BACKGROUND
the ability to synthesize rigid, mineralized structures is an essential trait to the majority of metazoan taxa.
vertebrates, echinoderms, mollusks, arthropods, brachiopods, bryozoans, annelids, cnidarians and sponges, amongst others, construct a spectacular diversity of endo- and exo-skeletons as well as sensory and protective structures from a range of minerals  <cit> .
the importance of this trait is highlighted by the observation that the so called 'cambrian explosion' was accompanied by the diversification of biomineralization mechanisms  <cit> , despite the fact that several lineages possessed this ability before the end of the proterozoic  <cit> .
it is currently unknown whether the molecular mechanisms used to create these structures have been inherited from an ancestral biomineralization repertoire, invented de novo, or are the result of an unprecedented lateral genetic transfer  <cit> .
the evolutionary origins, mode of construction, patterning and physical properties of the molluskan shell have held the attention of scientists for centuries, however the molecular mechanisms by which these structures are constructed are only now beginning to be elucidated  <cit> .
the mollusk shell is assembled extracellularly and is an ensemble of caco <dig> and organic macromolecules , which are secreted by the mantle epithelium.
the anterior edge of the mantle tissue underlies the lip of the shell and directs the ordered biomineralization of the different structural layers of the shell and controls the patterning of architectural and color features.
while the structure and function of a number of shell matrix proteins have recently been characterized  <cit> , the regulatory mechanisms that govern these shell-building processes remain largely unknown.
it has long been acknowledged that the diversity of shell types found in gastropod, bivalve and scaphopod mollusks are achieved through the ordered secretion of proteins and other molecules along the length of mantle  <cit> , however the full complexity and role of differential gene activity in the mantle remains undescribed.
the color, structure and geometric pattern of a sea shell is a historical record of the incorporation of proteins into the shell matrix and onto its surface, and directly reflects the gene expression activity of the mantle during the life of a mollusk  <cit> .
using the vetigastropod haliotis asinina  as a model, we sought to determine the complexity of the mantle transcriptome.
abalone shells are composed of three structurally-distinct layers:  the inner nacreous  layer, consisting of layers of aragonitic tablets encased within organic sheaths;  the calcitic prismatic layer, also containing organic macromolecules; and  the outer periostracum, a thin organic veneer that protects and decorates the shell  <cit> .
the anterior edge of the abalone mantle epithelium is convoluted and partitioned into discrete zones that produce each of these layers  <cit> .
within each of these zones are a number of cell types, which contribute to the construction and patterning of the shell  <cit> .
here we assess the complexity of gene expression in the h. asinina mantle, and explore the regulatory and structural factors that contribute to the construction of the shell.
previous studies have demonstrated that the organic component of the shell  is essential to its construction, and confers its remarkable physical properties.
for example, lustrin-a  <cit>  is thought to impart fracture resistant, elastomeric properties to the nacreous layer, while macromolecules isolated from calcitic or nacreous environments can direct the type of polymorph of caco <dig> that will be deposited in vitro  <cit> .
unfortunately, shell matrix proteins are often insoluble, highly acidic or complexed with minerals, making their purification very difficult  <cit> .
to gain a broader understanding of the molecular processes that underlie seashell construction, we have analyzed expressed sequence tags  from the mantle of juvenile h. asinina.
this approach allows for the identification of gene products that are not necessarily incorporated into the shell, but are nonetheless crucial for caco <dig> precipitation and other biomineralization events within the pallial space adjacent to the mantle.
other mantle-localized, secreted gene products not involved in biomineralization will also be detected by this methodology.
we have compared this est set with the recently sequenced l. scutum  genome in order to infer the degree of evolutionary conservation between shell building secretomes within one molluskan class.
RESULTS
structure of h. asinina shell and mantle
the juvenile shell of h. asinina is an ideal model system with which to study the molecular events of shell construction and patterning.
a complex but regular chromatic pattern adorns the outermost layer of the shell, the periostracum.
here, a series of dots  are laid down on top of the ridges of the shell.
the color of the dot will be blue if it overlies a brown/red background, or orange if it overlies a cream background .
this model system is such that it allows for the spatial mapping of gene expression profiles within the mantle to patterning and structural events at the leading edge of the shell.
the mineralogical composition of the shell is partitioned dorso-ventrally into two major layers; a dorsal calcitic layer and a ventral aragonitic layer .
the mantle epithelium, which secretes the proteins responsible for the construction of these structures, is convoluted at its anterior edge.
