Gene-gene interactions between mutants that accumulate abnormally high amounts of proglutelin in rice seed

Abstract

We had previously identified eight mutants, esp2 and g(G)lups1 to 7, which accumulated abnormally high amounts of proglutelin, the major storage protein in rice seeds. Analysis of their seed proteins by SDS-PAGE, their levels of the luminal chaperone BiP and gene-gene interactions indicated that these mutants fell into four classes. The most epistatic class consisted of esp2, which encodes a defective protein disulfide isomerase (PDI). A second class consisting of Glup1, glup2 and glup7 was hypostatic to esp2, and showed abnormally high levels of BiP, suggesting that maturation and export of proglutelins from the ER are inhibited in this class of mutants. The third class containing glup4, Glup5 and glup6 mutations was hypostatic to esp2, Glup1, glup2 and glup7. Since the glup4 allele encodes the small GTPase Rab5a, which participates in the trafficking of proglutelin from Golgi apparatus to the protein storage vacuole (PSV), this third class of mutants is likely affected in this process. Lastly, glup3, which encodes a vacuolar processing enzyme, which proteolytically processes proglutelin into acidic and basic subunits within the PSV, was hypostatic to the other mutants. Overall, these gene relationships are consistent with the sequential intracellular transport and processing of proglutelin and provide novel insights on the trafficking of proglutelin to the PSV.

GLUTELIN PRECURSOR ACCUMULATION3Encodes a Regulator of Post-Golgi Vesicular Traffic Essential for Vacuolar Protein Sorting in Rice Endosperm

“…Previous studies have identified eight 57H mutants in rice, including esp2 and glup1 to glup7 (Ueda et al, 2010). Compared with these previously reported rice 57H mutants, the gpa3 mutant displayed several distinct features.…”

“…Compared with these previously reported rice 57H mutants, the gpa3 mutant displayed several distinct features. First, gpa3 accumulated normal levels of PDI1-1 and BiP1, whereas those mutants that have defects in ER export of storage proteins usually exhibited elevated levels of these chaperones, such as esp2/pdi1-1 (Takemoto et al, 2002), glup1, glup2, and glup7 (Ueda et al, 2010). Second, despite the fact that these mutants all accumulate increased levels of proglutelins, there are distinct differences in their subcellular characteristics due to lesions in different trafficking processes.…”

“…To test whether gpa3 is defective in storage protein sorting at the ER level or in post-Golgi trafficking, we compared the levels of ER lumen BINDING PROTEIN1 (BiP1) and PDI1-1, two molecular chaperones known to be induced in 57H rice mutants defective in maturation and export of proglutelins from the ER (Ueda et al, 2010). Immunoblot analysis showed that the levels of BiP1 and PDI1-1 were comparable in the gpa3 mutant and the wild type ( Figure 1F).…”

“…These mutants were named endosperm storage protein (esp2) and glutelin precursor (glup1 to glup7; Ueda et al, 2010). Among them, ESP2 encodes PDI1-1 (for PROTEIN DISULFIDE ISOMERASE-LIKE1-1), involved in proglutelin maturation in the ER (Takemoto et al, 2002).…”

The Small GTPase Rab5a Is Essential for Intracellular Transport of Proglutelin from the Golgi Apparatus to the Protein Storage Vacuole and Endosomal Membrane Organization in Developing Rice Endosperm

“…The mutants were named endosperm storage protein mutant2 (esp2; Kumamaru et al, 1987Kumamaru et al, , 1988 and glutelin precursor mutant1 (glup1) to glup7 (Kumamaru et al, 2007;Satoh-Cruz et al, 2010a;Ueda et al, 2010). Gene-gene interaction analyses of the eight mutant lines indicate that esp2 is the most epistatic while glup3 is the most hypostatic among the eight mutant genes (Ueda et al, 2010). This relationship suggests that the factors encoded by ESP2 and GLUP3 function upstream and downstream, respectively, of the other GLUP genes in the pathway of the synthesis, trafficking, and accumulation of proglutelin.…”