Molecular Determinants of Calcitic Biomineralization.

鸡蛋壳（ES）的制造缺陷是一个人类健康和食品安全问题；营养丰富的食用鸡蛋是全世界数亿贫困人口的重要蛋白质、矿物质和维生素来源。从这个意义上说，蛋壳钙生物矿化的可重复性是人类社会关注的关键问题。在加拿大，据估计，每年大约有1万例沙门氏菌中毒病例是由于破裂和不完美的蛋壳被污染而进入食物链。在世界范围内，食源性沙门氏菌每年导致8030万例疾病（在美国每年有135万例）。在美国，沙门氏菌中毒每年导致26,500人住院，420人死亡。大约18%的沙门氏菌爆发是由于食用了由受污染的鸡蛋制成的食物。我的NSERC发现研究项目的总体目标是研究蛋壳作为生物矿化屏障的综合防御策略，以维持鸡蛋的无病原体状态。在2016-2022年期间[有16篇出版物和1本书章节证明]，我们对鸡、珍珠鸡和鸭的蛋壳生物矿化进行了详细的蛋白质组学和转录组学研究，除了从广泛的多组学进化角度进行分析外，还提供了对负责鸟类蛋壳形成的大自然工具包的见解。我们最近发表在《自然通讯》上的文章强调了蛋壳特异性c型凝集素基质蛋白（鸡的ovocleidin-17）在蛋壳矿化中的关键作用。我们发现了一种新的非晶态碳酸钙（ACC）介导的蛋壳矿化的囊泡途径，并评估了一种新的分子因子（EDIL3）的相关性。此外，还证实了骨桥蛋白对蛋壳纳米结构和硬度的调节作用。然而，蛋壳具有双重功能，既是生物矿化屏障，又是先天免疫系统的组成部分。解开免疫和生物矿化机制是理解生物体如何将生物矿化形成和免疫可塑性与关键物理保护的维持结合起来的重大挑战。在下一个资助周期中，我的NSERC Discovery研究的总体目标是研究绒毛膜尿囊膜（CAM）在ES减弱和微生物挑战期间的先天免疫作用，并进一步研究ACC在ES吸收（CAM介导的脱矿）中的作用。我们将使用转录组学（RNA-Seq）和蛋白质组学方法来研究CAM通过上调其保护机制来响应机械和微生物损伤的假设，以及稳定的ACC在胚胎生长过程中是一个容易动员的钙前体来源。这些研究将导致解决严重的食品安全问题，如食用鸡蛋的沙门氏菌污染。

Flaws in chicken eggshell (ES) fabrication are a human health and food safety issue; the nutritious table egg is an important source of protein, minerals and vitamins for hundreds of millions of individuals living in poverty worldwide. In this sense, the reproducible nature of calcitic biomineralization of the eggshell is of critical concern to human society. In Canada, it is estimated that about 10,000 annual cases of salmonella poisoning are due to contamination of cracked and imperfect eggshells that find their way into the food chain. Worldwide, foodborne salmonella is responsible for 80.3 million illnesses/yr (1.35 million cases/yr in the United States). In the USA, salmonella poisonings result in 26,500 hospitalizations/yr and 420 deaths/yr. Approximately 18% of salmonella outbreaks are due to consumption of food prepared from contaminated eggs.    My NSERC Discovery research program has an overall aim to investigate the integrated defense strategies that function at the eggshell as a biomineralized barrier in order to maintain the pathogen-free status of the egg.  During 2016-2022 [evidenced in 16 publications and 1 book chapter], our detailed proteomic and transcriptomic studies of eggshell biomineralization in chicken, guinea fowl and duck, in addition to analysis from a broad multi-omics evolutionary perspective, have provided insight into nature's toolkit that is responsible for avian eggshell formation. Our recent publication in Nature Communications emphasizes the critical role that eggshell-specific C-type lectin matrix proteins (ovocleidin-17 in chicken) play in eggshell mineralization. We identified a new vesicular pathway for amorphous calcium carbonate (ACC)-mediated eggshell mineralization and assessed the relevance of a new molecular actor (EDIL3). Moreover, the role of osteopontin in modulating eggshell nanostructure and hardness was confirmed.    However, the eggshell possesses dual functions as both a biomineralized barrier and as a component of the innate immune system. Disentangling immune and biomineralization mechanisms represents a significant challenge to understand how organisms integrate biomineral formation and immune plasticity with the maintenance of critical physical protection. The general goals of my NSERC Discovery research during the next grant cycle are to study the innate immune role of the chorioallantoic membrane (CAM) during ES weakening and microbial challenge, and to further investigate the role of ACC in ES resorption (CAM-mediated demineralization).  We will use transcriptomics (RNA-Seq) and proteomics approaches to investigate the hypotheses that the CAM responds to mechanical and microbial insults by up-regulating its protective mechanisms, and that stabilized ACC is a readily mobilized source of calcium precursor during embryonic growth. These studies will lead to solutions to serious food safety issues such as salmonella contamination of the table egg.