Insect Biochemistry and Molecular Biology 30 2000 711–718 www.elsevier.comlocateibmb
Identification of multiple peptides homologous to cockroach and cricket allatostatins in the stick insect Carausius morosus
Matthias W. Lorenz
a,c,
, Roland Kellner
b
, Klaus H. Hoffmann
a
, Gerd Ga¨de
c
a
Department of Animal Ecology 1, University of Bayreuth, Universita¨tsstrasse 30NW I, D-95440 Bayreuth, Germany
b
Biomed FoGBT, Merck KGaA, D-64271 Darmstadt,Germany
c
Zoology Department, University of Cape Town, Rondebosch 7701, South Africa Received 31 October 1999; received in revised form 31 December 1999; accepted 25 January 2000
Abstract
Eighteen peptides were isolated from brain extracts of the stick insect Carausius morosus. The peptides were purified in four steps by high-performance liquid chromatography, monitored by their ability to inhibit juvenile hormone biosynthesis by corpora
allata of the cricket Gryllus bimaculatus in vitro, and chemically characterised by Edman degradation and mass spectrometry. We obtained complete primary-structure information for nine peptides, four of which belong to the peptide family characterised by a
common C-terminal pentapeptide sequence –YXFGLamide. The remaining five belong to the W
2
W
9
amide peptide family, nonapep- tides characterised by having the amino acid tryptophan in positions 2 and 9. The amino-acid sequence of two other peptides could
not be completely resolved by means of Edman degradation; however, these peptides could be allocated to the –YXFGLamide and the W
2
W
9
amide family, respectively, by comparison of retention times, co-elution and mass spectrometry. Both classes of neuropep- tides strongly inhibit juvenile hormone biosynthesis in crickets but show no inhibiting effect on the corpora allata of the stick
insect.
2000 Elsevier Science Ltd. All rights reserved.
Keywords: Allatostatin; Peptide sequence; Juvenile hormone; Corpora allata; Stick insect; Cricket
1. Introduction
Juvenile hormones JHs regulate a variety of pro- cesses in insects, among which metamorphosis and
reproduction are the best understood. The rate of JH biosynthesis in the corpora allata CA is, in turn, con-
trolled by stimulatory allatotropin and inhibitory allatostatin factors. The latter have been the subject of
a large number of investigations in the last decade. The first allatostatins identified were isolated from brains of
the cockroach Diploptera punctata Woodhead et al., 1989; Pratt et al., 1991. Soon thereafter structurally
related neuropeptides with the C-terminal pentapeptide motif –YXFGLamide were identified in other cockroach
species Weaver et al., 1994; Belle´s et al., 1994, in flies Duve et al., 1993, mosquitoes Veenstra et al., 1997,
bees Kaatz, personal communication, crickets Lorenz
Corresponding author. Tel.: +
49-921-55-2655; fax: +
49-921-55- 2784.
E-mail address:
matthias.lorenzuni-bayreuth.de M.W.
Lorenz.
0965-174800 - see front matter
2000 Elsevier Science Ltd. All rights reserved. PII: S 0 9 6 5 - 1 7 4 8 0 0 0 0 0 4 2 - 4
et al., 1995a; Lorenz et al., 1999a, locusts Veelaert et al., 1996 and moths Davis et al., 1997; Duve et al.,
1997a,b, and even in the sister group of the insects, the crustacean Carcinus maenas Duve et al., 1997c.
However, the peptides isolated from flies, mosquitoes, bees, locusts and moths are not biologically active in
inhibiting JH biosynthesis in those species. In contrast, a peptide with a different structure to the –YXFGLamide
peptides was identified in the tobacco hornworm Mand- uca sexta, and it clearly inhibited JH biosynthesis
Kramer et al., 1991. The same peptide was also found in some other moth species Weaver et al., 1998.
Another group of neuropeptides with allatostatic activity in a homologous bioassay, which contains a
common W
2
W
9
amide motif, has been characterised from the brains of the cricket Gryllus bimaculatus Lorenz et
al., 1995b; Lorenz et al., 1999a. Peptides with structural similarities had previously been identified from Locusta
migratoria Schoofs et al., 1991 and M. sexta Blackburn et al., 1995 using a bioassay that monitored
their ability to inhibit contractions of smooth muscles.
We are interested in the development and reproductive
712 M.W. Lorenz et al. Insect Biochemistry and Molecular Biology 30 2000 711–718
physiology of the parthenogenetic stick insect Carausius morosus. Previous studies had revealed that, although JH
is apparently not required for normal vitellogenesis Pflugfelder, 1937; Bradley et al., 1995, the CA from
adult egg-carrying stick insects do synthesise and release JH III in vitro Lorenz et al., 1999b. In pilot studies we
were also able to extract and partially purify fractions from the brain of adult stick insects, which inhibited JH
biosynthesis of CA from G. bimaculatus in vitro Lorenz and Hoffmann, 1994; Ga¨de et al., 1997. The present
investigation was designed to purify and identify these factors, so that synthetic peptides could be obtained for
more intensive studies of their effects on the repro- ductive cycle of the stick insect.
2. Materials and methods
