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要旨:
The aliphatic nitroalkene (E)-1 nitropentadec-1-ene (NPD), reported in early seventies in soldiers of the
termite genus Prorhinotermes, was the first documented nitro compound produced by insects. Yet, its
biosynthetic origin has long remained unknown. Here, we investigated in detail the biosynthesis of NPD
in P. simplex soldiers. First, we track the dynamics in major metabolic pathways during soldier ontogeny,
with emphasis on likely NPD precursors and intermediates. Second, we propose a hypothesis of NPD
formation and verify its individual steps using in vivo incubations of putative precursors and intermediates.
Third, we use a de novo assembled RNA-Seq profiles of workers and soldiers to identify
putative enzymes underlying NPD formation. And fourth, we describe the caste- and age-specific
expression dynamics of candidate initial genes of the proposed biosynthetic pathway.
Our observations provide a strong support to the following biosynthetic scenario of NPD formation,
representing an analogy of the sphingolipid pathway starting with the condensation of tetradecanoic
acid with L-serine and leading to the formation of a C16 sphinganine. The C16 sphinganine is then oxidized
at the terminal carbon to give rise to 2-amino-3-hydroxyhexadecanoic acid, further oxidized to 2-amino-
3-oxohexadecanoic acid. Subsequent decarboxylation yields 1-aminopentadecan-2-one, which then
proceeds through six-electron oxidation of the amino moiety to give rise to 1-nitropentadecan-2-one.
Keto group reduction and hydroxyl moiety elimination lead to NPD. The proposed biosynthetic sequence
has been constructed from age-related quantitative dynamics of individual intermediates and confirmed
by the detection of labeled products downstream of the administered labeled intermediates. Comparative
RNA-Seq analyses followed by qRT-PCR validation identified orthologs of serine palmitoyltransferase
and 3-ketodihydrosphingosine reductase genes as highly expressed in the NPD production
site, i.e. the frontal gland of soldiers. A dramatic onset of expression of the two genes in the first days of
soldier's life coincides with the start of NPD biosynthesis, giving further support to the proposed biosynthetic hypothesis.
