PEENEY-WALLEY, THE MONEY-MAKER

by Klaus Werner Wolf,
Electron Microscopy Unit

A gene of the endemic Jamaican click beetle, Pyrophorus plagiophthalamus (Elateridae), is patented abroad and use for bioluminescence imaging in oncology, genetics and environmental sciences, among other things. This article describes the technology, which arose from basic research, and draws conclusions.

The term “bioluminescence” refers to a biochemical reaction in which an organic compound, luciferin, is oxidized in the presence of oxygen through the catalytic activity of the enzyme luciferase. In the process, short-lived intermediate compounds form, and their decay is accompanied by the emission of visible light.

Although details of the reaction, structure of the luciferin, and in particular the necessary cofactors vary, bioluminescence has evolved several times in the living world. Besides being present in numerous deep-sea organisms, it is found in bacteria, dinoflagellates (single-celled animals), fungi, coelenterates (shellfish), earthworms, millipedes, and insects. In the class of insects, representatives of the springtails (Collembola), flies (Diptera), and beetles (Coleoptera) show bioluminescence (Viviani, 2002).

This brief article concentrates on a Jamaican beetle, Pyrophorus plagiophthalamus, which shows intensive bioluminescence, and this property has been commercially exploited abroad. The beetle has paired, dorsal light-emitting organs on the thorax (Figure 1), in addition to a single, ventral luminescent organ. Males fly through forested areas at night in search of females. The males show continuous luminescence from their ventral organs to signal their presence to the females. The females are stationary in trees and bushes, and respond sporadically with their dorsal organs (Stolz et al., 2003).

P. plagiophthalamus is a click beetle (Order: Coleoptera; Family: Elaterida). This luminescent species is locally known as “peeney-walley.” Approximately 9,000 species belong to the Elateridae, a cosmopolitan family of insects. They can flick themselves into the air by an anatomically complex articulation between the first and second thoracic segment. The movement helps them to avoid predators, and is accompanied by a clicking sound.

Adult click beetles are found on flowers and under bark. The larvae live in soil, litter or rotting wood, and are phytophagous, saprophagous, or predatory.

Biologists use the term ‘endemic’ for organisms that are restricted to naturally delimited areas, such as a lake, a river, a mountain range or an island. Jamaica has a relatively large number of endemic species. Of the approximately 3,000 species of flowering plants, 28% are endemic. In some subgroups of the flowering plants, such as orchids and bromeliads, one-third of the species occur only in Jamaica and nowhere else in the world. Among ferns, 14% are endemic (82 of 597 species). In animals, the situation is similar. Of the 113 bird species that nest on the island, twenty (18%) are endemic. Among the twenty bat species found in Jamaica, three (15%) are endemic. Both because of the large absolute number of species and the high degree of endemism, researchers call Jamaica a biodiversity hotspot.

How is money made from the type of light emitted by P. plagiophthalamus? Profits are realized by exploiting the genetic material of the beetle, where the ability for bioluminescence is encoded. The genetic material consists of strings of nucleotides (organic compounds consisting of a sugar molecule, a base residue and a phosphate group). The stretches of nucleotides (deoxyribonucleic acid, DNA) are contained in the nucleus of the cell and give rise to proteins, which in turn are stretches of amino acids. These proteins are either enzymes or structural proteins forming a cell. The former are substances that facilitate chemical reactions within the cell. The abovementioned luciferase is an enzyme, and its genetic makeup (gene structure) is known. In other words, the sequence of nucleotides coding for luciferase has been determined.

For many years, the study of bioluminescence was in the realm of basic science. The curiosity of researchers was the driving force behind the numerous published studies, and immediate applications of the accumulated knowledge were not obvious at all.

This has changed with the advent of molecular biology. In this discipline there is a need to know whether specific genes are expressed in a particular cell type or under certain conditions. The coding sequence of each gene is preceded by a number of controlling nucleotides called promoters. Modern biotechnology permits the attachment of alien genes to a promoter of interest, and the insertion of this construct into the cell. After this manipulation of the cell has been completed, it would express the attached alien sequence and thereby indicate the activity of a gene of interest. The alien sequence is referred to as a ‘reporter gene.’

