Molecular characterisation of SALMFamide neuropeptides in sea urchins
1 School of Biological and Chemical Sciences, Queen Mary, University of
London, Mile End Road, London, E1 4NS, UK
2 Kristineberg Marine Research Station, Fiskebackskil, S 450 34,
Sweden
* Author for correspondence (e-mail: M.R.Elphick{at}qmul.ac.uk)
Accepted 3 October 2005
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Summary |
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An alternative strategy for identification of echinoid SALMFamides was provided by sequencing the genome of the sea urchin Strongylocentrotus purpuratus. Analysis of whole-genome shotgun sequence data using the Basic Local Alignment Search Tool (BLAST) identified a contig (347664) that contains a coding region for seven putative SALMFamide neuropeptides (PPVTTRSKFTFamide, DAYSAFSFamide, GMSAFSFamide, AQPSFAFamide, GLMPSFAFamide, PHGGSAFVFamide and GDLAFAFamide), which we have named SpurS1-SpurS7, respectively. Three of these peptides (SpurS1-3) have the C-terminal sequences TFamide or SFamide, which are identical or similar to the C-terminal region of the starfish SALMFamide S2. This may explain the occurrence of several S2-like immunoreactive peptides in extracts of Echinus esculentus.
Detailed analysis of the sequence of contig 347664 indicated that the SALMFamide gene in Strongylocentrotus purpuratus comprises two exons, with the first exon encoding a signal peptide sequence and the second exon encoding SpurS1-SpurS7. Characterisation of this gene is important because it is the first echinoderm neuropeptide precursor sequence to be identified and, more specifically, it provides our first insight into the structure and organisation of a SALMFamide gene in an echinoderm. In particular, it has revealed a hitherto unknown complexity in the diversity of SALMFamide neuropeptides that may occur in an echinoderm species because all previous studies, which relied on peptide purification and sequencing, revealed only two SALMFamide neuropeptides in each species examined. It now remains to be established whether or not the occurrence of more than two SALMFamides in Strongylocentrotus purpuratus is a feature that is peculiar to this species and to echinoids in general or is more widespread across the phylum Echinodermata. Identification of SpurS1-SpurS7 provides the basis for comparative analysis of the physiological actions of these peptides in sea urchins and for exploitation of the sea urchin genome sequence to identify the receptor(s) that mediate effects of SALMFamides in echinoderms.
Key words: echinoderm, Echinoidea, Strongylocentrotus purpuratus, Echinus esculentus, neuropeptide, SALMFamide, genome
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Introduction |
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The starfish SALMFamides S1 and S2 were the first echinoderm neuropeptides
to be sequenced and therefore it was of interest to investigate the occurrence
of SALMFamide neuropeptides in other echinoderms. Using the same strategy that
was used to isolate S1 and S2 from starfish species
(Elphick et al., 1991a),
Díaz-Miranda et al.
(1992
) succeeded in
identifying two SALMFamide neuropeptides in the sea cucumber Holothuria
glaberrima (class Holothuroidea):
Gly-Phe-Ser-Lys-Leu-Tyr-Phe-NH2 (GFSKLYFamide) and
Ser-Gly-Tyr-Ser-Val-Leu-Tyr-Phe-NH2 (SGYSVLYFamide). The anatomical
distribution and pharmacological actions of GFSKLYFamide have been examined in
Holothuria glaberrima
(Díaz-Miranda et al.,
1995
, Díaz-Miranda and
García-Arrarás, 1995
) and these studies indicate
that, as in starfish, SALMFamides act as muscle relaxants in holothurians.
Thus, it appears that SALMFamides have a general physiological role in
echinoderms as inhibitory neuromuscular transmitters
(Elphick and Melarange, 2001
).
Consistent with this notion, Ohtani et al.
