Development of asymmetry in the caenogastropods Amphissa columbiana and Euspira lewisii

Abstract. The asymmetry displayed by the body plan of gastropods has been directly or indirectly attributed to an evolutionary process called torsion. Torsion is defined as a rotation of 180° between the cephalopodium (head and foot) and visceropallium (visceral organs, mantle, mantle cavity, and shell). During development, the displacement of anatomical components occurs during a process called "ontogenetic torsion." Although ontogenetic torsion is central to theories of gastropod evolution, surprisingly few studies have documented actual tissue movements during the development of asymmetry in gastropods. We investigated the development of the mantle cavity and pleurovisceral nerve connective (visceral nerve loop) in the caenogastropods Amphissa columbiana and Euspira lewisii , because displacements of both of these structures are interpreted as major consequences of torsion. Scanning electron micrographs, histological sections, and immunofluorescence images showed that the developing vis-ceropallium twists by 90° relative to the cephalopodium, the mantle cavity initially forms on the right side, and displacements of the visceral nerve loop become evident on the left side before the right side. A developmental stage in which the mantle cavity is confined to the right side has also been reported in members of the Vetigastropoda and Heterobranchia. We suggest that further comparative studies should test the hypothesis that early development throughout the Gastropoda converges on an embryonic organization in which the mantle cavity and anus are located laterally, despite clade-specific differences in developmental patterns both before and after this stage.     

Early differentiating neuron in larval abalone (Haliotis kamtschatkana) reveals the relationship between ontogenetic torsion and crossing of the pleurovisceral nerve cords

Crossing of the pleurovisceral nerve cords in gastropods has supported the view that gastropods evolved by 180 degrees rotation between the ventral and dorsal body regions. Indeed, a rotation of this type occurs as a dramatic morphogenetic movement ("ontogenetic torsion") during the development of basal gastropods. According to a long-standing hypothesis, ontogenetic torsion in basal gastropods preserves an ancient developmental aberration that generated the contorted gastropod body plan. It follows from this reasoning that crossing of the pleurovisceral nerve cords during gastropod development should be mechanically coupled to ontogenetic torsion. The predicted mechanical coupling can now be examined because of the discovery of an early differentiating neuron in Haliotis kamtschatkana (Vetigastropoda) that expresses 5-hydroxytryptamine-like immunoreactivity. The neuron appeared to delineate the trajectory of the pleurovisceral nerve cords beginning before ontogenetic torsion. Before torsion, the neuronal soma is embedded in mantle epithelium at the ventral midline and two neurites extend anteriorly toward the apical sensory organ. Contrary to expectation, the two neurites of this cell did not cross-over during ontogenetic torsion because the soma of this mantle neuron shifted in the same direction as the rotating head and foot. Full crossing of the pleurovisceral nerve cords occurred gradually during later development as the mantle cavity deepened and expanded leftward. These results are consistent with a generalization emerging from comparative studies indicating a conserved developmental stage for gastropods in which the mantle cavity is localized to one side, despite a fully "post-torsional" orientation for other body components. Developmental morphology before this stage is much more variable among different gastropod clades.     

Ontogenetic torsion in two basal gastropods occurs without shell attachments for larval retractor muscles

Results of this study on two species of vetigastropods contradict the long-standing hypothesis, originally proposed by Garstang (1929), that the larval retractor muscles power the morphogenetic movement of ontogenetic torsion in all basal gastropods. In the trochid Calliostoma ligatum and the keyhole limpet Diodora aspera, the main and accessory larval retractor muscles failed to establish attachments onto the protoconch (larval shell) when the antibiotics streptomycin sulfate and penicillin G were added to cultures soon after fertilization. Defects in protoconch mineralization were also observed. Despite these abnormalities, developing larvae of these species accomplished complete or almost complete ontogenetic torsion, a process in which the head and foot rotate by 180 degrees relative to the protoconch and visceral mass. Analysis by using phalloidin-fluorophore conjugate and transmission electron microscopy showed that myofilaments differentiated within myocytes of the larval retractor muscles and adherens-like junctions formed between muscle and mantle epithelial cells in both normal and abnormal larvae. However, in abnormal larvae, apical microvilli of mantle cells that were connected to the base of the larval retractor muscles failed to associate with an extracellular matrix that normally anchors the microvilli to the mineralized protoconch. If morphogenesis among extant, basal gastropods preserves the original developmental alteration that created gastropod torsion, as proposed by Garstang (1929), then the alteration involved something other than the larval retractor muscles. Alternatively, the developmental process of torsion has evolved subsequent to its origin in at least some basal gastropod clades so that the original alteration is no longer preserved in these clades.     