It is logical to select genes for the reporter role that are easily detectable. This means, first of all, that the procedure to visualize the gene activity should be simple and cheap. Secondly, the reaction should be sensitive and the signal should be detectable without ambiguities.

The luciferase gene is in several respects an ideal reporter gene. It is easy and cheap to visualize the expression of the enzyme, because no expensive instrumentation is needed. The reaction is also extremely sensitive. Among the bioluminescent organisms, luminescence of click beetles is the brightest.

P. plagiophthalamus is not the brightest click beetle. The record holder is another member of the genus, P. noctilucus, but P. plagiophthalamus is probably close, and has an additional advantage. The beetle is unique in that there is polymorphism in the colour of light emitted. The ventral organs emit light from yellow-green to orange, and the paired dorsal organs from green to yellow-green, which renders it a very interesting model to evolutionary biologists (Stolz et al., 2003). The genes responsible for the emission of differently coloured light are known, and used as reporters. In contrast to other reporter genes, the polymorphism of luciferase in P. plagiophthalamus opens the possibility of monitoring two different genes of interest using the same detection method. The two genes of interest can be distinguished by the different wavelengths of visible light emitted by the reporter genes.

Reporter genes based on bioluminescence are widely used in medical research and in the life and environmental sciences.

A major application is in oncology (the study of tumours). Typically, such studies involve the injection of tumour cells expressing the luciferase gene into experimental animals. The spread of these cells within the organism (metastasis) is then followed. As a consequence of the high sensitivity of the method, even small numbers of cells can be detected, which is vital for the understanding of metastasis of specific cancerous cell lines. Such studies have increased our knowledge of tumour behaviour and contributed to the development of new therapies. Bioluminescence imaging of tumour cells allows an assessment of the efficiencies of therapies.

Other applications of bioluminescence imaging include basic genetic research, microbiology, food technology, and environmental monitoring.

A North American company has recognized the potential of the P. plagiophthalamus luciferase as a reporter gene, patented the procedure, and offers the technology in kits. Only recently, legislation is being drafted in Jamaica that governs possible royalties resulting from the export of biological material from the island. The beetles were collected and exported years ago, when only a permit was needed.

Are there any lessons to be learned from the above case? I see two major issues. First, legislation is overdue that regulates the export of biological material from Jamaica in terms of possible royalties. Pertinent legislation must, however, be very carefully designed. It could backfire if the exchange of specimens between researchers in Jamaica and those abroad is rendered difficult. Any measure that stifles biological research has to be avoided. It has to be kept in mind that there are just a few dozen biologists actively doing research in Jamaica. Exchange and collaboration with colleagues abroad, which necessarily involves the collection of material on the island, is vital, and should in no way be compromised by legislation.

Secondly, we learn how important basic research is. As is obvious with the phenomenon of bioluminescence, it can hardly be anticipated whether the accumulation of basic knowledge finally leads to applications. Without basic data gathered over long periods, including the study of the luminescence of a click beetle found only in Jamaica, its luciferase gene would never have been used in oncology. Given the high biodiversity and the high degree of endemism in Jamaica (see above), one wonders what else is out there that can be put into commercial use. Broad biological research involving laboratory and field work are the prerequisites for repeating locally what has been achieved abroad exploiting the luciferase of P. plagiophthalamus.

References

Stolz, U., S. Velez, K.V. Wood, M. Wood & J.L. Feder (2003). ‘Darwinian natural selection for orange bioluminescent colour in a Jamaican click beetle,’ Proc. Nat. Acad. Sci., 100: 14955-14959.

Viviani, V.R. (2002). ‘The origin, diversity, and structure function relationship of insect luciferase,’ Cell. Mol. Life Sci., 59: 1833-1850.

Acknowledgment

I am grateful to Dr Eric Garraway for lending a pinned specimen of Pyrophorus plagiophthalamus from the insect collection at the Department of Life Sciences.