between the two main folds  lies the periostracal groove , into which the periostracum is secreted and then extruded onto the dorsal surface of the shell.
we have identified a second minor fold within the mantle of h. asinina that we have termed the anterior crease of the outer fold .
although we cannot yet assign a specific function to this structure, the fact that it possesses an abundance of microvilli  suggests that it is actively secreting substances responsible for shell or periostracum construction.
mantle expressed sequence tags
we have sequenced  <dig> randomly selected clones from a cdna library constructed from the juvenile mantle tissue of the tropical abalone, haliotis asinina .
these sequences are available under genbank accession numbers dw <dig> to dq <dig>  this tissue was taken from juveniles maintained in warm water  that were rapidly depositing shell material, approximately  <dig> μm per day  <cit> .
from this survey, we have identified  <dig> unique est clusters  that are expressed in the mantle .
based on the presence of conserved signal sequences and similarity to secreted proteins in genbank  we conservatively estimate that 26% of these  encode secreted proteins.
amongst the  <dig> unigenes that lack both significant similarity to sequences lodged in public databases  and a signal sequence, some may be secreted via mechanisms alternative to the classical signal peptide pathway.
of the  <dig> intracellular unigenes that encode proteins with significant similarity to genbank sequences,  <dig> encode trafficking and mineral binding proteins, and are likely to represent mechanisms essential for the supply of shell building components.
for example, based upon in situ hybridization analyses  and previous studies on bivalves  <cit> , ferritin and calmodulin are likely to be playing fundamental and evolutionarily ancient roles in shell construction within the mollusca.
when compared with est surveys in other metazoan tissues , a markedly higher proportion  of the genes expressed in the abalone mantle encode secreted proteins.
by extrapolation, we estimate that hundreds of proteins are released extracellularly from the mantle, and contribute to the fabrication of the shell.
this estimate however needs to take into consideration non-biomineralizing secreted proteins that do not possess similarity with previously described genbank sequences.
this estimate is in marked contrast to the current number of biomineralizing proteins isolated from the shells of other gastropods and bivalves.
with efforts chiefly focused on the characterization of proteins from the nacreous layer  <cit> , fewer than  <dig> protein families have been shown to contribute to shell formation to date  <cit> .
strikingly, of the  <dig> unigenes that encode putative secreted proteins in the haliotis asinina mantle,  <dig>  do not share significant similarity to any sequences in genbank .
however, several novel secreted open reading frames  possess motifs comparable to proteins in other organisms, such as ggyglgl repeats – similar to the elasticity regions of spidroin, an elastomeric spider silk protein  <cit>  – and proline-rich repeats  as found in plant cell walls  <cit> .
in comparison,  <dig> of the  <dig> intracellular genes  do not match sequences in genbank.
interestingly, blast alignments between h. asinina mantle ests and genomic trace sequences from the gastropod l. scutum, which is estimated to be sequenced to 8× coverage and is currently being assembled  <cit> , reveals that only  <dig> of the  <dig>  novel secreted proteins in haliotis appear to have identifiable homologues in the lottia genome .
these  <dig> genes may be involved in conserved aspects of shell construction in gastropods and possibly other mollusks.
in contrast, the remaining  <dig> novel secreted proteins are likely to either represent genes that evolved after these gastropod lineages split, have been lost in the lottia lineage or a combination of these two scenarios.
likewise, many previously discovered molluskan shell matrix genes do not have clear homologues in the lottia genome .
the few shell matrix proteins, such as lustrin  <cit> , perlucin  <cit>  and mucoperlin  <cit> , are likely to be conserved components of molluskan shells, although their general function in shell fabrication and evolutionary origins are currently unknown.
together, these data suggest that the complex secretome involved in mollusk shell construction is encoded primarily by rapidly evolving genes.
localized expression of mantle genes
in order to infer the function of a subset of these ests, we selected  <dig> for spatial expression analysis on the basis of whether they were novel, predicted to be secreted, possessed repetitive domains and/or shared significant similarity with a gene likely to play a role in biomineralization .
of these,  <dig> are novel and one appears to have a homologue only in the lottia genome .
the spatial expression patterns of these  <dig> mantle ests reveals a diversity of territories and cell types in the mantle , and highlights the modular nature of the mantle tissue  <cit> .
some of these  are expressed homogenously in one or more zones responsible for the creation of different shell layers.
for example, lustrina, first isolated from haliotis rufescens, the gene product of which contributes to the elastomeric properties of the nacreous layer  <cit> , is expressed, along with two other novel genes, in the mantle territory responsible for the production of the inner nacreous layer .