(1999
) identified two
SALMFamides in the sea cucumber Stichopus japonicus (GYSPFMFamide and
FKSPFMFamide) using muscle contractility as a bioassay. Twenty peptides that
influence muscle activity in Stichopus were identified
(Iwakoshi et al., 1995
;
Ohtani et al., 1999
) but
GYSPFMFamide and FKSPFMFamide were the only peptides that were found to have a
direct inhibitory (relaxing) effect on muscle.
Discovery of two starfish SALMFamides (S1 and S2) and four holothurian
SALMFamides has provided an opportunity to identify structural features that
may be characteristic of this neuropeptide family. Sequence comparison
indicates that a conserved feature of these peptides is the C-terminal motif
Sx(L/F)xFamide, where x is variable. It remains to
be determined, however, whether or not this motif is a feature of SALMFamide
neuropeptides in other echinoderm classes (e.g. Ophiuroidea and Echinoidea). A
number of studies have investigated the occurrence and distribution of
SALMFamide-like peptides in ophiuroids and echinoids. For example,
SALMFamide-like immunoreactivity is present in the adult nervous system of the
brittle star species Ophiura ophiura and Amphipholis
squamata (Ghyoot et al.,
1994; De Bremaeker et al.,
1997
) and pharmacological tests using the starfish peptide S1
indicate that SALMFamides may regulate luminescence in Amphipholis
squamata (De Bremaeker et al.,
1999
). SALMFamide-like immunoreactivity has also been described in
the larval nervous system of echinoids, including the sand dollar
Dendraster excentricus and the sea urchin Psammechinus
miliaris (Thorndyke et al.,
1992
; Beer et al.,
2001
).
To facilitate comparative analysis of SALMFamide neuropeptide structure and function in echinoderms, it is important that SALMFamides are identified in species belonging to the class Ophiuroidea and/or the class Echinoidea. Therefore, in the present study we used two complementary approaches to characterise SALMFamide neuropeptides in sea urchins. SALMFamide radioimmunoassays in combination with high performance liquid chromatography (HPLC) were used to characterise and purify SALMFamides from whole-body extracts of the sea urchin Echinus esculentus. Then genomic sequence data for the sea urchin Strongylocentrotus purpuratus was analysed and a gene encoding a family of seven putative SALMFamide neuropeptides was identified. The data presented here provide the first description of an echinoderm neuropeptide precursor gene and the first insight into the structure of SALMFamide genes in echinoderms.
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Materials and methods |
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Analysis of Strongylocentrotus purpuratus genomic sequence data
The genome of the sea urchin Strongylocentrotus purpuratus has
been sequenced by the Baylor College of Medicine Human Genome Sequencing
Center (BCMHGSC) using Clone-Array Pooled Shotgun Sequencing (CAPSS;
Cai et al., 2001) in response
to a proposal by Eric Davidson and colleagues at the California Institute of
Technology (Cameron et al.,
2000
). To search the Strongylocentrotus purpuratus genome
for genes encoding putative sea urchin SALMFamide neuropeptides, the Basic
Local Alignment Search Tool (BLAST;
Altschul et al., 1990
) facility
on the BCMHGSC website was used
(http://www.hgsc.bcm.tmc.edu/blast/blast.cgi?organism=Spurpuratus).
As a query for a tBLASTn search of sea urchin genomic sequence data, a
hypothetical SALMFamide neuropeptide precursor sequence was used. This query
sequence comprised repeating copies of a peptide incorporating the amino acid
sequence of the starfish SALMFamide S1 followed by a glycine residue as a
C-terminal substrate for amidation and a lysine-arginine dipeptide sequence as
a dibasic cleavage site for endopeptidases (i.e.
GFNSALMFGKRGFNSALMFGKRGFNSALMFGKRGFNSALMFGKRGFNSALMFGKRGFNSALMFGKR). This
BLAST analysis of the sea urchin genome was performed during January 2005
using the dataset of Strongylocentrotus purpuratus contigs available
as of 23/11/2004.
