The binary protein interactome mapping of the Giardia lamblia proteasome lid reveals extra proteasomal functions of GlRpn11

Abstract

Giardia lamblia does not encode Rpn12 and Sem1, two proteins crucial for assembling the proteasome lid. To understand how the interactions between the giardial proteasome lid subunits may have changed to compensate for their absence, we used the yeast two-hybrid assay to generate a binary protein interaction map of the Giardia lid subunits. Most interactions within the Giardia proteasome lid are stronger than those within the Saccharomyces cerevisiae lid. These may compensate for the absence of Rpn12 and Sem1. A notable exception was the weaker interaction between GlRpn11 and GlRpn8, compared to the strong interaction between Rpn11-Rpn8 of yeast. The Rpn11-Rpn8 dimer provides a platform for lid assembly and their interaction involves the insertion of a methionine residue of Rpn11 into a hydrophobic pocket of Rpn8. Molecular modeling indicates that GlRpn8s pocket is wider, reconciling the experimental observation of its weak interaction with GlRpn11. This weaker interaction may have evolved to support extra proteasomal functions of GlRpn11, which localizes to multiple subcellular regions where other proteasome subunits have not been detected. One such location is the mitosome. Functional complementation in yeast shows that GlRpn11 can influence mitochondrial function and distribution. This, together with its mitosomal localization, indicates that GlRpn11 functions at the mitosome. Thus, this parasites proteasome lid has a simpler subunit architecture and structural attributes that may support dual functionalities for GlRpn11. Such parasite-specific proteasome features could provide new avenues for controlling the transmission of Giardia. 1 Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/613619v1_ufigs1.gif" ALT="Figure 0"> View larger version (33K): org.highwire.dtl.DTLVardef@989e69org.highwire.dtl.DTLVardef@1c8134aorg.highwire.dtl.DTLVardef@6a9794org.highwire.dtl.DTLVardef@5ac20_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIGiardia genome does not encode two proteasomal lid subunits: Rpn12 and Sem1 C_LIO_LIUnique interactions within the lid may compensate for the absence of these two C_LIO_LIGlRpn8:GlRpn11 weakly interacts to support GlRpn11s extra-proteasomal distribution C_LIO_LIGlRpn11 localizes at mitosomes, OZ of VD, and to the VFP C_LIO_LIThe 182-218 fragment of GlRpn11 may regulate mitosomal function C_LI