other evolutionarily conserved genes, including calmodulin and a calcium-binding protein, are expressed continuously along the length of the mantle within both the inner fold and the anterior crease of the outer fold , while others, such as ferritin, are expressed only in the outer fold .
the shared spatial expression of ferritin and calmodulin genes between the bivalvia and gastropoda suggests they play a conserved role in the formation or modification of the periostracum  <cit> .
the remaining  <dig> genes analyzed here are restricted to specific mantle territories and are expressed discontinuously along the length of the mantle; either in the inner fold alone , the inner mantle fold and the anterior crease of the outer fold , the anterior crease of outer fold alone , or the anterior zone of the outer fold the mantle .
the dynamic spatial expression of these genes along the length of the mantle, regardless of precise zone, implies that they are contributing to the patterning of shell structure and/or coloration  <cit> .
blue pigmentation gene
has-sometsuke  is the only gene in this study to be expressed in the anterior zone of the outer fold of the mantle  and the only gene to map precisely to a shell pigmentation pattern .
hassom transcripts are only detected in territories of the mantle that underlie regions of the shell that are producing blue dots and fields of red, which become blue upon de-calcification .
blue dots  correspond to zones of high hassom expression relative to regions of red .
the relative abundance  or absence  of this single gene product is consistent with a role in creating the juvenile shell pattern.
interestingly, blast alignments indicate the derived sometsuke protein shares weak similarity to the ependymins, a family of rapidly evolving extracellular glycoproteins previously only found in deuterostomes .
there is currently no evidence of this protein family encoded in ecdysozoan and basal metazoan genomes; we could not detect an ependymin gene within the lottia, nematostella, amphimedon or hydra genomes.
ependymin is highly expressed in vertebrate cerebrospinal fluid  <cit>  and may be involved in echinoderm tissue regeneration  <cit> .
with disparate roles in these three phyla and apparent loss from many genomes, it is difficult to infer the ancestral role of this protein in the bilateria.
however, various functional features of this protein, including its ability to bind calcium  <cit>  and to undergo polymerization into insoluble fibrils  <cit>  support a role for sometsuke in shell construction and patterning.
CONCLUSIONS
the spatial expression profiles of the genes surveyed here support the supposition that specific mantle zones influence the crystal morphology of discrete layers of the mature shell.
underlying these structural differences is a zone-specific secretome.
it appears likely that highly dynamic gene expression patterns along the length of a given mantle zone contribute to shell patterning.
in the case of juvenile h. asinina, these patterns include: the ridge and valley architecture; the periodic formation of respiratory pores, the regular deposition of blue and orange colored dots on the ridges, and the swathes of cream and red/brown fields that cover the shell.
the correspondence of hassom expression with shell coloration indicates that there are direct relationships between gene expression and shell patterns, which allows for understanding the molecular basis of structural and color patterning.
the modular design of the molluskan mantle  <cit> , along with distinct patterning mechanisms within each zone, allows for immense variation in shell structure and pattern in shell-building mollusks.
this morphogenetic system, combined with a complex and rapidly evolving secretome, as revealed here, is likely to have provided the foundation from which the incredible diversity of molluskan shell shapes and patterns has evolved.
despite the advantages of this approach to the rapid identification of novel biomineralizing proteins, it must be pointed out that common post-translational modifications  that are likely to greatly increase the diversity of the organic matrix, will not be detected by this approach, and further underscores the fact that we are some way from a detailed understanding of how nature generates these functional and beautiful structures.

