Functional and Developmental Impact of Cytosolic Protein N-Terminal Methionine Excision in Arabidopsis  

Abstract

Protein N-terminal methionine (Met) excision (NME) is carried out by two types of Met aminopeptidases (MAPs), MAP1 and MAP2, in eukaryotes. Three enzymes, MAP1A, MAP2A, and MAP2B, have been identified in the cytoplasm of Arabidopsis (Arabidopsis thaliana). MAP transcript quantification revealed a predominance of MAP2B and developmental and organ-specific regulation of both MAP1A and MAP2s. By combining reverse genetics and reverse chemogenomics in transgenic plant lines, we have devised specific and reversible switches for the investigation of the role of cytoplasmic NME in Arabidopsis and of the respective contributions of the two types of cytoplasmic MAPs throughout development. dsRNA interference and knockout (KO) plant lines targeting either MAP1A alone or both MAP2s simultaneously were constructed and shown to display wild-type phenotypes. In the MAP1A KO context, modulating MAP2 activity by treatment with various concentrations of the specific drug fumagillin impaired plant development, with particularly strong effects on the root system. Reciprocally, complete MAP2 inhibition in various MAP1A knocked-down genetic backgrounds also generated a gradient of developmentally abnormal plants, but the effects on the root system were milder than in the KO context. In the absence of MAP2 activity, the severity of the phenotype in the MAP1A knocked-down lines was correlated to the extent of MAP1A mRNA accumulation. Complete cytoplasmic NME inactivation blocked development after plant germination. Thus, in plants, (1) cytoplasmic NME is essential; (2) MAP1A and MAP2s are functionally interchangeable, which is not the case in fungi and animals, as a complete block of either MAP-type activity does not cause any visible molecular or phenotypic effect; and (3) a minimal level of cytoplasmic MAP is required for normal development.

Comparative Large Scale Characterization of Plant versus Mammal Proteins Reveals Similar and Idiosyncratic N-α-Acetylation Features

“…If it is not the case, this further supports the fact that MetAP2, the enzyme in charge of this cleavage, is more limiting in humans than in plants (71). This could explain why it is possible to inactivate this enzyme in plants and not in animals (72,73). Of note, these polypeptides retaining the first Met, although they do have a Thr or a Val at position 2, were found to be NAAed on the Met residue that does not correspond to a clear Nat target (14).…”

N-Myristoylation Regulates the SnRK1 Pathway inArabidopsis

“…On day 6 after imbibition (DAI 6), NMT2 was expressed significantly less strongly than NMT1 ( Figure 1C). The NMT1 transcript was more abundant than other transcripts of the cytosolic N-terminal maturation pathway, such as those of MAP1A, MAP2A, and MAP2B (Ross et al, 2005), whereas the NMT2 transcript was significantly less abundant. We next investigated the expression of the two NMT genes in various organs.…”

Interplay Between N-Terminal Methionine Excision and FtsH Protease Is Essential for Normal Chloroplast Development and Function in Arabidopsis

“…Furthermore, the cytoplasmic NME is essential for normal growth and development of Arabidopsis. NME orchestrates a crosstalk between two fundamental signaling pathways that are frequently deregulated under pathological conditions: thiol status and proteolysis (Ross et al, 2005;Frottin et al, 2009). In all studied compartments, the developmental defects induced by NME inhibition seem to be caused by increased cellular proteolytic activity, primarily induced by an increase in the number of proteins targeted for rapid degradation (Frottin et al, 2009).…”