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Results |
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Two S2-like immunoreactive peaks (1 and 2) were detected in the HPLC fractionated 40% ACN/TFA Sep-Pak eluate (Fig. 1B) and two S2-like immunoreactive peaks (3 and 4) were detected in the HPLC fractionated 60% ACN/TFA Sep-Pak eluate (Fig. 1D). Peaks 1, 2, 3 and 4 were also detected by the S1 antiserum SLII (Fig. 1A,C) but were much less immunoreactive with this antiserum than the S2-antiserum BGI (note the differences in the scales of the y axes in Fig. 1). This observation is consistent with the lower levels of S1-like immunoreactivity detected in the Sep-Pak eluates (Table 1). SLII did, however, detect some additional immunoreactive peaks, which eluted before and after peaks 1-4 but were not detected by BGI (Fig. 1).
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As peak 3 contained more SALMFamide-immunoreactivity than any of the other immunoreactive fractions it was selected for further purification by HPLC. Moreover, if, as suggested above, peaks 2 and 3 contain an identical immunoreactive peptide then it is likely that this is by far the most abundant SALMFamide neuropeptide in Echinus. Peak 3 was purified by repeated HPLC fractionation on the C8 column using ACN/TFA or ACN/30 mmol l-1 sodium phosphate (pH 7.0; ACN/PO4) as elutants. A single peak of immunoreactivity was detected throughout purification, indicating that peak 3 contains only one immunoreactive peptide. Seven HPLC steps were required to purify this immunoreactive peptide and the final HPLC chromatogram is shown in Fig. 2.
Purified peak 3 was subjected to automated Edman degradation sequencing using an Applied Biosystems sequencer. Peak 3 eluted in four fractions in HPLC step 7 (Fig. 2) and was sequenced twice; first the second major immunoreactive fraction alone, and then the first, third and fourth fractions combined. Both samples were subjected to seven cleavage cycles and the amino acid sequence Met-Arg-Tyr-His could be clearly resolved from both sets of sequencing data (not shown). The yield of the predominant (cleaved) amino acid fell from about 7 pmol in the first three cycles to about 2 pmol in the fourth cycle and no further sequence data could be obtained from later cycles.
Peak 4 was also subjected to further purification; however, it was not possible to purify this peak to homogeneity (data not shown). In view of this inherent difficulty in purifying neuropeptides from whole-body extracts of sea urchins, we adopted an alternative in silico approach by analysing DNA sequence data made available by the recent sequencing of the genome of the sea urchin Strongylocentrotus purpuratus.
Identification of a gene encoding SALMFamide neuropeptides in Strongylocentrotus purpuratus
Analysis of Strongylocentrotus purpuratus genomic sequence data
using the tBLASTn method with the query
GFNSALMFGKRGFNSALMFGKRGFNSALMFGKRGFNSALMFGKRGFNSALMFGKRGFNSALMFGKR identified
a 16 425 base contig (347664) containing a sequence of 153 bases encoding a
polypeptide that shared significant similarity with the query sequence (E
value=8e-04; Fig. 3A).
Moreover, the polypeptide sequence contained four putative SALMFamide
neuropeptides: GMSAFSFamide, AQPSFAFamide, GLMPSFAFamide, PHGGSAFVFamide
(Fig. 3A). Two of these
peptides have the motif SxF/LxFamide (where x is
variable), which is a characteristic of the SALMFamide neuropeptides that have
been identified in other echinoderms (see Introduction). These data indicated,
therefore, that contig 347664 contains a gene encoding a sea urchin SALMFamide
neuropeptide precursor.