Turning snails into slugs: induced body plan changes and formation of an internal shell

The archetypal body plan of conchiferan molluscs is characterized by an external calcareous shell, though internalization of shells has evolved independently in a number of molluscan clades, including gastropod families. In gastropods, the developmental process of torsion is regarded as a hallmark that is associated with a new anatomical configuration. This configuration is present in extant prosobranch gastropod species, which predominantly bear external shells. Here, we show that short-term exposure to platinum during development uncouples at least two of the processes associated with torsion of the freshwater snail Marisa cornuarietis. That is, the anus of the treated snails is located anteriorly, but the gill and the designated mantle tissue remains in a posterior location, thus preventing the formation of an external shell. In contrast to the prosobranchian archetype, platinum treatment results in the formation of a posterior gill and a cone-shaped internal shell, which persists across the lifetime. This first finding of artificially induced snail-slug conversion was also seen in the pulmonate snail Planorbarius corneus and demonstrates that selective alteration of embryonic key processes can result in fundamental changes of an existing body plan and-if altered regulation is inherited-may give rise to a new one.     

Torsion in Patella caerulea (Mollusca, Patellogastropoda): ontogenetic process, timing, and mechanisms

Abstract. Torsion is a process in gastropod ontogenesis where the visceral body portion rotates 180° relative to the head/foot region. We investigated this process in the limpet Patella caerulea by using light microscopy of living larvae, as well as scanning electron microscopy (SEM) of larvae fixed during the torsion process. The completion of the 180° twist takes considerably less time in larvae of Patella caerulea than previously described for other basal gastropod species. At a rearing temperature of 20–22°C, individuals complete ontogenetic torsion in ?2 h. Furthermore, the whole process is monophasic, i.e., carried out at a constant speed, without any evidence of distinct ‘fast” or ‘slow” phases. Both larval shell muscles—the main and the accessory larval retractor—are already fully contractile before the onset of torsion. During the torsion process both retractors perform cramp‐like contractions at ～30 s intervals, which are followed by hydraulic movements of the foot. However, retraction into the embryonic shell occurs only after torsion is completed. The formation of the larval operculum is entirely in‐dependent from ontogenetic torsion and starts before the onset of rotation, as does the mineralization of the embryonic shell. The reported variability regarding the timing (mono‐ versus biphasic; duration) of torsion in basal gastropod species precludes any attempt to interpret these data phylogenetically. The present findings indicate that the torsion process in Patella caerulea, and probably generally in basal gastropods, is primarily caused by contraction of the larval shell muscles in combination with hydraulic activities. In contrast, the adult shell musculature, which is independently formed after torsion is completed, does not contribute to ontogenetic torsion in any way. Thus, fossil data relying on muscle scars of adult shell muscles alone appear inappropriate to prove torted or untorted conditions in early Paleozoic univalved molluses. Therefore, we argue that paleontological studies dealing with gastropod phylogeny require data other than those based on fossilized attachment sites of adult shell muscles.     

Evolution of the cephalopod head complex by assembly of multiple molluscan body parts: Evidence from Nautilus embryonic development

Cephalopod head parts are among the most complex occurring in all invertebrates. Hypotheses for the evolutionary process require a drastic body-plan transition in relation to the life-style changes from benthos to active nekton. Determining these transitions, however, has been elusive because of scarcity of fossil records of soft tissues and lack of some of the early developmental stages of the basal species. Here we report the first embryological evidence in the nautiloid cephalopod Nautilus pompilius for the morphological development of the head complex by a unique assembly of multiple archetypical molluscan body parts. Using a specialized aquarium system, we successfully obtained a series of developmental stages that enabled us to test previous controversial scenarios. Our results demonstrate that the embryonic organs exhibit body plans that are primarily bilateral and antero-posteriorly elongated at stereotyped positions. The distinct cephalic compartment, foot, brain cords, mantle, and shell resemble the body plans of monoplacophorans and basal gastropods. The numerous digital tentacles of Nautilus develop from simple serial and spatially-patterned bud-like anlagen along the anterior-posterior axis, indicating that origins of digital tentacles or arms of all other cephalopods develop not from the head but from the foot. In middle and late embryos, the primary body plans largely change to those of juveniles or adults, and finally form a "head" complex assembled by anlagen of the foot, cephalic hood, collar, hyponome (funnel), and the foot-derived epidermal covers. We suggest that extensions of the collar-funnel compartment and free epidermal folds derived from multiple topological foot regions may play an important role in forming the head complex, which is thought to be an important feature during the body plan transition.     

