The gene NaPMT3 in Nicotiana attenuata encodes putrescine N-methyltransferase 3 (PMT3), an enzyme integral to the biosynthesis of nicotine, a predominant alkaloid in tobacco species. PMT3 catalyzes the methylation of putrescine to N-methylputrescine, marking the first committed step in nicotine synthesis.

Function and Catalytic Activity

PMT3 facilitates the transfer of a methyl group from S-adenosyl-L-methionine (SAM) to putrescine, producing N-methylputrescine and S-adenosyl-L-homocysteine. This reaction is pivotal in directing putrescine towards alkaloid biosynthesis rather than polyamine synthesis. The enzyme's activity is characterized by the following reaction:

[ \text{S-adenosyl-L-methionine} + \text{putrescine} \rightarrow \text{S-adenosyl-L-homocysteine} + \text{N-methylputrescine} ]

This enzymatic function is essential for the production of nicotine, as N-methylputrescine serves as a precursor in the subsequent steps of the nicotine biosynthetic pathway.

Biological Processes and Pathways

PMT3 operates within the nicotine biosynthesis pathway, primarily localized in the roots of Nicotiana attenuata. The pathway involves:

1. Putrescine Formation: Derived from ornithine via ornithine decarboxylase or from arginine via arginine decarboxylase.

2. Methylation by PMT3: Converts putrescine to N-methylputrescine.

3. Oxidation to N-methyl-Δ¹-pyrrolinium Cation: N-methylputrescine is oxidized by N-methylputrescine oxidase (MPO).

4. Condensation with Nicotinic Acid: Forms nicotine through a series of enzymatic reactions.

The expression of PMT genes, including PMT3, is upregulated by jasmonate signaling, particularly in response to herbivore attack or mechanical wounding. This regulation underscores the role of PMT3 in plant defense mechanisms. (pubmed.ncbi.nlm.nih.gov)

Localization

PMT3 is predominantly expressed in the root tissues of Nicotiana attenuata. Within the roots, its expression is localized to the cortex, endodermis, and xylem. This spatial distribution aligns with the site of nicotine biosynthesis, as nicotine is synthesized in the roots and subsequently transported to aerial parts of the plant. (pubmed.ncbi.nlm.nih.gov)

Regulation and Expression

The expression of PMT3 is inducible by methyl jasmonate, a plant hormone associated with stress responses. Studies in related species, such as Nicotiana sylvestris, have demonstrated that the 5'-flanking regions of PMT genes contain elements responsive to jasmonate, facilitating the upregulation of these genes upon exposure to the hormone. (pubmed.ncbi.nlm.nih.gov)

Structural and Evolutionary Considerations

PMT3 belongs to the class I-like S-adenosylmethionine-dependent methyltransferase family. It contains key domains characteristic of this family, including the PABS domain (IPR030374), which is involved in binding S-adenosylmethionine, and the Putrescine N-methyltransferase domain (IPR025803), essential for its catalytic activity. Evolutionarily, PMT enzymes are believed to have diverged from spermidine synthases, adapting to fulfill specialized roles in alkaloid biosynthesis. (en.wikipedia.org)

Conclusion

NaPMT3 in Nicotiana attenuata encodes an enzyme crucial for the initial step in nicotine biosynthesis, catalyzing the methylation of putrescine to N-methylputrescine. Its expression is root-specific and regulated by jasmonate signaling, highlighting its role in the plant's defense response. The enzyme's activity and regulation are integral to understanding nicotine production in tobacco species.