EAGER: Experimental Techniques for Discerning Female-specific Morphology in Non-avian Theropod Dinosaurs

鸟类包括的物种比任何其他现存的脊椎动物都多，是大约6500万年前白垩纪末期大规模灭绝后幸存下来的唯一恐龙谱系。鸟类的长期成功可能部分归因于它们独特的生殖生理，其中包括一种特殊的骨骼组织，储存在长骨的内腔中，被称为髓质骨(MB)。甲基溴作为生产蛋壳的钙储备，长期以来被认为是鸟类特有的一种关键的生殖适应。然而，这种组织最近被认为存在于少数灭绝物种中，包括非鸟类恐龙和恐龙近亲。鉴于已经初步确定MB的各个群体之间的进化距离很远(跨越在鸟类起源之前1亿多年存在分歧的谱系)，在这些分类群中证实(或驳斥)MB并仔细研究这些骨骼组织的化学特征和微观结构是确定这些鉴定的有效性和这种组织的进化意义的关键第一步。拟议的项目将具有变革性，开发一种未经测试的、独立的、用于从化学上区分化石中甲基溴的替代物，以及一种非破坏性的手段，用于识别化石记录中的甲基溴的微结构，可用于探索鸟类血统的生殖生活史和生理学。与这项研究相关的公众宣传和教育将在北卡罗来纳州自然科学博物馆进行。髓质骨(MB)是现存鸟类骨髓腔内的一种短暂的、雌激素依赖的骨内组织。这种特殊的生殖组织在组织学上是高度血管化的，显示出编织的骨基质，并且比其他类型的骨更密集地矿化。虽然最初被认为是鸟类的一种关键适应，但后来在非鸟类兽脚类和鸟足类恐龙中发现了甲基溴，在系统发育上与现存的鸟类直系相去甚远，最近在恐龙之外，在该支系最接近灭绝的近亲翼龙(非恐龙类飞行爬行动物)中发现了甲基溴。到目前为止，化石中甲基溴的鉴定完全依赖于通过组织学(破坏性)采样与鸟类中甲基溴的结构比较(位置、质地、分布和微观结构)。然而，活着的鸟类中的MB也可以通过其独特的分子特征进行识别，因为该组织比皮质骨包含更多的非胶原蛋白，并利用更多的硫酸糖胺聚糖，特别是皮质骨中不存在的角蛋白硫酸盐。该项目将提供有关现存鸟类体内甲基溴化学和微结构变化的新信息，并将利用这些数据产生新的微CT和组织化学技术，以识别化石记录中的甲基溴。然后，该方案将被用于测试已发表的关于已灭绝的始祖龙中MB的同源性，最终为髓骨的进化和鸟类特有的生殖生理提供一个知情的系统发育背景。这项研究是由美国国家科学基金会国际科学与工程办公室共同资助的，以支持该项目中对中国和蒙古的化石研究。

Birds include more species than in any other group of extant vertebrates, and were the only lineage of dinosaurs to survive the mass extinction at the end of the Cretaceous Period about 65 million years ago. The long-standing success of birds may be attributable in part to their unique reproductive physiology, which includes a specialized skeletal tissue deposited in the inner cavity of long bones known as medullary bone (MB). MB acts as a calcium reserve for the production of eggshells and was long thought to be a key reproductive adaptation unique to birds. However, the tissue has recently been purported to occur in a handful of extinct species, including both non-avian dinosaurs and dinosaurian relatives. Given the vast evolutionary distance across the groups in which MB has been tentatively identified (across lineages that diverged over 100 million years prior to the origin of birds), substantiation (or refutation) of MB in these taxa and a careful study of the chemical characteristics and microstructure of these skeletal tissues is a critical first step to determine the validity of these identifications and the evolutionary significance of this tissue. The proposed project will be transformative in developing an untested, independent proxy for chemically distinguishing MB in fossils, and a non-destructive means for identifying the microstructure of MB in the fossil record that can be used to explore reproductive life history and physiology of the bird lineage in deep time. Public outreach and education associated with this research will be conducted at the North Carolina Museum of Natural Sciences. Medullary bone (MB) is an ephemeral, estrogen dependent, endosteal tissue lining the marrow cavity in extant avialans (birds). This specialized reproductive tissue is highly vascular histologically, exhibits a woven bone matrix, and is more densely mineralized than other bone types. Although originally considered a key avian adaptation, MB has since been identified in non-avian theropod and ornithopod dinosaurs, far removed phylogenetically from the direct lineage of extant avialans, and most recently outside of Dinosauria, in one of the clade's closest extinct relatives Pterosauria (non-dinosaurian flying reptiles). To date, identifications of MB in fossils have relied exclusively on structural comparisons (location, texture, distribution, and microstructure) with MB in birds by means of histological (destructive) sampling. However, MB in living birds can also be identified by its unique molecular signature because the tissue incorporates more non-collagenous proteins than cortical bone, and utilizes more sulfated glycosaminoglycans, particularly keratan sulfate, which is not present in cortical bone. This project will provide new information on chemical and microstructural variation in MB within extant avialans and will use these data to generate novel µCT and histochemical techniques for identifying MB in the fossil record. This protocol will then be used to test the homology of published accounts of MB in extinct Archosauria, ultimately providing an informed phylogenetic context for the evolution of medullary bone and avian-specific reproductive physiology. This research is co-funded through the NSF Office of International Science and Engineering in support of research on fossils in China and Mongolia in this project.