Further twists in gastropod shell evolution

The manner in which a gastropod shell coils has long intrigued laypersons and scientists alike. In evolutionary biology, gastropod shells are among the best-studied palaeontological and neontological objects. A gastropod shell generally exhibits logarithmic spiral growth, right-handedness and coils tightly around a single axis. Atypical shell-coiling patterns (e.g. sinistroid growth, uncoiled whorls and multiple coiling axes), however, continue to be uncovered in nature. Here, we report another coiling strategy that is not only puzzling from an evolutionary perspective, but also hitherto unknown among shelled gastropods. The terrestrial gastropod Opisthostoma vermiculum sp. nov. generates a shell with: (i) four discernable coiling axes, (ii) body whorls that thrice detach and twice reattach to preceding whorls without any reference support, and (iii) detached whorls that coil around three secondary axes in addition to their primary teleoconch axis. As the coiling strategies of individuals were found to be generally consistent throughout, this species appears to possess an unorthodox but rigorously defined set of developmental instructions. Although the evolutionary origins of O. vermiculum and its shell's functional significance can be elucidated only once fossil intermediates and live individuals are found, its bewildering morphology suggests that we still lack an understanding of relationships between form and function in certain taxonomic groups.     

Siphonariid development: Quintessential euthyneuran larva with a mantle fold innovation (Gastropoda; Panpulmonata)

Recent phylogenetic revisions of euthyneuran gastropods (“opisthobranchs” and “pulmonates”) suggest that clades with a planktotrophic larva, the ancestral life history for euthyneurans, are more widely distributed along the trunk of the euthyneuran tree than previously realized. There is some indication that the planktotrophic larva of euthyneurans has distinctive features, but information to date has come mainly from traditional “opisthobranch” groups. Much less is known about planktotrophic “pulmonate” larvae. If planktotrophic larvae of “pulmonates” share unique traits with those of “opisthobranchs,” then a distinctive euthyneuran larval-type has been the developmental starting template for a spectacular amount of evolved morphological and ecological disparity among adult euthyneurans. We studied development of a siphonariid by preparing sections of larval and postmetamorphic stages for histological and ultrastructural analysis, together with 3D reconstructions and data from immunolabeling of the larval apical sensory organ. We also sought a developmental explanation for the unusual arrangement of shell-attached, dorso-ventral muscles relative to the mantle cavity of adult siphonariids. Adult siphonariids (“false limpets”) have a patelliform shell but their C-shaped shell muscle partially embraces a central mantle cavity, which is different from the arrangement of these components in patellogastropods (“true limpets”). It is not obvious how shell muscles extending into the foot become placed anterior to the mantle cavity during siphonariid development from a veliger larva. We found that planktotrophic larvae of Siphonaria denticulata are extremely similar to previously described, planktotrophic “opisthobranch” larvae. To emphasize this point, we update a list of distinctive characteristics of planktotrophic euthyneuran larvae, which can anchor future studies on the impressive evolvability of this larval-type. We also describe how premetamorphic and postmetamorphic morphogenesis of larval mantle fold tissue creates the unusual arrangement of shell-muscles and mantle cavity in siphonariids. This result adds to the known postmetamorphic evolutionary innovations involving mantle fold tissue among euthyneurans.     

The functional significance of aperture form in gastropods

Aperture form of marine prosobranch gastropods has evolved under the influence of a number of different selective forces, including: generation of shell form; protection from predation; accommodation of the foot during clamping behavior: and accommodation of water currents in and out of the mantle cavity. Aperture form correlates positively with foot shape in most gastropods and foot shape, in turn, correlates moderately well with substrate preference. Almost all gastropods that have non-round apertures elongate the aperture parallel to the foot so that water currenth tend to flow anteriorly to posteriorly. Fresh-water pulmonates have responded to somewhat different stresses. They exhibit clamping behavior and thus show correspondence between foot shape and aperture shape. They show less apertural strengthening as crab (or crayfish) predation is less of a factor and presumably because calcium carbonate is less available. They also lack anterior-posterior apertural elongation due to the absence of water currents through their mantle cavity. Due to the absence of mantle cavity water currents and clamping behavior, terrestrial gastropods do not show the apertural modifications associated with these two factors. In addition. few adaptations of apertural form are present to resist predation. Instead, many of the apertural modifications of terrestrial pulmonates seem to be concer-ned with the problems of water loss during estivation.     

