Dataset Information

Transcriptome and proteome analysis of Pinctada margaritifera calcifying mantle and shell: focus on biomineralization.

ABSTRACT: BACKGROUND: The shell of the pearl-producing bivalve Pinctada margaritifera is composed of an organic cell-free matrix that plays a key role in the dynamic process of biologically-controlled biomineralization. In order to increase genomic resources and identify shell matrix proteins implicated in biomineralization in P. margaritifera, high-throughput Expressed Sequence Tag (EST) pyrosequencing was undertaken on the calcifying mantle, combined with a proteomic analysis of the shell. RESULTS: We report the functional analysis of 276 738 sequences, leading to the constitution of an unprecedented catalog of 82 P. margaritifera biomineralization-related mantle protein sequences. Components of the current "chitin-silk fibroin gel-acidic macromolecule" model of biomineralization processes were found, in particular a homolog of a biomineralization protein (Pif-177) recently discovered in P. fucata. Among these sequences, we could show the localization of two other biomineralization protein transcripts, pmarg-aspein and pmarg-pearlin, in two distinct areas of the outer mantle epithelium, suggesting their implication in calcite and aragonite formation. Finally, by combining the EST approach with a proteomic mass spectrometry analysis of proteins isolated from the P. margaritifera shell organic matrix, we demonstrated the presence of 30 sequences containing almost all of the shell proteins that have been previously described from shell matrix protein analyses of the Pinctada genus. The integration of these two methods allowed the global composition of biomineralizing tissue and calcified structures to be examined in tandem for the first time. CONCLUSIONS: This EST study made on the calcifying tissue of P. margaritifera is the first description of pyrosequencing on a pearl-producing bivalve species. Our results provide direct evidence that our EST data set covers most of the diversity of the matrix protein of P. margaritifera shell, but also that the mantle transcripts encode proteins present in P. margaritifera shell, hence demonstrating their implication in shell formation. Combining transcriptomic and proteomic approaches is therefore a powerful way to identify proteins involved in biomineralization. Data generated in this study supply the most comprehensive list of biomineralization-related sequences presently available among protostomian species, and represent a major breakthrough in the field of molluskan biomineralization.

SUBMITTER: Joubert C

PROVIDER: S-EPMC3091754 | biostudies-literature | 2010

REPOSITORIES: biostudies-literature

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Transcriptome and proteome analysis of Pinctada margaritifera calcifying mantle and shell: focus on biomineralization.

Joubert Caroline C Piquemal David D Marie Benjamin B Manchon Laurent L Pierrat Fabien F Zanella-Cléon Isabelle I Cochennec-Laureau Nathalie N Gueguen Yannick Y Montagnani Caroline C

BMC genomics 20101101

<h4>Background</h4>The shell of the pearl-producing bivalve Pinctada margaritifera is composed of an organic cell-free matrix that plays a key role in the dynamic process of biologically-controlled biomineralization. In order to increase genomic resources and identify shell matrix proteins implicated in biomineralization in P. margaritifera, high-throughput Expressed Sequence Tag (EST) pyrosequencing was undertaken on the calcifying mantle, combined with a proteomic analysis of the shell.<h4>Res ...[more]

PMID: 21040589

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Project description:Molluscan larval ontogeny is a highly conserved process typical of 3 principal developmental stages. A characteristic unique to each of these stages is shell design, termed prodissoconch I, prodissoconch II and dissoconch. These shells vary in morphology, mineralogy and microstructure. The discrete temporal transitions in shell biomineralization between these larval stages are utilized in this study to investigate transcriptional involvement in several distinct biomineralization events. Scanning electron microscopy and X-ray diffraction analysis of P. maxima larvae and juveniles collected throughout post-embryonic ontogenesis, document the mineralogy and microstructure of each shelled stage as well as establishing a timeline for transitions in biomineralization. P. maxima larval samples most representative of these biomineralization distinctions and transitions were analyzed for differential gene expression on the microarray platform PmaxArray 1.0. A number of transcripts are reported as differentially expressed in correlation to the mineralization events of P. maxima larval ontogeny. Some of those isolated are known shell matrix genes while others are novel, these are discussed in relation to potential shell formation roles. This interdisciplinary investigation has married the shell developments of P. maxima larval ontogeny with corresponding gene expression profiles, furthering the elucidation of shell biomineralization. Keywords: Temporal expression profiling by array Microarray is used to examine the temporal differential expression of transcripts from several bivalve larval development stages including 24hrs post fertilization, 3 days, 17 days, 20 days, 23 days, 26 days, 30 days, 35 days, 40 days. Differential expression profiles for transcripts of all the temporal samples was determined based on comparison to a common reference of unfertilized eggs. Each temporal larval sample included in the study has at least 3 replicate hybridizations. Dye flips have been incorporated in the replicates. A total of 46 microarray hybridizations were performed in this investigation for differential expression analysis.