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To facilitate identification of a 5' exon(s), the sequence of contig
347664 was analysed using GenScan 1.0, an online tool for predicting the
locations and exon-intron structures of genes in genomic DNA sequences
(http://genes.mit.edu/GENSCAN.html;
Burge and Karlin, 1997). This
analysis revealed the presence of a putative exon located between bases 11828
and 11986 of contig 347664 encoding a polypeptide sequence of 53 residues,
which was then analysed for the presence of a signal peptide sequence using
SignalP 3.0. The results provided very strong evidence for the presence of a
signal peptide in the N-terminal region of the 53 residue sequence, with
hidden Markov models giving a signal peptide probability of 1.0
(Fig. 3B). Furthermore, SignalP
3.0 predicts that the signal peptide would be cleaved during precursor
processing between residues 25 and 26 (probability=0.945, using hidden Markov
models; Fig. 3B).
Collectively, these data indicate that SALMFamide neuropeptides in
Strongylocentrotus purpuratus are encoded by a gene comprising two
exons, with exon 1 encoding the N-terminal signal peptide of the putative
precursor protein and exon 2 encoding seven structurally related SALMFamide
neuropeptides. The sequence of this gene is shown in
Fig. 4, with the locations of
the two exons indicated by the underlying predicted 266 amino acid residue
precursor protein sequence. The exon-intron boundaries are based on
predictions from analysis of the sequence of contig 347664 using GenScan 1.0
and that conform to the classical 5'-donor and 3'-acceptor
consensus rule (gt/ag). Analysis of genomic DNA sequence 5' to the
predicted start codon (atg) of exon 1 revealed a putative TATA-box-like
promoter (tttatt). The tttatt sequence and the start codon are separated by 24
bases, which is within the normal range for TATA-box containing promoters
(Arkhipova, 1995). At the
3' end of the gene there is consensus polyadenylation signal sequence
(aataaa) located 47 bases downstream from the stop codon.
The predicted sequence of the 266 amino acid residue SALMFamide precursor protein is shown in Fig. 5A. Seven putative SALMFamide neuropeptides are predicted to be generated from this precursor following cleavage at monobasic (K) or dibasic (KR, RR) cleavage sites by endopeptidases and C-terminal amidation by peptidyl-glycine alpha-amidating monooxygenase. Based on their relative positions in the Strongylocentrotus purpuratus SALMFamide precursor, we have named these putative neuropeptides: SpurS1 (PPVTTRSKFTFamide), SpurS2 (DAYSAFSFamide), SpurS3 (GMSAFSFamide), SpurS4 (AQPSFAFamide), SpurS5 (GLMPSFAFamide), SpurS6 (PHGGSAFVFamide) and SpurS7 (GDLAFAFamide; see Fig. 5A,B).
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Discussion |
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Our attempts to purify and sequence SALMFamide neuropeptides from
whole-body acetone extracts of the sea urchin Echinus esculentus had
limited success, with only a partial N-terminal sequence (MRYH) being obtained
for one of the SALMFamide-like immunoreactive peptides present in this
species. The difficulty we had in obtaining pure samples of sea urchin
SALMFamides in sufficient quantities to determine full-length sequences is
probably a consequence of using whole-body extracts. In our previous studies
on starfish, radial nerve cord extracts were used, providing a highly enriched
source of neuropeptides (Elphick et al.,
1991a,b
).
However, this approach was not feasible for Echinus because, unlike
in starfish where the nerve cords are located accessibly along the external
midline of the ambulacrum in each arm, in sea urchins the nerve cords are
embedded within a calcareous exoskeleton, making dissection a much more
difficult and time-consuming procedure. Therefore, we decided that alternative
strategies were more appropriate for identification of SALMFamide
neuropeptides in sea urchins. By analysing whole-genome shotgun sequence data
for the sea urchin Strongylocentrotus purpuratus we have succeeded in
identifying a gene encoding a SALMFamide neuropeptide precursor. This is the
first neuropeptide precursor gene to be characterised in the phylum
Echinodermata and our data provide the first insight into the structural
organisation of echinoderm SALMFamide genes.
