Cytochrome P450 CYP6L1 is specifically expressed in the reproductive tissues of adult male German cockroaches, Blattella germanica (L.)
Introduction
Cytochrome P450s (P450s) are an important enzymatic system found in all organisms examined, including bacteria, fungi, plants and animals. Each animal has approximately 80 (Caenorhabditis elegans, (Consortium, 1998)) to 90 (Drosophila melanogaster, (Adams et al., 2000)) P450s, and some can metabolize numerous substrates (Bernhardt, 1995, Guengerich, 1995, Nebert and Gonzalez, 1987). P450s are named CYP followed by a number, letter and number indicating the family, subfamily and individual isoform (Nelson et al., 1996). In this system, individual P450s with >40% amino acid sequence identity are usually grouped into the same family, and those with >55% identity grouped into the same subfamily. However, there are several exceptions. For example, CYP6A1 and CYP6B2 are <40% identical, but are both grouped into family 6 because “sequences surrounding the conserved cysteine residue make it clear that these two genes are evolutionarily related” (Nelson et al., 1993). Given that this nomenclature system is based on the overall sequence similarity, and that a minor change in a single amino acid can alter substrate specificity (Lindberg and Negishi, 1989), substrate preference cannot be assumed for a certain P450 based on its name.
Insect P450s catalyze the metabolism of plant allelochemicals and insecticides, as well as physiologically important endogenous compounds including juvenile hormones (JHs), ecdysteroids and pheromones (Berenbaum, 1999, Feyereisen, 1999, Hodgson, 1985, Scott et al., 1998). Among the insect P450s, CYP6B1 (Ma et al., 1994) and CYP6B4 (Hung et al., 1997) were shown to metabolize plant allelochemicals (furanocoumarins). CYP6A1 (Andersen et al., 1994), CYP6A2 (Dunkov et al., 1997), CYP6D1 (Wheelock and Scott, 1992, Zhang and Scott, 1996) and CYP12A1 (Guzov et al., 1998) have been shown to metabolize insecticides. CYP4C7 was demonstrated to metabolize JHs and JH precursors (Sutherland et al., 1998). CYP6A1 also metabolizes JHs and their analogs (Andersen et al., 1997). The cloning of these P450s offers a great opportunity to better understand the roles of P450s in herbivore–plant interactions, insecticide resistance, insect development and physiology.
P450s have been implicated in insect reproduction (Hodgson, 1985, Yu and Terriere, 1974). However, no individual insect P450 has yet been associated with reproduction. Herein, we report cloning of the full-length cDNA of the first sex-specific insect P450, CYP6L1, which is exclusively expressed in the reproductive system of male adult German cockroaches. This finding has important implications for understanding the roles of P450s in insect reproduction and may offer a potential target for the development of novel insect control agents.
Section snippets
Insects
Two strains of German cockroaches were used: Baygon-R (Siegfried and Scott, 1991) and CSMA (Scott and Matsumura, 1981). Cockroaches were reared as described previously (Siegfried and Scott, 1991).
Dissection
To examine the expression of CYP6L1 in different body regions of male and female adult cockroaches the abdomens were separated from the remainder of the bodies on dry ice using forceps. The expression of CYP6L1 in the abdomens of adult male cockroaches was also examined. For this, cockroaches were
cDNA cloning and characterization
The full-length cDNA sequence (accession #: AF227531) of MCHA is shown in Fig. 1. MCHA was named CYP6L1 by the P450 Nomenclature Committee. CYP6L1 has an open reading frame of 1509 nucleotides with a deduced protein of 503 amino acids and a molecular mass of 57 kDa. A BLAST search (Altschul et al., 1997) with the deduced amino acid sequence shows that CYP6L1 is most similar to the members of P450 family 6 from insects. A comparison of CYP6L1 with some members of family 6 is shown in Table 2.
Discussion
Although expression patterns for most insect P450s are not yet clear, available information shows that insect P450s have great diversity in their expression patterns with regard to life stages and tissues (Scott et al., 1998) (Table 2). CYP6L1 is unique because it is the first sex-specific insect P450 reported. Furthermore, CYP6L1 stands out because, although a number of mammalian P450s are expressed at different levels in males and females (Waxman and Chang, 1995), CYP6L1 is the first P450
Acknowledgements
We thank Drs P. Danielson and J. Fogleman for suggestions with the 3′ and 5′ RACE techniques, Drs C. Gilbert, J. Helmann, J. Ewer and M. Wolfner for valuable discussions. We also thank Dr D. Nelson for naming the P450. This project was supported by a grant from Sigma Xi (to ZW) and Hatch Project 139414.
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