 the ability to synthesize rigid, mineralized structures is an essential trait to the majority of metazoan taxa. vertebrates, echinoderms, mollusks, arthropods, brachiopods, bryozoans, annelids, cnidarians and sponges, amongst others, construct a spectacular diversity of endo- and exo-skeletons as well as sensory and protective structures from a range of minerals. the importance of this trait is highlighted by the observation that the so called'cambrian explosion' was accompanied by the diversification of biomineralization mechanisms, despite the fact that several lineages possessed this ability before the end of the proterozoic. it is currently unknown whether the molecular mechanisms used to create these structures have been inherited from an ancestral biomineralization repertoire, invented de novo, or are the result of an unprecedented lateral genetic transfer. the evolutionary origins, mode of construction, patterning and physical properties of the molluskan shell have held the attention of scientists for centuries, however the molecular mechanisms by which these structures are constructed are only now beginning to be elucidated. the mollusk shell is assembled extracellularly and is an ensemble of caco and organic macromolecules, which are secreted by the mantle epithelium. the anterior edge of the mantle tissue underlies the lip of the shell and directs the ordered biomineralization of the different structural layers of the shell and controls the patterning of architectural and color features. while the structure and function of a number of shell matrix proteins have recently been characterized, the regulatory mechanisms that govern these shell-building processes remain largely unknown. it has long been acknowledged that the diversity of shell types found in gastropod, bivalve and scaphopod mollusks are achieved through the ordered secretion of proteins and other molecules along the length of mantle, however the full complexity and role of differential gene activity in the mantle remains undescribed. the color, structure and geometric pattern of a sea shell is a historical record of the incorporation of proteins into the shell matrix and onto its surface, and directly reflects the gene expression activity of the mantle during the life of a mollusk. using the vetigastropod haliotis asinina as a model, we sought to determine the complexity of the mantle transcriptome. abalone shells are composed of three structurally-distinct layers: the inner nacreous layer, consisting of layers of aragonitic tablets encased within organic sheaths; the calcitic prismatic layer, also containing organic macromolecules; and the outer periostracum, a thin organic veneer that protects and decorates the shell. the anterior edge of the abalone mantle epithelium is convoluted and partitioned into discrete zones that produce each of these layers. within each of these zones are a number of cell types, which contribute to the construction and patterning of the shell. here we assess the complexity of gene expression in the h. asinina mantle, and explore the regulatory and structural factors that contribute to the construction of the shell. previous studies have demonstrated that the organic component of the shell is essential to its construction, and confers its remarkable physical properties. for example, lustrin-a is thought to impart fracture resistant, elastomeric properties to the nacreous layer, while macromolecules isolated from calcitic or nacreous environments can direct the type of polymorph of caco that will be deposited in vitro. unfortunately, shell matrix proteins are often insoluble, highly acidic or complexed with minerals, making their purification very difficult. to gain a broader understanding of the molecular processes that underlie seashell construction, we have analyzed expressed sequence tags from the mantle of juvenile h. asinina. this approach allows for the identification of gene products that are not necessarily incorporated into the shell, but are nonetheless crucial for caco precipitation and other biomineralization events within the pallial space adjacent to the mantle. other mantle-localized, secreted gene products not involved in biomineralization will also be detected by this methodology. we have compared this est set with the recently sequenced l. scutum genome in order to infer the degree of evolutionary conservation between shell building secretomes within one molluskan class.  structure of h. asinina shell and mantle the juvenile shell of h. asinina is an ideal model system with which to study the molecular events of shell construction and patterning. a complex but regular chromatic pattern adorns the outermost layer of the shell, the periostracum. here, a series of dots are laid down on top of the ridges of the shell. the color of the dot will be blue if it overlies a brown/red background, or orange if it overlies a cream background. this model system is such that it allows for the spatial mapping of gene expression profiles within the mantle to patterning and structural events at the leading edge of the shell. the mineralogical composition of the shell is partitioned dorso-ventrally into two major layers; a dorsal calcitic layer and a ventral aragonitic layer. the mantle epithelium, which secretes the proteins responsible for the construction of these structures, is convoluted at its anterior edge. between the two main folds lies the periostracal groove, into which the periostracum is secreted and then extruded onto the dorsal surface of the shell. we have identified a second minor fold within the mantle of h. asinina that we have termed the anterior crease of the outer fold. although we cannot yet assign a specific function to this structure, the fact that it possesses an abundance of microvilli suggests that it is actively secreting substances responsible for shell or periostracum construction. mantle expressed sequence tags we have sequenced randomly selected clones from a cdna library constructed from the juvenile mantle tissue of the tropical abalone, haliotis asinina. these sequences are available under genbank accession numbers dw to dq this tissue was taken from juveniles maintained in warm water that were rapidly