was taken with a pose time twice as long as the other pictures. G (GFP fluorescence) and H (DAPI staining) are control observations of GFP alone expressed from the GPD promoter; the hyphae is taken 1 centimetre away from the growing edge as in C and D. I (GFP fluorescence), J (DAPI staining) and K (superposition of I and J) show the sub-nuclear localisation of the foci in two adjacent nuclei. Clearly, the GFP foci are located in a depression of the DAPI staining, where the nucleolus is supposed to be located.