Golgi Reassembly and Stacking Protein (GRASP) Participates in Vesicle-Mediated RNA Export in Cryptococcus neoformans

Abstract

Golgi reassembly and stacking protein (GRASP) is required for polysaccharide secretion and virulence in <i>Cryptococcus neoformans</i>. In fungal species, extracellular vesicles (EVs) participate in the export of polysaccharides, proteins and RNA. In the present work, we investigated if EV-mediated RNA export is functionally connected with GRASP in <i>C. neoformans</i> using a <i>grasp</i>Δ mutant. Since GRASP-mediated unconventional secretion involves autophagosome formation in yeast, we included the <i>atg7</i>Δ mutant with defective autophagic mechanisms in our analysis. All fungal strains exported EVs but deletion of <i>GRASP</i> or <i>ATG7</i> profoundly affected vesicular dimensions. The mRNA content of the <i>grasp</i>Δ EVs differed substantially from that of the other two strains. The transcripts associated to the endoplasmic reticulum were highly abundant transcripts in <i>grasp</i>Δ EVs. Among non-coding RNAs (ncRNAs), tRNA fragments were the most abundant in both mutant EVs but <i>grasp</i>Δ EVs alone concentrated 22 exclusive sequences. In general, our results showed that the EV RNA content from <i>atg7</i>Δ and WT were more related than the RNA content of <i>grasp</i>Δ, suggesting that GRASP, but not the autophagy regulator Atg7, is involved in the EV export of RNA. This is a previously unknown function for a key regulator of unconventional secretion in eukaryotic cells.