The SALMFamide precursor gene in Strongylocentrotus purpuratus
appears to comprise two exons, with the first exon encoding an N-terminal
signal peptide sequence and the second exon encoding seven putative SALMFamide
neuropeptides, which we have named SpurS1-SpurS7
(Fig. 5). The occurrence of a
signal peptide sequence that is encoded by a different exon to the exon
encoding putative neuropeptides is of particular interest because this is a
feature that has been reported before in some, but not all, neuropeptide
precursor genes. For example, in the mollusc Lymnaea stagnalis the
precursor for the neuropeptide FMRFamide is encoded by an mRNA derived from
two exons, with exon 1 encoding the signal peptide and exon 2 encoding
multiple copies of FMRFamide (Kellett et
al., 1994). In contrast, in Drosophila melanogaster the
precursor for FMRFamide-like peptides is encoded by a single exon
(Schneider and Taghert, 1990
).
These differences in the structural organisation of neuropeptide precursor
genes invite functional explanations. In the case of the Lymnaea
FMRFamide gene, an explanation for the presence of a separate exon that
encodes the signal peptide is provided by the occurrence of additional
neuropeptide-encoding exons (3,4,5) located downstream from the
FMRFamide-encoding exon (2). Consequently, transcripts of this gene can be
alternatively spliced to give rise to two different mRNAs: one encoding the
signal peptide (exon 1) and the FMRFamide products of exon 2 and the other
encoding the signal peptide (exon 1) and the alternative neuropeptides encoded
by exons 3, 4 and 5 (Kellett et al.,
1994
). Although the occurrence of a separate signal
peptide-encoding exon does not in itself necessarily imply the existence of
multiple neuropeptide-encoding exons that are alternatively spliced, it is
possible that in Strongylocentrotus purpuratus there are additional
neuropeptide-encoding exon(s) located downstream of the exon encoding
SpurS1-SpurS7. Therefore, we analysed contig 347664 for the presence of
additional neuropeptide-encoding exons but we did not find evidence of any
such coding regions in the
2.5 kb sequence located downstream of the exon
encoding SpurS1-SpurS7.
In addition to the seven neuropeptides (SpurS1-SpurS7) that are likely to
be generated by proteolytic processing of the SALMFamide precursor protein in
Strongylocentrotus purpuratus, other peptides need to be considered
as potential biologically active molecules that may be co-released with the
SALMFamide peptides (Fig. 5A).
Firstly, located between the N-terminal signal peptide and the dibasic
cleavage site that precedes SpurS1, there is an 82 amino acid residue
sequence. A noteworthy feature of this sequence is that it comprises a high
proportion of acidic residues (18x E or D, i.e. 22%), indicating
that this part of the precursor protein probably functions as an acidic
spacer. Similarly, located between SpurS1 and SpurS2 there is a 54 amino acid
residue sequence comprised of
19% acidic residues and located between
SpurS6 and SpurS7 there is a 19 amino acid residue sequence, which contains 5
acidic residues (i.e.
26%). Although the primary functions of these parts
of the precursor protein may be to simply function as acidic spacers, all
three regions do also contain potential monobasic and/or dibasic cleavage
sites, which if targeted by endopeptidases may generate smaller peptide
fragments that could have biological activity as secreted molecules.
Discovery of the SALMFamide precursor gene in Strongylocentrotus
purpuratus has provided an opportunity to analyse the full complement of
SALMFamide neuropeptides that occurs in an echinoderm species. Previous
studies, which have relied on biochemical purification of SALMFamide
neuropeptides, have revealed only two SALMFamides in each species examined
(Elphick et al.,
1991a,b
;
Díaz-Miranda et al.,
1992
; Ohtani et al.,
1999
). Therefore, the discovery of a gene encoding seven putative
SALMFamide neuropeptides in Strongylocentrotus purpuratus is
indicative of an unprecedented level of SALMFamide diversity in this species.
