Du Jingwen, Hincke Maxwell T, Rose-Martel Megan, Hennequet-Antier Christelle, Brionne Aurelien, Cogburn Larry A, Nys Yves, Gautron Joel
Department of Cellular and Molecular Medicine, University of Ottawa, 451 Smyth Road, Ottawa, K1H 8 M5, Canada.
INRA, UR83 Recherches Avicoles, F-37380, Nouzilly, France.
BMC Genomics. 2015 Oct 15;16:792. doi: 10.1186/s12864-015-2013-3.
The avian eggshell membranes surround the egg white and provide a structural foundation for calcification of the eggshell which is essential for avian reproduction; moreover, it is also a natural biomaterial with many potential industrial and biomedical applications. Due to the insoluble and stable nature of the eggshell membrane fibres, their formation and protein constituents remain poorly characterized. The purpose of this study was to identify genes encoding eggshell membrane proteins, particularly those responsible for its structural features, by analyzing the transcriptome of the white isthmus segment of the oviduct, which is the specialized region responsible for the fabrication of the membrane fibres.
The Del-Mar 14 K chicken microarray was used to investigate up-regulated expression of transcripts in the white isthmus (WI) compared with the adjacent magnum (Ma) and uterine (Ut) segments of the hen oviduct. Analysis revealed 135 clones hybridizing to over-expressed transcripts (WI/Ma + WI/Ut), and corresponding to 107 NCBI annotated non-redundant Gallus gallus gene IDs. This combined analysis revealed that the structural proteins highly over-expressed in the white isthmus include collagen X (COL10A1), fibrillin-1 (FBN1) and cysteine rich eggshell membrane protein (CREMP). These results validate previous proteomics studies which have identified collagen X (α-1) and CREMP in soluble eggshell extracts. Genes encoding collagen-processing enzymes such as lysyl oxidase homologs 1, 2 and 3 (LOXL1, LOXL2 and LOXL3), prolyl 4 hydroxylase subunit α-2 and beta polypeptide (P4HA2 and P4HB) as well as peptidyl-prolyl cis-trans isomerase C (PPIC) were also over-expressed. Additionally, genes encoding proteins known to regulate disulfide cross-linking, including sulfhydryl oxidase (QSOX1) and thioredoxin (TXN), were identified which suggests that coordinated up-regulation of genes in the white isthmus is associated with eggshell membrane fibre formation.
The present study has identified genes associated with the processing of collagen, other structural proteins, and disulfide-mediated cross-linking during eggshell membrane formation in the white isthmus. Identification of these genes will provide new insight into eggshell membrane structure and mechanisms of formation that will assist in the development of selection strategies to improve eggshell quality and food safety of the table egg.
禽蛋壳膜包裹着蛋清，为蛋壳钙化提供结构基础，这对禽类繁殖至关重要；此外，它还是一种天然生物材料，具有许多潜在的工业和生物医学应用。由于蛋壳膜纤维不溶性和稳定性，其形成过程及蛋白质成分仍未得到充分表征。本研究旨在通过分析输卵管白色峡部段的转录组来鉴定编码蛋壳膜蛋白的基因，特别是那些负责其结构特征的基因，该区域是负责制造膜纤维的特殊区域。
使用Del-Mar 14 K鸡微阵列研究了与母鸡输卵管相邻的蛋白分泌部（Ma）和子宫部（Ut）相比，白色峡部（WI）中转录本的上调表达。分析发现135个克隆与过表达的转录本杂交（WI/Ma + WI/Ut），对应107个NCBI注释的非冗余原鸡基因ID。该综合分析表明，在白色峡部高度过表达的结构蛋白包括Ⅹ型胶原蛋白（COL10A1）、原纤蛋白-1（FBN1）和富含半胱氨酸的蛋壳膜蛋白（CREMP）。这些结果验证了先前蛋白质组学研究在可溶性蛋壳提取物中鉴定出Ⅹ型胶原蛋白（α-1）和CREMP的结果。编码胶原蛋白加工酶的基因，如赖氨酰氧化酶同源物1、2和3（LOXL1、LOXL2和LOXL3）、脯氨酰4-羟化酶亚基α-2和β多肽（P4HA2和P4HB）以及肽基脯氨酰顺反异构酶C（PPIC）也过表达。此外，还鉴定出编码已知调节二硫键交联的蛋白质的基因，包括巯基氧化酶（QSOX1）和硫氧还蛋白（TXN），这表明白色峡部基因的协同上调与蛋壳膜纤维形成有关。
本研究鉴定了与白色峡部蛋壳膜形成过程中胶原蛋白、其他结构蛋白加工以及二硫键介导的交联相关的基因。这些基因的鉴定将为蛋壳膜结构及其形成机制提供新的见解，有助于制定提高食用蛋蛋壳质量和食品安全的选择策略。
