The Arabidopsis AtSTE24 Is a CAAXProtease with Broad Substrate Specificity

Abstract

Following prenylation, the proteins are subject to two prenyl-dependent modifications at their C-terminal end, which are required for their subcellular targeting. First, the three C-terminal residues of the CAAX box prenylation signaling motif are removed, which is followed by methylation of the free carboxyl group of the prenyl cysteine moiety. An Arabidopsis homologue of the yeast CAAX protease STE24 (AFC1) was cloned and expressed in rce1⌬ ste24⌬ mutant yeast to demonstrate functional complementation. The petunia calmodulin CaM53 is a prenylated protein terminating in a CTIL CAAX box. Coupled methylation proteolysis assays demonstrated the processing of CaM53 by AtSTE24. In addition, AtSTE24 promoted plasma membrane association of the GFP-RacU88402 fusion protein, which terminates with a CLLM CAAX box. Interestingly, a plant homologue of the second and major CAAX protease in yeast and animal cells, RCE1, was not identified despite the availability of vast amounts of sequence data. Taken together, these data suggest that AtSTE24 may process several prenylated proteins in plant cells, unlike its yeast homologue, which processes only a-mating factor, and its mammalian homologue, for which prenyl-CAAX substrates have not been established. Transient expression of GFPAtSTE24 in leaf epidermal cells of Nicotiana benthamiana showed that AtSTE24 is exclusively localized in the endoplasmic reticulum, suggesting that prenylated proteins in plants are first targeted to the endoplasmic reticulum following their prenylation.

AtFACE-2, a functional Prenylated Protein Protease from Arabidopsis thaliana Related to Mammalian Ras-converting Enzymes

“…This finding and the absence of an obvious plant Rce1p ortholog has led to the proposal that plants lack FACE-2/Rce1-like enzymes. According to this hypothesis, AtSTE24 is proposed to have evolved into a broader biological role as the only CAAX protease in these organisms (19). This hypothesis is consistent with the observation that A. thaliana lacks Ras GTPases (20), which have thus far been shown to be FACE-2/Rce1-specific targets in other organisms.…”

“…AtSTE24 isolated from Arabidopsis thaliana has been functionally characterized and shown to have broad substrate specificity (19). This finding and the absence of an obvious plant Rce1p ortholog has led to the proposal that plants lack FACE-2/Rce1-like enzymes.…”

The Endoplasmic Reticulum Is a Reservoir for WAVE/SCAR Regulatory Complex Signaling in the Arabidopsis Leaf

“…This is an important discovery because it suggests that the endogenous signals and protein machineries that mediate the initial steps of NAP1-and W/SRC-positive regulation are located at the ER surface, rather than residing exclusively at the plasma membrane. Given that SPK1 and ROP localize to the ER and accumulate at ERES (Bracha et al, 2002;Zhang et al, 2010), it is possible that SPK1 and ROP-GTP are direct positive regulators of NAP1-containing complexes at the specialized domains of the ER. Key questions now are centered on the precise nature of NAP1-positive regulation, how it relates to W/SRC assembly, and its full activation at specific cellular locations that have importance during ARP2/3-dependent growth.…”

Enlarged meristems and delayed growth in plp mutants result from lack of CaaX prenyltransferases

“…era1-4, originally called wig-1, and clv3-2 have been described (6,25,28 Additional Methods. The following methods were carried our essentially as described: phenotypic analysis and microscopy (8,27), in vitro and in vivo prenylation assays (34)(35)(36)(37), immunoblots (27,34), and abscisic acid (ABA) germination assays (38). Detailed descriptions can be found in Supporting Materials and Methods, which is published as supporting information on the PNAS web site.…”