Major muscle systems in the larval caenogastropod,Ilyanassa obsoleta,display different patterns of development

This study describes the anatomical and developmental aspects of muscular development from the early embryo to competent larval stage in the gastropod Ilyanassa obsoleta. Staining of F‐actin revealed differential spatial and temporal patterns of several muscles. In particular, two major muscles, the larval retractor and pedal retractor muscles originate independently and display distinct developmental patterns similar to observations in other gastropod species. Additionally, together with the larval retractor muscle, the accessory larval muscle developed in the embryo at the trochophore stage. Therefore, both these muscles develop prior to ontogenetic torsion. The pedal retractor muscle marked the most abundant growth in the mid veliger stage. Also during the middle stage, the metapodial retractor muscle and opercular retractor muscle grew concurrently with development of the foot. We show evidence that juvenile muscles, such as the buccal mass muscle and siphon muscle develop initially during the late veliger stage. Collectively, these findings substantiate that larval myogenesis involves a complex sequence of events that appear evolutionary conserved within the gastropods, and set the stage for future studies using this model species to address issues concerning the evolution and eventual fates of larval musculature in molluscs. J. Morphol., 2009. © 2009 Wiley‐Liss, Inc.     

GARSTANG'S HYPOTHESIS AND GASTROPOD TORSION

Recent experiments purporting to show that torsion in gastropodveligers does not function defensively against attacks by planktonicpredators are inconclusive. There is still much to be said forGarstang's suggestion that torsion is advantageous to the veligerin allowing it to withdraw head first in to the mantle cavity,sealing the shell aperture with the foot and operculum, andthat the ability to do so may indeed have originated with asingle gene mutation in evolution. (Received 25 April 1986;     

Developmental modularity and phenotypic novelty within a biphasic life cycle: morphogenesis of a cone snail venom gland

The venom gland of predatory cone snails (Conus spp.), which secretes neurotoxic peptides that rapidly immobilize prey, is a proposed key innovation for facilitating the extraordinary feeding behaviour of these gastropod molluscs. Nevertheless, the unusual morphology of this gland has generated controversy about its evolutionary origin and possible homologues in other gastropods. I cultured feeding larvae of Conus lividus and cut serial histological sections through the developing foregut during larval and metamorphic stages to examine the development of the venom gland. Results support the hypothesis of homology between the venom gland and the mid-oesophageal gland of other gastropods. They also suggest that the mid-region of the gastropod foregut, like the anterior region, is divisible into dorsal and ventral developmental modules that have different morphological, functional and ontogenetic fates. In larvae of C. lividus, the ventral module of the middle foregut transformed into the anatomically novel venom gland of the post-metamorphic stage by rapidly pinching-off from the main dorsal channel of the mid-oesophagus, an epithelial remodelling process that may be similar to other cases where epithelial tubes and vesicles arise from a pre-existing epithelial sheet. The developmental remodelling mechanism could have facilitated an abrupt evolutionary transition to the derived morphology of this important gastropod feeding innovation.     

Larval muscle contraction fails to produce torsion in a trochoidean gastropod.

The causes and effects of ontogenetic torsion in gastropods have been debated intensely for more than a century (1-19). Occurring rapidly and very early in development, torsion figures prominently in shaping both the larval and adult body plans. We show that mechanical explanations of the ontogenetic event that invoke contraction of larval retractor muscles are inadequate to explain the observed consequences in some gastropods. The classic mechanical explanation of Crofts (4, 5) and subsequent refinements of her explanation have been based on species with rigid larval shell properties (18, 19) that cannot be extrapolated to all gastropods. We present visual evidence of the lack of rigidity of the uncalcified larval shell in a basal trochid gastropod, Margarites pupillus (Gould), and provide photographic confirmation of our prediction that larval retractor muscle contraction is insufficient to produce more than local deformation or dimpling at the site of muscle insertion. These findings do not refute muscular contraction as a primary cause of ontogenetic torsion in gastropods that calcify their larval shells prior to the onset of torsion, nor do they refute the monophyly of torsion. They do, however, suggest that torsion may be a loosely constrained developmental process with multiple pathways to the more constrained end result (20, 21).     