One noteworthy feature of the SALMFamide precursor in Strongylocentrotus
purpuratus is that SpurS2-SpurS6 are positioned in tandem without spacers
and separated only by dibasic cleavage sites. In contrast SpurS1 and SpurS7,
located N-terminally and C-terminally with respect to SpurS2-SpurS6, are
separated from SpurS2-SpurS6 by spacer sequences. One possible explanation for
this pattern in precursor structure is that SpurS2-SpurS6 arose more recently
than SpurS1 and SpurS7 as a result of a series of intragenic tandem
duplications. If this is correct, then the number of SALMFamides in
Strongylocentrotus purpuratus may not necessarily be representative
of all sea urchins. Nevertheless, our biochemical analysis of Echinus
esculentus extracts indicates the presence of at least three SALMFamides
in this species (Fig. 1). Further studies are now required to address this issue. In particular, it will
be interesting to determine the sequences of SALMFamide precursor genes in
other echinoids and in species belonging to other echinoderm classes.
A striking feature of our analysis of SALMFamide-like peptides in Echinus esculentus was that the immunoreactive peptides detected in this species exhibited much greater reactivity with antisera to the starfish SALMFamide neuropeptide S2 than with antisera to the starfish SALMFamide neuropeptide S1 (Table 1, Fig. 1). Identification of a SALMFamide precursor gene in Strongylocentrotus purpuratus may facilitate determination of a possible structural basis for this feature of sea urchin SALMFamides. In particular, it is noteworthy that SpurS1 shares the C-terminal sequence TFamide with S2, whilst SpurS2 and SpurS3 have the C-terminal sequence SFamide, where the penultimate threonine residue of S2 and SpurS1 is replaced with a structurally similar amino acid, serine. The presence of these three putative peptides in Strongylocentrotus purpuratus, which share more C-terminal structural similarity with S2 (TFamide) than with S1 (MFamide), may explain why SALMFamide-like peptides in Echinus esculentus are more immunoreactive with S2 antisera than with S1 antisera.
Discovery of the SALMFamide precursor gene in Strongylocentrotus purpuratus has also enabled further evaluation of the structural features that are characteristic of members of the SALMFamide neuropeptide family. Four of the putative neuropeptides in Strongylocentrotus purpuratus (SpurS1, SpurS2, SpurS3, SpurS6) have the same consensus sequence as the starfish and sea cucumber SALMFamides (i.e. SxL/FxFamide, where x is variable; Fig. 5B). However, SpurS4, SpurS5 and SpurS7 have a proline or leucine residue substituted for the serine residue in the consensus sequence, although they do share the sequence FxFamide with SpurS1, SpurS2, SpurS3, SpurS6 and the Stichopus SALMFamides GYSPFMFamide and FKSPFMFamide. These data indicate that SxL/FxFamide should continue to be recognised as the characteristic motif for SALMFamide neuropeptides in echinoderms. However, identification of the SALMFamide precursor in Strongylocentrotus purpuratus has revealed that deviations from this consensus sequence can occur in at least some of the SALMFamides that are found in any one species of echinoderm.
Based on the sequence conservation that occurs in the C-terminal region of SALMFamide neuropeptides and cross-reactivity with antibodies to the C-terminal region of S2, it is likely that SALMFamide neuropeptides in Echinus share C-terminal sequence similarity with Strongylocentrotus SALMFamides and with other echinoderm SALMFamides. In contrast, the partial N-terminal sequence MRYH obtained for peak 3 from Echinus does not share any obvious sequence similarity with SpurS1-SpurS7 or with other echinoderm SALMFamides. This is not surprising because this region of SALMFamide neuropeptides in Strongylocentrotus and in other echinoderms is highly variable in sequence (see Fig. 5B).