depositing shell material, approximately μm per day. from this survey, we have identified unique est clusters that are expressed in the mantle. based on the presence of conserved signal sequences and similarity to secreted proteins in genbank we conservatively estimate that 26% of these encode secreted proteins. amongst the unigenes that lack both significant similarity to sequences lodged in public databases and a signal sequence, some may be secreted via mechanisms alternative to the classical signal peptide pathway. of the intracellular unigenes that encode proteins with significant similarity to genbank sequences, encode trafficking and mineral binding proteins, and are likely to represent mechanisms essential for the supply of shell building components. for example, based upon in situ hybridization analyses and previous studies on bivalves, ferritin and calmodulin are likely to be playing fundamental and evolutionarily ancient roles in shell construction within the mollusca. when compared with est surveys in other metazoan tissues, a markedly higher proportion of the genes expressed in the abalone mantle encode secreted proteins. by extrapolation, we estimate that hundreds of proteins are released extracellularly from the mantle, and contribute to the fabrication of the shell. this estimate however needs to take into consideration non-biomineralizing secreted proteins that do not possess similarity with previously described genbank sequences. this estimate is in marked contrast to the current number of biomineralizing proteins isolated from the shells of other gastropods and bivalves. with efforts chiefly focused on the characterization of proteins from the nacreous layer, fewer than protein families have been shown to contribute to shell formation to date. strikingly, of the unigenes that encode putative secreted proteins in the haliotis asinina mantle, do not share significant similarity to any sequences in genbank. however, several novel secreted open reading frames possess motifs comparable to proteins in other organisms, such as ggyglgl repeats – similar to the elasticity regions of spidroin, an elastomeric spider silk protein – and proline-rich repeats as found in plant cell walls. in comparison, of the intracellular genes do not match sequences in genbank. interestingly, blast alignments between h. asinina mantle ests and genomic trace sequences from the gastropod l. scutum, which is estimated to be sequenced to 8× coverage and is currently being assembled, reveals that only of the novel secreted proteins in haliotis appear to have identifiable homologues in the lottia genome. these genes may be involved in conserved aspects of shell construction in gastropods and possibly other mollusks. in contrast, the remaining novel secreted proteins are likely to either represent genes that evolved after these gastropod lineages split, have been lost in the lottia lineage or a combination of these two scenarios. likewise, many previously discovered molluskan shell matrix genes do not have clear homologues in the lottia genome. the few shell matrix proteins, such as lustrin, perlucin and mucoperlin, are likely to be conserved components of molluskan shells, although their general function in shell fabrication and evolutionary origins are currently unknown. together, these data suggest that the complex secretome involved in mollusk shell construction is encoded primarily by rapidly evolving genes. localized expression of mantle genes in order to infer the function of a subset of these ests, we selected for spatial expression analysis on the basis of whether they were novel, predicted to be secreted, possessed repetitive domains and/or shared significant similarity with a gene likely to play a role in biomineralization. of these, are novel and one appears to have a homologue only in the lottia genome. the spatial expression patterns of these mantle ests reveals a diversity of territories and cell types in the mantle, and highlights the modular nature of the mantle tissue. some of these are expressed homogenously in one or more zones responsible for the creation of different shell layers. for example, lustrina, first isolated from haliotis rufescens, the gene product of which contributes to the elastomeric properties of the nacreous layer, is expressed, along with two other novel genes, in the mantle territory responsible for the production of the inner nacreous layer. other evolutionarily conserved genes, including calmodulin and a calcium-binding protein, are expressed continuously along the length of the mantle within both the inner fold and the anterior crease of the outer fold, while others, such as ferritin, are expressed only in the outer fold. the shared spatial expression of ferritin and calmodulin genes between the bivalvia and gastropoda suggests they play a conserved role in the formation or modification of the periostracum. the remaining genes analyzed here are restricted to specific mantle territories and are expressed discontinuously along the length of the mantle; either in the inner fold alone, the inner mantle fold and the anterior crease of the outer fold, the anterior crease of outer fold alone, or the anterior zone of the outer fold the mantle. the dynamic spatial expression of these genes along the length of the mantle, regardless of precise zone, implies that they are contributing to the patterning of shell structure and/or coloration. blue pigmentation gene has-sometsuke is the only gene in this study to be expressed in the anterior zone of the outer fold of the mantle and the only gene to map precisely to a shell pigmentation pattern. hassom transcripts are only detected in territories of the mantle that underlie regions of the shell that are producing blue dots and fields of red, which become blue upon de-calcification. blue dots correspond to zones of high hassom expression relative to regions of red. the relative abundance or absence of this single gene product is consistent with a role in creating the juvenile shell pattern. interestingly, blast alignments indicate the derived sometsuke protein shares weak similarity to the ependymins, a family of rapidly evolving extracellular glycoproteins previously only found in deuterostomes. there is currently no evidence of this protein family encoded in ecdysozoan and basal metazoan genomes; we could not detect an ependymin gene within the lottia, nematostella, amphimedon or hydra genomes. ependymin is highly expressed in vertebrate cerebrospinal fluid and may be involved in echinoderm tissue regeneration. with disparate roles in these three phyla and apparent loss from many genomes, it is difficult to infer the ancestral role of this protein in the bilateria. however, various functional features of this protein, including its ability to bind calcium and to undergo polymerization into insoluble fibrils support a role for sometsuke in shell construction and patterning.  