Pelagiella exigua,an early Cambrian stem gastropod with chaetae: lophotrochozoan heritage and conchiferan novelty

Exceptionally well-preserved impressions of two bundles of bristles protrude from the apertures of small, spiral shells of Pelagiella exigua, recovered from the Kinzers Formation (Cambrian, Stage 4, ‘Olenellus Zone’, c. 512 Ma) of Pennsylvania. These impressions are inferred to represent clusters of chitinous chaetae, comparable to those borne by annelid parapodia and some larval brachiopods. They provide an affirmative test in the early metazoan fossil record of the inference, from phylogenetic analyses of living taxa, that chitinous chaetae are a shared early attribute of the Lophotrochozoa. Shells of Pelagiella exhibit logarithmic spiral growth, microstructural fabrics, distinctive external sculptures and muscle scars characteristic of molluscs. Hence, Pelagiella has been regarded as a stem mollusc, a helcionelloid expressing partial torsion, an untorted paragastropod, or a fully torted basal member of the gastropod crown group. The inference that its chaeta-bearing appendages were anterior–lateral, based on their probable functions, prompts a new reconstruction of the anatomy of Pelagiella, with a mainly anterior mantle cavity. Under this hypothesis, two lateral–dorsal grooves, uniquely preserved in Pelagiella atlantoides, are interpreted as sites of attachment for a long left ctenidium and a short one, anteriorly on the right. The orientation of Pelagiella and the asymmetry of its gills, comparable to features of several living vetigastropods, nominate it as the earliest fossil mollusc known to exhibit evidence of the developmental torsion characteristic of gastropods. This key adaptation facilitated an evolutionary radiation, slow at first and rapid during the Ordovician, that gave rise to the remarkable diversification of the Gastropoda.     

Towards a phylogeny of gastropod molluscs: an analysis using morphological characters

Morphological (including ultrastructural) and developmental characters utilized in recent literature are critically reviewed as the basis to reassess the phylogenetic relationships of gastropods. The purpose of this paper is to provide a framework of characters for future studies and a testable phylogenetic hypothesis. This is one of the first attempts to use such characters to assess the relationships of all major clades using parsimony methods. The analysis uses 117 characters and includes 40 taxa, predominantly ‘prosobranchs’. Five outgroup taxa are included, representing four conchiferan groups and Poly-placophora. Of the 117 characters reviewed and included in the analyses, nine are shell characters (four of these are shell structure), two opercular, two muscular, four ctenidial, 12 renopericardial and 24 reproductive (including 17 based on sperm and spermatogenesis), 27 of the digestive system, 32 of the nervous system and sense organs; the remainder are developmental (3) and of the foot and hypobranchial gland. In the initial analysis the data set included a mixture of binary and multistate characters with all characters unordered. These data were also analysed after scaling so that each character had equal weight. A third data set was constructed in which all characters were coded as binary characters. These analyses resulted in some implausible character transformations, mainly-involving the regaining of lost pallial structures. Additional analyses were run on all three sets of data after removing five characters showing the most unlikely transformations. These analyses resulted in generally similar topologies. The robustness of the clades was tested using clade decay. The adaptive radiation of gastropods and their life history traits are briefly described and discussed and the terminology for simultaneous hermaphroditism refined. A scenario for the evolution of torsion equated with the fossil record is proposed and the effects of torsion and coiling on gastropods are discussed along with asymmetry imposed by limpet-shaped body forms. It is suggested that the first gastropods were ultradextral. The idea that heterochrony has played a major part in gastropod evolution is developed and discussed, particularly the paedomorphic stamp imposed on the apogastropods. The veliger larvae of caenogastropods and heterobranchs are contrasted and found to differ in many respects. The evolution of planktotrophy within gastropods is discussed. Recent phylogenetic hypotheses for gastropods based on molecular data are generally in broad agreement with the present results. On the basis of our analyses we discuss the major monophyletic groups within gastropods. Gastropods appear to be a monophyletic clade, and divide into two primary groups, the Eogastropoda (incorporating the patellogastropods and their (probably sinistrally coiled) ancestors and the Orthogastropoda – the remainder of the gastropods. Orthogastropoda comprises several well defined clades. The vetigastropod clade encompasses most of the groups previously included in the paraphyletic Archaeogastropoda (fissurellids, trochoideans, scissurelloideans, halioroideans pleurotomarioideans) as well as lepeto-driloidean and lepetelloidean limpets and seguenzids. The location of the hot vent taxa Peltospiridae and Neomphalidae varies with each analysis, probably because there is a lack of ultrastructural data for these taxa and parallelism in many characters. They either form a paraphyletic or monophyletic group at or near the base of the vetigastropods or a clade with the neritopsines and cocculinoideans. The neritopsines (Neritoidea etc.) consistently form a clade with the cocculinoidean limpets, but their position on the tree also differs depending on the data set used and (in the case of the scaled data) whether or not the full suite of characters is used. They are either the sister to the rest of the orthogastropods or to the apogastropods. Caenogastropods [Mesogastropoda (+ architaenioglossan groups) + Neogastropoda] are consistently monophyletic as are the heterobranchs (‘Heterostropha’+ Opisthobranchia + Pulmo-nata). The caenogastropods and heterobranchs also form a clade in all the analyses and the name Apogastropoda is redefined to encompass this group. New taxa are proposed, Sorbeoconcha for the caenogastropods exclusive of the architaenioglossan taxa, and Hypsogastropoda for the ‘higher caenogastropods“– the Sorbeoconcha exclusive of the Cerithioidea and Campaniloidea.     

