Table_1_Heat Shock Protein 40 (HSP40) in Pacific White Shrimp (Litopenaeus vannamei): Molecular Cloning, Tissue Distribution and Ontogeny, Response to Temperature, Acidity/Alkalinity and Salinity Stresses, and Potential Role in Ovarian Development.DOCX

Heat shock proteins (HSPs), a family of conserved proteins that are produced by cells in response to stresses, are known as molecular chaperones with a range of housekeeping and cellular protective functions. The 40 kD heat shock protein (HSP40) is a co-chaperone for HSP70 in the regulation of ATP hydrolysis. Unlike its well-documented cofactor HSP70, little is currently known regarding the biological functions of HSP40 in crustacean species such as penaeid shrimp. In the present study, the cDNA encoding HSP40 (Lv-HSP40) was identified from the Pacific white shrimp Litopenaeus vannamei, a highly significant commercial culture species. The structural organization indicates that Lv-HSP40 belongs to the type-I HSP40s. The muscle, gill, and hepatopancreas are the main sites of Lv-HSP40 transcript expression. Within these tissues, Lv-HSP40 mRNA were predominantly exhibited in the myocytes, epithelial cells and hepatopancreatic cells, respectively. Under acute thermal stress in the culture environment, Lv-HSP40 transcript levels are significantly induced in these three tissues, while low pH stress only upregulates Lv-HSP40 mRNA in the hepatopancreas and gill. During ontogenesis, Lv-HSP40 transcript levels are high at early embryonic stages and drop sharply at late embryonic and early larval stages. The ovary is another major organ of Lv-HSP40 mRNA expression in female shrimp, and Lv-HSP40 transcripts were mainly presented in the follicle cells but only weekly detected in the oocytes. Ovarian Lv-HSP40 mRNA levels increase continuously during gonadal development. Silencing of the Lv-HSP40 gene by RNA interference may effectively delay ovarian maturation after unilateral eyestalk ablation. The roles of Lv-HSP40 in ovarian development are speculated to be independent of its cofactor HSP70, and the vitellogenesis factor vitellogenin (Vg) and vitellogenin receptor (VgR). Our study, as a whole, provides new insights into the roles of HSP40 in multiple physiological processes in L. vannamei: (1) HSP40 is a responding factor during stressful conditions; and (2) HSP40 participates in embryonic and ovarian development.

热休克蛋白（heat shock proteins, HSPs）是一类保守蛋白，由细胞在应激条件下合成，作为分子伴侣具备多种持家功能与细胞保护作用。40 kDa热休克蛋白（heat shock protein 40, HSP40）是热休克蛋白70（heat shock protein 70, HSP70）的辅助伴侣蛋白，参与调控ATP水解过程。相较于被广泛研究的辅助因子HSP70，目前学界对甲壳类动物（如对虾）中HSP40的生物学功能仍知之甚少。
本研究从商业化养殖的重要物种凡纳滨对虾（Litopenaeus vannamei）中克隆得到编码HSP40的cDNA，命名为Lv-HSP40。结构特征分析表明，Lv-HSP40属于I型HSP40家族。Lv-HSP40的转录本主要在肌肉、鳃和肝胰腺中表达；在这些组织中，其mRNA分别主要分布于肌细胞、上皮细胞和肝胰腺细胞。
在养殖环境中的急性热应激下，上述三种组织内的Lv-HSP40转录本水平均显著上调；而低pH胁迫仅能诱导肝胰腺与鳃组织中Lv-HSP40 mRNA的表达升高。
在个体发育过程中，Lv-HSP40的转录本在胚胎发育早期呈现高表达，在胚胎发育晚期及早期幼虫阶段则急剧下降。
卵巢是雌性凡纳滨对虾中Lv-HSP40 mRNA表达的另一主要组织，其转录本主要分布于滤泡细胞，仅在卵母细胞中微量表达。在性腺发育过程中，卵巢内Lv-HSP40的mRNA水平持续升高。通过RNA干扰（RNA interference）沉默Lv-HSP40基因，可有效延缓单侧眼柄切除后的卵巢成熟进程。研究推测，Lv-HSP40在卵巢发育中的作用与其辅助伴侣HSP70以及卵黄生成因子卵黄原蛋白（vitellogenin, Vg）、卵黄原蛋白受体（vitellogenin receptor, VgR）无关。
本研究整体为HSP40在凡纳滨对虾的多种生理过程中的作用提供了新的认识：（1）HSP40是应激条件下的响应因子；（2）HSP40参与胚胎发育与卵巢发育过程。