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Sox4 belonged to the SoxC subfamily of the Sox family, which play important roles in the development of the vertebrate gonad and nervous system. A Sox4 homologue was cloned from brain of Paramisgurnus dabryanus by using homologous cloning and rapid amplification of cDNA ends (RACE), designated as PdSox4 . The full-length cDNA was 2163bp, containing the 759bp 5'-untranslated region, 267bp 3'-untranslated region and encoding a putative protein of 378 amino acids with a characteristic high mobility group box (HMG-box) DNA-binding domain of 79 amino acids with the specific motif (RPMNAFMVW). Alignment and phylogenetic analyses indicated that PdSox4 shares highly identical sequence with Sox4 homologues from different species. The signal peptide analysis predicted that PdSox4 is a non-secretory protein. The hydropathy profile of PdSox4 protein revealed that this protein is hydrophilic in nature. The expression profiles of PdSox4 in different developmental stages and various adult tissues of sexs were analyzed by quantitative real-time RT-PCR (qRT-PCR) and In situ hybridization (ISH). The results showed that PdSox4 was ubiquitously expressed during embryogenesis and various adult tissues, especially in central nervous system. Tissue distribution analyses revealed that PdSox4 was expression in developing germ cells. Taken together, these preliminary findings suggested that PdSox4 is highly conserved during vertebrate evolution and involved in a wide range of developmental processes including embryogenesis, neurogenesis and gonad development. [ABSTRACT FROM AUTHOR]

Teleost fins can regenerate accurate position-matched structure and function after amputation. However, we still lack systematic transcriptional profiling and methodologies to understand the molecular basis of fin regeneration. After histological analysis, we established a suppression subtraction hybridization library containing 418 distinct sequences expressed differentially during the process of blastema formation and differentiation in caudal fin regeneration. Genome ontology and comparative analysis of differential distribution of our data and the reference zebrafish genome showed notable subcategories, including multi-organism processes, response to stimuli, extracellular matrix, antioxidant activity, and cell junction function. KEGG pathway analysis allowed the effective identification of relevant genes in those pathways involved in tissue morphogenesis and regeneration, including tight junction, cell adhesion molecules, mTOR and Jak-STAT signaling pathway. From relevant function subcategories and signaling pathways, 78 clones were examined for further Southern-blot hybridization. Then, 17 genes were chosen and characterized using semi-quantitative PCR. Then 4 candidate genes were identified, including F11r , Mmp9 , Agr2 and one without a match to any database. After real-time quantitative PCR, the results showed obvious expression changes in different periods of caudal fin regeneration. We can assume that the 4 candidates, likely valuable genes associated with fin regeneration, deserve additional attention. Thus, our study demonstrated how to investigate the transcript profiles with an emphasis on bioinformatics intervention and how to identify potential genes related to fin regeneration processes. The results also provide a foundation or knowledge for further research into genes and molecular mechanisms of fin regeneration. [ABSTRACT FROM AUTHOR]

Abstract: Heat shock protein 60 (Hsp60) is a kind of highly conserved immunogenic molecule involved in a wide range of biochemical processes in response to external stressors. Its multifunction in regulating immune responses and modulating signal pathway interests us in investigating its role in fin regeneration that has become an excellent and interesting model for studying the molecular basis of morphogenesis. We firstly clarified basical process and crucial period of caudal fins regeneration in Paramisgurnus dabryanus by histological analysis. Then we cloned full-length cDNA of hsp60 from P. dabryanus (designated as PdHsp60) by RACE method. The cDNA contains a 124 bp 5′UTR, a 1731 bp open reading frame (ORF) encoding 576 amino acids and a 510bp 3′UTR (Accession no.: KF544774). The phylogenetic tree shows that the PdHsp60 fits within the hsp60 clade. And quantitative RT-PCR detected the PdHsp60 began to increase rapidly its expression at 1 dpa and reached its peak at 2 dpa. Next, spatial distribution analysis of PdHsp60 in fins showed that PdHsp60 located mainly in the deeper lay of regenerated epidermis when PdHsp60 expressed most. After the PdHsp60 had been cloned into the pET-32a vector, SDS-PAGE and Western blotting analysis confirmed that the PdHsp60 protein was efficiently expressed in Escherichia coli BL21. These findings have revealed that PdHsp60, a highly conserved gene related to the innate immune system and stress response during vertebrate evolution, is involved in response to wounding stimulation—in the formation of wound epidermis which occurs as the first phase of fin regeneration after fin amputation in caudal fin regeneration. [Copyright &y& Elsevier]