The diversity of putative SALMFamide neuropeptides encoded by the
SALMFamide precursor gene in Strongylocentrotus purpuratus raises
questions about their functions and relative activities. Previous studies on
starfish and sea cucumbers (Elphick et
al., 1995; Díaz-Miranda
and García-Arrarás, 1995
;
Melarange et al., 1999
;
Ohtani et al., 1999
) suggest
that at least some of the SALMFamides present in sea urchins may act as muscle
relaxants. To address this issue, we have performed preliminary experiments in
which the starfish SALMFamides S1 and S2 were tested on tube feet from
Echinus esculentus. Both peptides caused tube foot relaxation when
tested at a concentration of 10 µmol l-1 (N=3; data not
shown), consistent with the relaxing effects that S1 and S2 have on starfish
tube feet (Melarange and Elphick,
2003
). These data provide further evidence that SALMFamide
neuropeptides act as muscle relaxants throughout the phylum Echinodermata, as
discussed previously by Elphick and Melarange
(2001
).
It will be interesting to determine whether or not all seven of the
putative Strongylocentrotus SALMFamide neuropeptides (SpurS1-SpurS7)
act as muscle relaxants and if they do, to determine the relative potencies of
these peptides. Unfortunately, this issue has not been addressed for the two
pairs of SALMFamides identified in sea cucumbers
(Díaz-Miranda et al.,
1992; Ohtani et al.,
1999
but it has been found that the starfish SALMFamide S2 is
approximately ten times more potent than S1 as a muscle relaxant in
Asterias rubens (Melarange et
al., 1999
; Elphick and
Melarange, 2001
). Ongoing studies are investigating the structural
basis for this difference in potency of S1 and S2 (Otara et al.,
2004
,
2005
) and there now exists an
opportunity to extend these studies to the much larger repertoire of
SALMFamide neuropeptides that appear to be present in Strongylocentrotus
purpuratus.
One important aspect of SALMFamide neuropeptides about which little is
known is their mode of action in target tissues
(Elphick and Melarange, 2001).
A series of tests with the starfish SALMFamides S1 and S2 have investigated
cyclic-AMP (cAMP) and cyclic-GMP (cGMP) as potential mediators of the effects
of these peptides; however, no changes in the levels of cAMP or cGMP in target
tissues were observed when exposed to S1 or S2 at concentrations that exert
physiological effects (Melarange and
Elphick, 2003
). Indeed, nothing is known about the molecular
properties of the receptor(s) that mediate effects of SALMFamides in
echinoderms. Identification of the SALMFamide gene in Strongylocentrotus
purpuratus now provides a basis for addressing this issue. In particular,
by analogy with neuropeptides in other animal phyla, it seems likely that
SALMFamides exert at least some of their physiological effects by activating
one or more G-protein coupled receptors (GPCRs). For example, by exploiting
data provided by genome sequencing, a GPCR that is activated by FMRFamide-like
peptides was recently identified in Drosophila
(Cazzamali and Grimmelikhuijzen,
2002
). Similarly, it may therefore be possible to exploit the
complete genome sequence of Strongylocentrotus purpuratus to identify
a SALMFamide receptor(s) in this species.
Finally, our exploitation of Strongylocentrotus purpuratus genomic
sequence data to identify a SALMFamide gene paves the way for identification
of other neuropeptide precursor genes in this species. In other invertebrate
species where genomes have been sequenced it has been possible to identify
large, and possibly complete, complements of genes encoding neuropeptides.
However, these studies have so far been focused on species belonging to
protostomian phyla (Hewes and Taghert,
2001; Li et al.,
1999
; Nathoo et al.,
2001
; Riehe et al.,
2002
; Vanden Broeck,
2001
). The genome sequence of Strongylocentrotus
purpuratus provides an opportunity for genome-wide analysis of
neuropeptide structure and function in a deuterostomian invertebrate belonging
to the phylum Echinodermata. For example, there exists the intriguing prospect
of identifying genes encoding neuropeptides such as gonad-stimulating
substance, an echinoderm hormone that was first discovered in starfish more
than 45 years ago (Chaet and McConnaughy,
1959
) but whose molecular identity still remains unknown.
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Acknowledgments |
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