Genetic and Molecular Regulation of Beak Tip (Rhinotheca) Shape in Layer Hens

喙是一个复杂的器官，在形状和大小上变化很大。鸡的喙由两根骨头组成，上面覆盖着一层角蛋白（构成毛发、指甲、羽毛、爪子和喙的蛋白质）。鸡嘴为蛋鸡提供了吃、喝和探索环境的手段。它还能对其他母鸡造成伤害，这是一种被称为伤害性啄的有害行为。有害啄食对母鸡的福利和生产构成严重挑战，特别是在非笼养鸡群中，其患病率可达80%。伤害性啄食的爆发可能导致同类相食和死亡，在极端情况下死亡率为50%或更高。尽管近几十年来在管理方面取得了进步，但啄鸟疫情仍然不能可靠地控制或预防。喙修剪是减少啄食相关损害的最有效方法；然而，这种做法也引发了自身的福利问题。这导致了许多国家禁止剪喙。在英国，2016年曾考虑过一项禁令，但遭到拒绝；然而，DEFRA的目标是“尽快合理地停止常规的喙修剪。为了实现这一目标，需要尽一切努力减少有害的啄食。”为了实现可持续的解决方案，需要采取多因素的办法。在改善鸟类管理方面取得了一些成功，以尽量减少未修剪过喙的鸟群中的疫情。通过基因选择不太容易发生这种行为的蛋鸡，也成功地降低了产蛋率。这项工作提出了一个不同的方向来补充这些努力。最近的研究表明，喙本身可以是一个工具，以减少啄相关的伤害，并提高成功的机会，没有喙处理蛋鸡。我们的研究小组已经开发并发表了识别喙形状的方法，并发现遗传层线内部和之间的大变化，表现为尖度，角度和整体喙的大小。除了这些形状和大小的外部测量之外，使用喙作为减少伤害的工具需要详细了解喙在胚胎中是如何发育的以及它的形状是如何确定的。目前，人们还不完全了解远端喙尖（鼻鞘）是如何发育的。也就是说，构成鼻鞘的细胞是否在细胞命运、分子身份和蛋白质组成方面是专门的？我们的总体目标是通过基因干预，包括选择和基因编辑，使喙形修改在商业规模。我们的目标是通过建立两组具有相反喙形的蛋鸡来做到这一点。在用机器学习对鸟类的喙进行成像和分类后，它们将被分成不同的组。每组的繁殖对将用于胚胎生产。来自这些群体的胚胎将进行组织学和原位杂交，以详细描述鼻鞘角化前细胞分化的变化。利用罗斯林研究所的特殊鸟类系，命运图（用于研究组织的胚胎起源）将被创造出来，以确定形成喙尖的细胞来自哪里。最后，了解鼻鞘细胞的起源并使用育种组胚胎，我们将创建一个体外角质细胞模型，以探索遗传和传递的扰动如何改变这些专门细胞的蛋白质生产。这将使测试与喙发育和组成有关的遗传变异成为可能。这一努力的成功需要了解鼻鞘形状的遗传、分子和细胞决定因素，以避免影响喙的功能和其他多效性效应。通过了解在分子水平上控制喙形的因素，我们获得的知识和工具可能导致能够预测和/或操纵基因组变化如何改变鸡喙形。在未来，新的育种技术可以利用这些知识。

The beak is a complex organ that varies considerably in shape and size. Chicken beaks are formed of two bones covered by a layer of keratin (protein that makes up hair, nails, feathers, claws, and beaks). The beak provides laying hens with the means to eat, drink, and explore their environment. It is also capable of inflicting injury on other hens, a harmful behaviour known as injurious pecking. Injurious pecking poses a serious challenge to hen welfare and production, particularly in non-cage flocks where its prevalence can reach 80%. Outbreaks of injurious pecking can result in cannibalism and death, with 50% mortality or more in extreme cases. Despite the advancements in management made in recent decades, pecking outbreaks still cannot be reliably controlled or prevented. Beak trimming is the most effective method of reducing pecking-related damage; however, the practice raises its own welfare issues. This has led to beak trimming bans in numerous countries. In the UK, a ban was considered in 2016 and declined; however, DEFRA aims "to stop routine beak trimming as soon as reasonably possible. To achieve this, every effort is needed to reduce injurious pecking." To move towards a sustainable solution, a multifactorial approach is needed. There has been some success in improving bird management to minimise outbreaks in non-beak trimmed flocks. There has also been success in reducing the incidence of IP by genetically selecting laying hens that are less apt to engage in the behaviour. This work proposes a different direction to supplement these efforts. Recent research suggests that the beak itself can be a tool to reduce pecking-related damage and improve the chances of successfully housing non-beak treated laying hens. Our research group has developed and published methods to identify beak shapes and found large variation within and between genetic layer lines, manifesting as pointedness, angle, and overall beak size. Beyond these external measures of shape and size, using the beak as a tool to reduce damage requires a detailed understanding of how the beak develops within the embryo and how its shape is determined. Presently, it is not fully understood how the distal beak tip (rhinotheca) develops. Namely, are the cells that make up the rhinotheca specialised with respect to cellular fate, molecular identity, and protein composition? Our overall goal is to enable beak shape modification at a commercial scale through genetic interventions including selection and gene editing. We aim to do this by establishing two groups of laying hens with opposing beak shapes. Birds will be placed into groups after their beaks are imaged and categorised using machine learning. Breeding pairs from each group will be used for embryo production. The embryos from these groups will undergo histology and in situ hybridisation to detail changes of cell differentiation that precede rhinotheca keratinisation. Using specialised avian lines from the Roslin Institute, fate maps (used to study embryonic origins of tissues) will be created to establish where the cells that contribute to the beak tip originate from. Finally, understanding where the rhinotheca cells originate and using the breeding group embryos, we will create an in vitro keratinocyte model to explore how perturbations, genetic and delivered, alter these specialised cells' protein production. This will enable testing of genetic variants relevant to beak development and composition. Success in this endeavour requires understanding the genetic, molecular, and cellular determinants of rhinotheca shape to avoid compromising beak function and other pleiotropic effects. By understanding factors that control beak shape at the molecular level, we gain knowledge and tools that may lead to being able to predict and/or manipulate how changes in the genome alter chicken beak shape. In the future, new breeding technologies could capitalise on this knowledge.