Expression Patterns ofArabidopsis DRGGenes: Promoter‐GUS Fusions, Quantitative Real‐Time PCR, and Patterns of Protein Accumulation in Response to Environmental Stresses

Abstract

DRGs are very highly conserved GTP-binding proteins. All eukaryotes contain DRG1 and DRG2 orthologs. Arabidopsis has three DRGs: AtDRG1 (At4g39520), AtDRG2 (At1g17470), and AtDRG3 (At1g72660). DRG2 and DRG3 encode proteins that are 95% identical; identity between DRG1 and DRG2/3 is 55%. The focus of this article is expression of Arabidopsis DRGs. DRG1 and DRG2 promoter-GUS constructs showed similar spatial expression in seedlings and mature organs, but gene-specific differences were noted. Quantitative real-time PCR experiments indicated similar levels of DRG1 and DRG2 mRNA accumulation in most tissues. DRG3 transcripts were very low in all tissues. Heat stress at 37°C led to a 10-fold increase in DRG1 transcripts and a 1000-fold increase in DRG3 transcripts. DRG1 antibodies recognized a 43-kD protein, and DRG2 antibodies recognized bands at 30, 43, and 45 kD. Plants were exposed to stresses (salt, heat, cold, UV light, osmotic, and other stresses) and examined by Western blotting. Only heat stress caused detectable changes. Heat did not affect DRG1, but DRG2 and a 72-kD protein recognized by DRG2 antibodies both increased. The modest changes in DRG mRNA and protein levels seen here suggest that other types of regulation, such as altered subcellular localization, may be important for their cellular functions.

Association of DRG1 and DRG2 with Ribosomes from Pea,Arabidopsis, and Yeast

“…The AtDRG1 promoter was more active in leaf veins, and the AtDRG2 promoter was more active in petals, siliques, and leaf trichomes. As assayed by quantitative RT-PCR, AtDRG1 and AtDRG2 mRNA levels were relatively high and similar to each other in every tissue examined (Stafstrom 2008). In contrast, AtDRG3 expression was extremely low in the same tissues.…”

“…OBG proteins, the bacterial cousins of DRGs, are associated with ribosomes and implicated in mediating stress responses (Jiang et al 2007). In Arabidopsis, heat stress stimulates AtDRG3 mRNA accumulation and also alters accumulation patterns of proteins recognized by DRG antibodies (Stafstrom 2008). We are continuing to study relationships between plant stress, ribosome activity, and DRG localization.…”

“…AtDRG1 is a DRG1 ortholog, and AtDRG2 and AtDRG3 are DRG2 orthologs. AtDRG1 and AtDRG2 promoter-GUS fusions revealed similar but subtly different spatial expression patterns in seedlings and mature organs (Stafstrom 2008). For example, both promoters showed strong GUS expression in root apices, developing root primordia, the root stele, cotyledons, pollen, and stigmas.…”

“…Microarray experiments also demonstrated that AtDRG1 and AtDRG2 expression is quite uniform under essentially all conditions tested and that AtDRG3 is strongly stimulated by heat stress and stimulated ;10-fold by several other stresses and in pollen and developing seeds (Schmid et al 2005). Whereas DRG1 protein levels were similar in all of these tissues, DRG2 protein levels were quite variable and notable low in older leaves (Stafstrom 2008). The basis for tissue-specific and developmental differences in relative mRNA and protein abundance is unknown.…”

“…The DRG1 and DRG2 antibodies used in this study were described previously (Devitt et al 1999;Stafstrom 2008). DRG1 antibodies recognized a single band with an apparent molecular mass of ;43 kDa protein (the deduced mass is 41.1 kDa); smaller bands (possible degradation products) were seen only occasionally.…”