the spatial expression profiles of the genes surveyed here support the supposition that specific mantle zones influence the crystal morphology of discrete layers of the mature shell. underlying these structural differences is a zone-specific secretome. it appears likely that highly dynamic gene expression patterns along the length of a given mantle zone contribute to shell patterning. in the case of juvenile h. asinina, these patterns include: the ridge and valley architecture; the periodic formation of respiratory pores, the regular deposition of blue and orange colored dots on the ridges, and the swathes of cream and red/brown fields that cover the shell. the correspondence of hassom expression with shell coloration indicates that there are direct relationships between gene expression and shell patterns, which allows for understanding the molecular basis of structural and color patterning. the modular design of the molluskan mantle, along with distinct patterning mechanisms within each zone, allows for immense variation in shell structure and pattern in shell-building mollusks. this morphogenetic system, combined with a complex and rapidly evolving secretome, as revealed here, is likely to have provided the foundation from which the incredible diversity of molluskan shell shapes and patterns has evolved. despite the advantages of this approach to the rapid identification of novel biomineralizing proteins, it must be pointed out that common post-translational modifications that are likely to greatly increase the diversity of the organic matrix, will not be detected by this approach, and further underscores the fact that we are some way from a detailed understanding of how nature generates these functional and beautiful structures. 
the spatial expression profiles of mantle genes that belong to the secretome is restricted to discrete mantle zones, with each zone responsible for the fabrication of one of the structural layers of the shell. the evolutionary origins of the ability to synthesize calcified structures across various metazoan taxa remain obscure, with only a small number of protein families identified from molluskan shells. almost 85% of the secretome encodes novel proteins; remarkably, only 19% of these have identifiable homologues in the full genome of the patellogastropod lottia scutum. here we show that over 25% of the genes expressed in the mantle of the vetigastropod haliotis asinina encode secreted proteins, indicating that hundreds of proteins are likely to be contributing to shell fabrication and patterning. the unexpected complexity and evolvability of this secretome and the modular design of the molluskan mantle enables diversification of shell strength and design, and as such must contribute to the variety of adaptive architectures and colors found in mollusk shells. we also describe the expression of two novel genes involved in nacre deposition. the recent sequencing of a wide range of metazoan genomes coupled with the analysis of gene expression in non-model animals has allowed us to investigate the evolution and process of biomineralization in gastropod mollusks. for example, has-sometsuke maps precisely to pigmentation patterns in the shell, providing the first case of a gene product to be involved in molluskan shell pigmentation.
 instructions to fabricate mineralized structures with distinct nanoscale architectures, such as seashells and coral and vertebrate skeletons, are encoded in the genomes of a wide variety of animals. in mollusks, the mantle is responsible for the extracellular production of the shell, directing the ordered biomineralization of caco and the deposition of architectural and color patterns. the evolutionary origins of the ability to synthesize calcified structures across various metazoan taxa remain obscure, with only a small number of protein families identified from molluskan shells. the recent sequencing of a wide range of metazoan genomes coupled with the analysis of gene expression in non-model animals has allowed us to investigate the evolution and process of biomineralization in gastropod mollusks.  here we show that over 25% of the genes expressed in the mantle of the vetigastropod haliotis asinina encode secreted proteins, indicating that hundreds of proteins are likely to be contributing to shell fabrication and patterning. almost 85% of the secretome encodes novel proteins; remarkably, only 19% of these have identifiable homologues in the full genome of the patellogastropod lottia scutum. the spatial expression profiles of mantle genes that belong to the secretome is restricted to discrete mantle zones, with each zone responsible for the fabrication of one of the structural layers of the shell. patterned expression of a subset of genes along the length of the mantle is indicative of roles in shell ornamentation. for example, has-sometsuke maps precisely to pigmentation patterns in the shell, providing the first case of a gene product to be involved in molluskan shell pigmentation. we also describe the expression of two novel genes involved in nacre deposition.  the unexpected complexity and evolvability of this secretome and the modular design of the molluskan mantle enables diversification of shell strength and design, and as such must contribute to the variety of adaptive architectures and colors found in mollusk shells. the composition of this novel mantle-specific secretome suggests that there are significant molecular differences in the ways in which gastropods synthesize their shells. 