Autecology of Labrocuspis, a Middle Devonian omphalotrochid gastropod

Labrocuspis kobayashii , a Middle Devonian euomphalacean omphalotrochid gastropod from Japan has a wide callus pad on its base similar to that seen in the living trochid Umbonium. L. kobayashii could support the shell over the cephalopedal mass in a similar fashion to that seen in the Umbonium species. This reconstruction indicates that the labral projection in the outer lip was above the head, and the opisthocyrt basal lip and the sinus in the outer lip were loci of inhalant and exhalant currents, respectively. Therefore, the animal is assumed to have had only one ctenidium (the left). The presence of a wide callus pad is indicative of an infaunal life habit for the Umbonium species, Labrocuspis and some Paleozoic gastropods; clamping of the shell against the foot is also suggested for some omphalotrochid gastropods. These features indicate an additional adaptive strategy employed by euomphalaceans.     

Arresting mantle formation and redirecting embryonic shell gland tissue by platinum2+ leads to body plan modifications in Marisa cornuarietis (Gastropoda,Ampullariidae)

To evaluate the threat that anthropogenic substances pose to animals when they are emitted into the environment, tests like the invertebrate embryo toxicity test with the ramshorn snail Marisa cornuarietis have been developed. These tests are used to investigate substances like the heavy metal platinum (Pt) that is used in catalytic converters and is gradually released in car exhausts. In 2010, our group reported that high Pt concentrations cause body plan alterations in snails and prevent the formation of an external shell during M. cornuarietis embryogenesis. Now, this study presents scanning‐electron micrographs and histological sections of platinum²⁺ (Pt²⁺)‐treated and untreated M. cornuarietis embryos and compares “normally” developing and “shell‐less” embryos during embryogenesis, to reveal the exact course of events that lead to this body plan shift. Both groups showed similar development until the onset of torsion 70‐ to 82‐h postfertilization. In the Pt²⁺‐exposed embryos, the rudimentary shell gland (=anlage of both shell gland and mantle, which usually evaginates, grows, and eventually covers the visceral sac) does not spread across the visceral sac but remains on its ventral side. Without the excessive growth of the shell gland, a horizontal rotation of the visceral sac relative to head and foot does not occur, as being normal during the process of torsion. J. Morphol., 2012. © 2012 Wiley Periodicals, Inc.     

Developmental role of dpp in the gastropod shell plate and co-option of the dpp signaling pathway in the evolution of the operculum

The operculum is a novel structure in gastropod molluscs. Because the operculum shows notable similarities to the shell plate, we asked whether there were an evolutionary link between these two secretory organs. We found that some of the genes involved in shell-field development are expressed in the operculum, such as dpp and grainyhead, whereas engrailed and Hox1 are not. Specific knockdown of dpp by injection of double-stranded RNA (dsRNA) resulted in malformation of the shell plate. The shell plate was smaller due to failure of activation of cell proliferation in the shell-field margin. The expressions of grainyhead and chitin synthase 1 in the shell field margin were suppressed by dpp-dsRNA. However, matrix secretion was not completely abolished, and the expressions of ferritin, engrailed or Hox1 were not affected by dpp-dsRNA, indicating that dpp is partly involved in the developmental pathway for shell matrix secretion. We also present evidence that dpp performs a key role in operculum development. Indeed, dpp-dsRNA impaired matrix secretion in the operculum as well as expression of grainyhead. Based on these observations that dpp is important for development of both the shell plate and operculum, we conclude that co-option of dpp to the posterior part of the foot contributed to the innovation of the operculum in gastropods.