Comparative Transcriptomic Analysis Reveals Bioluminescence-Related Genes in Firefly Pyrocoelia pectoralis

• A third firefly species transcriptome confirming the same luminous-tissue gene signature. RNA-seq comparison of luminous and non-luminous larval tissues of Pyrocoelia pectoralis (the endangered Chinese firefly, Lampyrinae, sister to Photinus pyralis with a divergence time of ~51 Mya per Fu 2024). The differentially-expressed gene profile recapitulates what Fallon 2018 found in P. pyralis and Zhang 2020 found in Lamprigera yunnana and Abscondita terminalis (Luciolinae): a consistent, cross-species, cross-subfamily signature for “what a firefly lantern looks like at the transcript level.” This is the first dedicated luminous-vs-non-luminous tissue transcriptome for P. pectoralis specifically, even though the genome has been available since Fu 2017 and the chromosome-level assembly since Fu 2024.
• Upregulated in lanterns: cystathionine gamma-lyase (CGL), 4-hydroxyphenylpyruvate dioxygenase (HPPD), β-glucosidase 2 (BGL2), and multiple 4-coumarate:CoA ligases (4CLs). CGL produces L-cysteine from cystathionine, confirming active cysteine biosynthesis in lanterns. HPPD converts 4-hydroxyphenylpyruvate to homogentisate, the first committed step of the tyrosine-catabolism → benzoquinone-acetic-acid route Zhang 2020 proposed as an alternative BQ source. BGL2 hydrolyzes arbutin to release hydroquinone, the canonical Oba 2013 / Kanie 2018 step. Multiple 4CL paralogs were upregulated and explicitly proposed as candidates for the formation of the 2-S-cysteinylhydroquinone intermediate. All four findings reproduce Zhang 2020's transcriptomic results in a phylogenetically distant Lampyrinae species.
• Downregulated in lanterns: cysteine dioxygenase type 1 (CDO1). CDO1 is the entry point of cysteine catabolism (cysteine → cysteinesulfinic acid, ultimately to taurine and inorganic sulfate). Its downregulation in lanterns confirms the Zhang 2020 finding that lantern tissue suppresses cysteine catabolism while upregulating cysteine biosynthesis, channeling the L-cysteine pool toward luciferin biosynthesis rather than degradation. This is the cleanest single piece of evidence that lantern tissue is metabolically optimized to retain L-cysteine, and the reciprocal regulation (CGL up + CDO1 down) is now seen in three independent firefly species.
• The cross-species convergence is the headline. Five firefly species across three subfamilies (Lampyrinae: P. pyralis, P. pectoralis; Luciolinae: L. yunnana, A. terminalis, A. lateralis) now have published lantern transcriptomes, and the same core gene set keeps showing up: BGL, HPPD, polyphenol oxidases / laccases, 4CL, ACOT, luciferase, LST, with CGL up and CDO1 down on the cysteine side. This level of cross-species convergence is the strongest available indicator that the candidate gene assignments are reproducible biological signal rather than per-species artifact, and it is the basis on which any heterologous reconstitution effort can confidently pick which beetle homologs to clone.
• Methodological scope: larval rather than adult tissue. Worth flagging that this is a larval transcriptome, while Oba 2013 and Kanie 2018 established that luciferin biosynthesis occurs primarily in the pupal stage in L. lateralis. Wang 2025 confirms that the same gene-expression signature is present in larval lantern tissue (which is bioluminescent, fireflies glow as larvae before they flash as adults), so the biosynthetic machinery is at least transcriptionally engaged in larvae too. This is consistent with Strause et al. 1979's observation that luciferin synthesis rate is more abundant in larval and pupal stages than in adults.
• Limited new mechanistic insight; this is a confirmation paper, not a paradigm-shifting one. No knockout experiments, no in vitro enzyme assays, no novel pathway proposals beyond the existing literature. The contribution is replication of the Zhang 2020 / Fallon 2018 transcriptomic signature in another Lampyrinae species, plus formal documentation of the P. pectoralis lantern transcriptome itself as a resource for the field. Useful to cite for “this finding has been replicated across multiple firefly species” claims, less useful as a primary mechanistic citation.

Bottom line for the project: This paper strengthens the bibliography by giving cross-species replication for the four enzymatic gene assignments in the construct design. Three concrete uses. First, for TU3 (BGLU46+SKL): β-glucosidase 2 upregulation in P. pectoralis lanterns adds a third independent firefly species (after P. pyralis via Fallon 2018 and L. yunnana / A. terminalis via Zhang 2020) confirming that BGL is part of the lantern biosynthetic machinery. Cross-species convergence on the same gene class strengthens the case for using a BGL ortholog (Arabidopsis BGLU46 in this case) as TU3, and weakens any reviewer objection that the gene assignment rests on a single species' transcriptomics. Second, the cysteine-anabolism-up / cysteine-catabolism-down pattern (CGL up, CDO1 down) reproduced here matters for the heterologous host design: it tells you that fireflies don't accomplish cysteine availability through specialized lantern-only biosynthesis machinery, just by turning off catabolism. Tobacco runs active cysteine biosynthesis through its sulfur assimilation pathway and does not have a strong lantern-equivalent CDO1-driven cysteine sink, so by default N. tabacum leaf cells should already be in a “high cysteine availability” state that favors the spontaneous BQ + L-cys → luciferin chemistry, the engineering doesn't need to add cysteine biosynthesis genes. Third, the 4CL upregulation result is intellectually interesting because plants natively have 4CL as a core phenylpropanoid pathway enzyme, so endogenous tobacco 4CL might in principle participate in the heterologous luciferin pathway by activating phenolic intermediates to CoA esters, which could either help or hurt depending on flux competition. Worth keeping on a watch list for Phase 2 troubleshooting if the four-TU construct underperforms expectations: if 4CL-dependent intermediates accumulate in tobacco, native plant 4CL activity might be a relevant background variable. Cite Wang 2025 alongside Zhang 2020 and Fallon 2018 wherever cross-species transcriptomic convergence is the argument being made, three papers, three subfamilies, same gene list, same regulatory direction.