Project description:BACKGROUND: Biomineralization is a process encompassing all mineral containing tissues produced within an organism. One of the most dynamic examples of this process is the formation of the mollusk shell, comprising a variety of crystal phases and microstructures. The organic component incorporated within the shell is said to dictate this architecture. However general understanding of how this process is achieved remains ambiguous. The mantle is a conserved organ involved in shell formation throughout molluscs. Specifically the mantle is thought to be responsible for secreting the protein component of the shell. This study employs molecular approaches to determine the spatial expression of genes within the mantle tissue to further the elucidation of the shell biomineralization. RESULTS: A microarray platform was custom generated (PmaxArray 1.0) from the pearl oyster Pinctada maxima. PmaxArray 1.0 consists of 4992 expressed sequence tags (ESTs) originating from mantle tissue. This microarray was used to analyze the spatial expression of ESTs throughout the mantle organ. The mantle was dissected into five discrete regions and analyzed for differential gene expression with PmaxArray 1.0. Over 2000 ESTs were determined to be differentially expressed among the tissue sections, identifying five major expression regions. In situ hybridization validated and further localized the expression for a subset of these ESTs. Comparative sequence similarity analysis of these ESTs revealed a number of the transcripts were novel while others showed significant sequence similarities to previously characterized shell related genes. CONCLUSIONS: This investigation has mapped the spatial distribution for over 2000 ESTs present on PmaxArray 1.0 with reference to specific locations of the mantle. Expression profile clusters have indicated at least five unique functioning zones in the mantle. Three of these zones are likely involved in shell related activities including formation of nacre, periostracum and calcitic prismatic microstructure. A number of novel and known transcripts have been identified from these clusters. The development of PmaxArray 1.0, and the spatial map of its ESTs expression in the mantle has begun characterizing the molecular mechanisms linking the organics and inorganics of the molluscan shell.

Project description:BackgroundColor polymorphism in the nacre of pteriomorphian bivalves is of great interest for the pearl culture industry. The nacreous layer of the Polynesian black-lipped pearl oyster Pinctada margaritifera exhibits a large array of color variation among individuals including reflections of blue, green, yellow and pink in all possible gradients. Although the heritability of nacre color variation patterns has been demonstrated by experimental crossing, little is known about the genes involved in these patterns. In this study, we identify a set of genes differentially expressed among extreme color phenotypes of P. margaritifera using a suppressive and subtractive hybridization (SSH) method comparing black phenotypes with full and half albino individuals.ResultsOut of the 358 and 346 expressed sequence tags (ESTs) obtained by conducting two SSH libraries respectively, the expression patterns of 37 genes were tested with a real-time quantitative PCR (RT-qPCR) approach by pooling five individuals of each phenotype. The expression of 11 genes was subsequently estimated for each individual in order to detect inter-individual variation. Our results suggest that the color of the nacre is partially under the influence of genes involved in the biomineralization of the calcitic layer. A few genes involved in the formation of the aragonite tablets of the nacre layer and in the biosynthesis chain of melanin also showed differential expression patterns. Finally, high variability in gene expression levels were observed within the black phenotypes.ConclusionsOur results revealed that three main genetic processes were involved in color polymorphisms: the biomineralization of the nacreous and calcitic layers and the synthesis of pigments such as melanin, suggesting that color polymorphism takes place at different levels in the shell structure. The high variability of gene expression found within black phenotypes suggests that the present work should serve as a basis for future studies exploring more thoroughly the expression patterns of candidate genes within black phenotypes with different dominant iridescent colors.

Dataset Information

Transcriptome and proteome analysis of Pinctada margaritifera calcifying mantle and shell: focus on biomineralization.

Publications

Transcriptome and proteome analysis of Pinctada margaritifera calcifying mantle and shell: focus on biomineralization.

Similar Datasets

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