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Embryonic stem cells are the stuff of dreams. They can transform into any other type of tissue in the body and that has made them the subject of intense debate between scientists and pro-life advocates. Embryos are thought to hold cures for a plethora of diseases, including genetic disorders, cancer and immune disorders. Embryos are also used in other fields of basic research such as cell signalling, as well as applied research areas such as the aquaculture industry where animal embryos are being developed as monitors for environmental pollution. To this end, gastropod embryos have attracted attention but their growth and development is complex, taking them through several unique phases. These changes include reorganisation of the mantle cavity, visceral mass and some organs in a process known as torsion. Subsequent shell formation requires changes in body shape and the deposition of minerals and pigments within a protein matrix. A third change is head-foot differentiation. Although these growing transformations are well-documented, the genetic and proteomic changes that occur along the way are less clear. There must be many genes involved to initiate and control the various stages. The gene expression pattern will be reflected in the proteomic profile but there have been no reported proteomics studies of gastropod embryos, say Taiwanese researchers. So, Jian-Wen Qiu and Jin Sun from the Hong Kong Baptist University, Yu Zhang and Pei-Yuan Qian from the Hong Kong University of Science and Technology and Vengatesen Thiyagarajan from the University of Hong Kong, undertook a study of the channelled apple snail, Pomacea canaliculata. This snail is native to South America but has been found in other continents, where it is becoming a problem as a rice pest. The reproductive season of P. canaliculata extends up to 10 months and each female can lay up to 8000-9000 eggs in that time. For the Taiwanese study, this ensured a plentiful supply of embryos for study. They concentrated on the transition from stage II to stage III. In the former, head-foot differentiation is incomplete, the foregut and midgut are being connected and the mantle is just being formed. In stage III, most of the external and internal organs are fully formed and the embryo has a complete but non-pigmented shell and an operculum, the small lid that closes the shell opening when the snail retracts its soft parts. It took 5 and 9 days, respectively, for fertilised eggs to develop into stage II and III. The proteins were extracted from the embryos at each stage and separated by 2D gel electrophoresis. A total of 718 and 635 protein spots were observed for stage II and III, respectively and the most abundant 125 that were present in all of the replicate samples were selected for analysis by MALDI MS/MS following in-gel digestion with trypsin. Due to a dearth of genomic and proteomic data on gastropods, the proteins were identified by cross-species de novo sequencing using the NCBI non-redundant database. Proteins that were not identified this way were subjected to LC-electrospray tandem MS of the tryptic digest using the MSDB database, using MASCOT and MS-BLAST searching. A total of 65 proteins from the 125 were identified. They included 6 that appeared only in stage II, 1 only in stage III, 2 spots that were up-regulated, 6 that were down-regulated and 50 that did not show significant differences in expression level. Of the 15 differentially expressed proteins, 5 were recognised as housekeeping proteins but the remainder had other functions. Some of the unidentified proteins could be gene products that are specific to gastropods in general or P. canaliculata in particular. The 65 were placed into 11 functional groups, many being related to energy, metabolism, transcription, protein synthesis and protein modification. The perceived functions of several were discussed by the researchers. Proteins found in stage I but not stage II included proliferating cell nuclear antigen and putative septin 10 (both involved in cell cycle and DNA processing), actin-related protein 3 (biogenesis of cellular components), dihydrolipamide S-acetyltransferase (energy) and ERp 57 (protein fate). Conversely, the protein found solely in stage III was cytoplasmic intermediate filament protein, involved in cellular biogenesis. The results of this study will form the basis of future proteomics investigations covering various aspects of snail life, such as their responses to environmental stress and their interactions with parasites. The de novo sequences could also be used to gain a better understanding of the functions of novel genes using cloning and expression studies. Related links:
The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.
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Image: Endemic Species Research Unit, Taiwan
The snail eggs attached to a plant |
